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Pathology 17/10/2022

General Pathology 

Chagas disease pathology: history and challenges

Zilton A Andrade

Experimental Pathology Laboratory, Gonçalo Moniz Research Center/Fiocruz

Email: zilton@cpqgm.fiocruz.br

Soon after Chagas in 1909 made the Trypanosoma cruzi discovery and the disease it caused, Gaspar Vianna, in 1911, made its pathology study, as a crowning achievement for Chagas’ great scientific achievement. The studies that followed with Magarinos Torres, Crowell, Jorg and Mazza meant that the fundamental data on the Chagas disease pathology were already consolidated by the middle of the 20th century. Since then, the contributions have not ceased, have highlighted the disease pathogenesis great complexity, and have served as a stimulus for our experimental science development.

The historical development of knowledge about Chagas disease presents a curious aspect. The interest in each of the forms in which the disease presents itself has arisen separately in time, which is due to diverse causes. It seems that at a given time, interest is concentrated on one disease phase or clinical form. This account uses this fact to describe briefly the progress in the Chagas disease pathology study, and attempts to show the reasons for this concentration of interest for different clinical forms at different times.

Acute form of Chagas disease

We can consider a first period, from 1909 to the end of the 1930s, when interest in the disease study was centered on its acute form. Although Carlos Chagas early on began investigating disease various clinical forms, including a nervous form, the publications that appeared gave the impression that the disease was concentrated in rural areas. The eye sign description, or Romaña’s sign (1935), as well as the cutaneous chagomas and lipo-chagomas, made it easier to recognize the case and served to illustrate the various publications from the 1930s onward. The diffuse myocarditis study, with its causative agent within the contractile fibers, was already accompanied by pathogenetic interpretations. When interest in the new disease study had inexplicably waned among Brazilians, it was the acute cases repeated publicationn by Mazza’s group in Argentina that brought new encouragement to the studies resumption.

The chronic cardiac form

Beginning in the mid-1930s, and extending through the next 30 years, interest focused on the disease chronic cardiac form. The great motivating factor for this trend was the electrocardiograph entry into increasingly common use in hospitals and physicians’ offices, coupled with the progressive detection of a chronic heart disease cases frightening number, even among the urban population. From the studies of Francisco Laranja and collaborators, cardiologists in the American countries large cities, where the disease is endemic, were surprised to find a chronic myocardial pathology, progressive and severe, that exhibited a varied hue of arrhythmias, isolated or conjugated, such as had not yet been seen in any other heart disease. The cardiac pathology was thoroughly investigated in necropsy material (Figure 1) and the findings observed were correlated with the clinical and electrocardiographic manifestations, with thromboembolic phenomena appearing prominently. The rarity with which the parasites were found in the sections, in contrast to the acute form, raised discussions for the histopathological diagnosis and pathogenesis interpretation.

Figure 1  Heart Detail of a cardiac patient with Chagas’ disease.

The Digestive form

Around the 1960s there was a surge of interest for the disease digestive form. Obtaining purified and well standardized antigens and introducing new techniques have improved serology to such an extent that many have more confidence in the results. From there it was a step to test a long-held theory held by doctors in Central Brazil, who worked in an area where Chagas disease, megaesophagus and megacolon were endemic: that digestive megas were a Chagas disease manifestation. They then added the clinical and epidemiological evidence to the well-reliable serological studies, and saw that mega-oesophagus and megacolon carriers did indeed systematically exhibit positive serology for T. cruzi. This was followed by studies on pathogenesis, headed by Fritz Köberle, from Ribeirão Preto, centered on the new pathology possibility that affected, diffusely and primarily, the autonomic nervous system. Although this theory fundamental postulates (which indicated Chagas disease as a disease of the autonomic nervous system, with chronic heart disease being a neurogenic heart disease) have not been confirmed, the anatomical studies have contributed decisively to the recognition of the Chagas disease digestive form, precisely because of the broad debates that they have generated.

The indeterminate chronic form

Since 1983, when the official program to combat Triatoma infestans with residual insecticides was installed, we have witnessed a drastic reduction in transmission, which is reflected in the infection acute forms near disappearance, a low serological reactivity of schoolchildren in endemic areas, and a decrease in morbidity and mortality in chronic forms, as described by Dias and collaborators. The indeterminate form has now become the most common form of all. Although it is a relatively benign form, it is surprising how little we still know about the significance and pathogenesis of this Chagas disease form. That is why its study in these times of transmission control is a priority for those interested in the disease problems.

In fact, interest in the Chagas disease indeterminate form arose from the early days of studies, when infected individuals were found, but without the disease. The initial idea was that they were “potential cardiacs” who would suddenly die. This notion revealed the lack of a secure criterion for separating the indeterminate form from asymptomatic chagasic patients. This criterion was established by the Brazilian Society of Tropical Medicine in its 1985 meeting, which represented a breakthrough for longitudinal, clinical, and pathological studies. Anatomical studies have revealed that asymptomatic Chagas sufferers, who came to die suddenly, actually belonged to the disease chronic cardiac form. True indeterminate form carriers usually show only mild focal myocarditis on microscopic examination. Experimental studies have revealed that such foci of myocarditis have a cyclical evolution, arising after parasitic stimulation, and resolving by apoptosis of the inflammatory cells and the excess extracellular matrix re-absorption. This evolution can occur for a prolonged time, without major repercussions on the patient, as long as there is no, so far unpredictable, change in the immunopathological pattern, as reviewed by Andrade et al. in 1997. The significance elucidation of the indeterminate form and its transition mechanism to the chronic cardiac form constitute one of the great challenges for the Chagas disease pathology study today.

Chagas disease acute phase

Tania Cremonini de Araujo-Jorge

Laboratory of Innovations in Therapies, Teaching and Bioproducts, Oswaldo Cruz Institute/Fiocruz

Email: taniaaj@ioc.fiocruz.br

Chagas disease chronic phase

Joseli Lannes-Vieira

Laboratory of Interactions Biology, Oswaldo Cruz Institute/Fiocruz

Email: lannes@ioc.fiocruz.br

Chagas disease remains a major health problem in Latin American countries, where the WHO estimates that there are approximately 16-18 million infected patients, 25-35% of whom will develop cardiovascular changes in the infection chronic phase. Carlos Chagas, Gaspar Vianna, Dias, and Laranja, in their works on American trypanosomiasis, pointed out the importance of the cardiac alterations derived from the Trypanosoma cruzi pathogenic action, in the acute phase and, above all, in the infection chronic phase, as reviewed in the text by Rassi and Kropf.

Studies over the past 80 years have confirmed that the chronic Chagas’ infection pathology is primarily characterized by the cardiac form, with dilated cardiomyopathy associated with myocarditis, fibrosis, and cardiac dysfunction. The non-invasive functional evaluation methods use for chagasic heart disease is fundamental for the study of patients with suspected functional impairment or who are engaged in work that requires too much or continuous physical effort.

About 10% of individuals chronically infected with T. cruzi develop the digestive form that can result in mega colon and/or mega esophagus, which are often associated with the cardiac form, constituting the mixed chronic or cardio-digestive form. The Chagas disease nervous form existence is still controversial, but in patients with Chagas’ infection and immunosuppressed by co-infection or treatment with immune suppressing drugs, reactivation is often expressed as a severe meningoencephalitis.

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Cardiac form 

Anatomopathological aspects in the different clinical forms of human infection

Maria de Lourdes Higuchi

Heart Institute (InCor)

Email: anplourdes@incor.usp.br

Acute cardiac form

At the entry site (inoculation chagoma) there is local swelling that histologically shows the presence of an intense inflammatory reaction consisting of macrophages, fibroblasts, and lymphocytes, as well as vascular proliferation, congestion, and edema mainly in the subcutaneous area, representing panniculitis nodules and granulomatous reactions with the parasites presence. Foci of inflammation without evident parasites can also be seen, which are called metastatic chagomas. Vianna’s pioneering studies show that while the parasitized cell was intact there was no inflammatory reaction around it. The parasites move into the blood and parasitize the different organs where amastigotes can be found in histological sections in almost all cells: macrophages, endothelial cells, smooth muscle, cardiac and skeletal fibers, fibroblasts, etc. Patients may die in the acute phase due to direct complications of the parasite infection, myocarditis, meningoencephalitis, or secondary complications such as bronchopneumonia. The most common complication is myocarditis whose lesion is characterized by intense lymphohistiocytic inflammation, edema dissociating the fibers that also show an aspect of myocytolysis and edema, and the parasites are easily found. The heart is flaccid, congested and moderately enlarged in volume, with the acute myocarditis appearance.

Similar changes are found in skeletal muscles. In the intestine and esophagus there is parasitism of muscle and Auerbach’s plexus, and lymphomononuclear infiltrate along the intramural nerve plexuses with neurons destruction. In the brain, inflammation lesions with congestion, perivascular hemorrhages, and perivascular lymphocytic infiltration have also been described, with parasitism in glia cells and macrophages of the meninges, and more rarely in neurons.

Latent or indeterminate cardiac form

The patients vast majority who are infected with Trypanosoma cruzi do not die during the acute phase, which can be oligo- or asymptomatic. About 10 years or more after the acute phase, some of the individuals start to have cardiac or digestive symptoms, constituting the symptomatic chronic phase. However, most patients do not develop signs or symptoms attributable to the Chagas disease chronic phase, individuals with the indeterminate form. Myocardial biopsy studies performed in the 1980s showed that in 15% of individuals with the disease indeterminate form show mild inflammation in the myocardium, small foci of myocardial aggression, while symptomatic patients show more frequent and more intense myocarditis in activity.

Chronic cardiac form

It is the most important Chagas disease form because it is responsible for cardiac involvement that leads to alterations in heart rhythm, thromboembolic phenomena, and congestive heart failure. Histologically there is often an intense chronic myocarditis by mononuclear cells, fibrosing, with great myocardiocytes hypertrophy, and the finding of T. cruzi nests is infrequent. The heart characteristics vary according to the death cause. In individuals who died of sudden death usually associated with arrhythmias and who did not have congestive heart failure, the heart may be little or moderately enlarged in volume, while those who die with CHF have hearts that are greatly enlarged in volume, dilated, and with intense hypertrophy. In these, intracavitary thrombosis often occurs, most frequently in the right atrium and left ventricle tip. The lesion considered Chagas disease pathognomonic is called the tip lesion, represented by tip thinning where there is total or myocardium partial disappearance that is replaced by fibrosis, and may be associated with mural thrombosis (Figure 1).

Figure 1 – Heart sagittal section of a chagasic patient who died of congestive heart failure, showing ventricular cavities dilatation, left ventricular tip thinning and of the right ventricle, with thrombosis. 

Comparative studies of the patients hearts who died suddenly or with CHF have shown qualitatively similar lesions, but more intense in the second group. An interesting finding was the granulomatous lesions presence in 50% of the cases in the first group and in only 11% of the second hearts group. The heart of patients who die with CHF is round, globose, with congested venous vessels, dilated heart chambers, especially on the right side. The whitish fibrotic nodules presence along the coronary arteries is frequent, which was called coronary rosarium and was initially considered Chagas disease pathognomonic. It was later found that such a lesion can occur in other diseases that course with cardiomegaly. In addition to tip thinning that can lead to aneurysmal dilatation, other myocardial thinning areas can occur, one of the most frequent being the basal portion of the left ventricle posterolateral wall (Figure 2).

Figure 2 – Myocardial thinning of the L ventricle posterolateral and basal regions, with replacement by fibrosis. Frequent cardiac arrhythmia focus of the sustained ventricular tachycardia type.

Studies on the pathogenesis of these localized fibrotic lesions, with myocardial wall thinning, lead in the ischemic pathogenetic mechanism direction, which will be detailed further on. In the Chagas disease chronic phase, the intensity of the inflammatory infiltrate is unrelated to the parasites presence and quantity. The infiltrate tends to be predominantly lymphocytic and multifocal, distributed throughout the myocardium, and is accompanied by intense fibrosis that sometimes surrounds isolated cardiac fibers. The mast cells presence may also be important, and lymphocytes attacking non-parasitized cardiac fibers suggests the existence of factors other than the parasite inducing the inflammatory lesion perpetuation.

Chronic Chagas’ disease and the parasite in the myocardium

In view of the parasite absence or scarcity in the myocardium, some theories have been raised to explain the chronic Chagas’ cardiopathy development. Köberle, in the 1960s, proposed that, like the digestive changes, the cardiac changes could be due to parasympathetic neuronal denervation caused by T. cruzi leading to an exacerbation of the sympathetic nervous system action. In this theory, myocardial inflammation would not play a major role in the cardiac decompensation development.

Humoral and cellular autoimmunity are theories that predominated in the 1970s where the humoral factor presence described as EVI factor in the Chagas disease carriers serum that would react against endocardium, vessels, and interstitium of myocardium and skeletal muscle of chronic Chagas disease carriers. However, no immunoglobulins were detected in cardiac biopsies from Chagas disease patients with heart disease, suggesting that the EVI factor present in the serum may not have a direct pathogenetic role in self-injury. Antibodies against myocardial muscarinic and adrenergic receptors present in the Chagas disease carriers serum are another humoral participation evidence in the Chagas disease pathogenesis. Autoantibodies against b adrenergic receptors can interfere with myocardial cell cardiac and mechanical activities. Lymphocytes from T. cruzi infected animals show specific cytotoxicity against rabbit myocardial cells. Common antigens between T. cruzi and cardiac fibers generating specific autoreactive T lymphocytes were found both in animals with experimentally induced disease and in Chagas disease carriers. The autoimmune theory suggests that myocarditis in the Chagas disease chronic phase perpetuates independently of the parasite presence. However, the myocarditis multifocal nature and the fibrosis formation in certain regions, such as the left ventricular apex and posterior wall, in chronic Chagas’ disease is difficult to explain in this context. Autopsy studies have shown that patients, even in the indeterminate form, had foci of chronic inflammatory infiltrate in the myocardium.The T. cruzi presence in the chronic phase in a quantity disproportionate to the intensity of the inflammatory infiltrate has been emphasized since the initial descriptions by Carlos Chagas. However, with more sensitive techniques use for the detection of both DNA and parasite antigens, it has been possible to demonstrate with greater frequency and intensity the parasite presence in association with inflammation in the myocardium and other organs in the Chagas disease chronic phase. T. cruziantigens were found in 100% of the Chagas disease patients hearts who died of heart failure when many samples were tested. It was observed that there was no direct correlation between inflammation intensity and parasite antigen amount, but there was a positive association between parasite antigen presence and moderate or intense myocardial inflammation (Figure 3). These studies suggest that antigens from the parasite are a trigger of the inflammatory response and that by a probable immune alteration present in the host, this response also attacks uninfected cardiac fibers. Experimental studies in mice showed similar results.

Figure 3 – Chagasic patient heart who died in CHF, showing focus of inflammatory infiltrate in the myocardium surrounding amastigotes and T. cruzi antigens detected in brown by immunohistochemistry technique.

Thus, it appears that the parasite presence in the myocardium causes immune system unbalanced activation, with consequent self-aggression of normal cardiac fibers. A strong evidence of the parasite importance in the Chagas disease chronic phase occurred with the reactivation cases detection of the disease secondary to immunosuppression. Patients with AIDS and Chagas disease in the chronic phase, even in the indeterminate form, can present clinical pictures of panniculitis, myocarditis, or Chagas’ meningoencephalitis, which are often fatal. In these cases the lesions histological appearance is similar to that described in the disease acute phase, with the numerous parasite nests presence, intense inflammation and necrosis (Figure 4).

Figure 4 – Paniculitis in a chronic chagasic heart transplant patient presenting numerous macrophages loaded with T. cruzi amastigotes. Evidence that the parasite in the chronic phase is controlled by the host’s immune system, and that immunosuppression can lead to a breakdown in this control.

It was later found that Chagas disease reactivation can follow any immunosuppression type, such as that which occurs in cases of malignant neoplasia and particularly in transplantation. Chagas disease reactivation in the heart transplant patient due to Chagas disease is a real and current problem. Its frequency varies from 0 to 20% of patients in the most current series, but it is rarely a death cause. The Chagas disease reactivation diagnosis is a controversial issue, and is classically done by direct parasite detection in blood or tissue, associated or not with a clinical picture of an acute infectious process. In the heart transplanted patient, skin nodules are common, where a macrophagic inflammatory process with numerous parasites is identified, and also the acute chagasic myocarditis appearance. The latter presents particular diagnostic difficulty with acute cellular rejection in endomyocardial biopsy. Chagas’ myocarditis and acute rejection are very similar histopathologically, and the distinction between the two can only be made by finding parasites in the former. This is done by routinely looking for parasite nests in sequential histological sections and by immunohistochemistry for T. cruzi antigens. More recently the PCR technique has been used for this purpose, with promising results. The Chagas disease reactivation occurrence in immunosuppressed patients shows that the parasite is present in the disease chronic phase, even in the indeterminate form, probably in latent form, under constant immunological surveillance. During immunosuppression, the immune surveillance disruption would lead to intense parasite proliferation and tissue invasion, similar to what occurs in the acute phase.

The parasite influencing the immune response

Experimental and human studies provide strong evidence that T. cruzi, like many other parasitic infectious agents, induces changes in the host’s immune system that allow it to evade immune attacks during and after entry into its cells. Few IL-2+ positive lymphocytes are observed in myocardial biopsies from chronic Chagas disease patients with CHF, an interleukin that is essential in activating the immune system. Chronic Chagas’ myocarditis has a CD8+ T cells predominance, which on the other hand are the cells that increase in the T. cruzi antigens presence, while the number of CD4+ T cells remains consistently low. Such evidence suggests T. cruzi immunodepressive action, which is confirmed experimentally where IL-2 administration restored the immune response in mice subjected to T. cruzi infection. Also in this context, it was observed that the control of experimental chagasic infection. in mice is mediated by CD4+ T cells and by a Th2 type response.

High correlation between the PDGF-A+ positive cells number and PDGF-B positive cells is present in the myocardium of patients with chronic Chagas’ disease. However, there is no correlation between PDGF-A and TGF-b1; PDGF-A and GM-CSF, and PDGF-B and TGF-b1. GM-CSF and TGF-b1 are important factors in controlling T. cruzi infection and are present in small amounts in chronic Chagas’ disease.

As yet unpublished studies we are developing suggest that other primitive microorganisms such as chlamydia and archaea may be in symbiosis with T. cruzi, and this particular situation could explain the increased inflammation, including of lymphocytes against non-parasitized cardiac fibers, present in patients with Chagas’ heart disease.

The parasite and changes in the microcirculationT. cruzi infection of endothelial cells favors platelet adhesion and aggregation, which could cause lesions in the microcirculation. A study of the heart in chronic Chagas’ myocardiopathy showed microcirculation dilation and epicardial coronary arteries larger diameter when compared to idiopathic dilated myocardiopathy. Studies we have performed suggest that the fibrosis in chronic Chagas’ myocardiopathy pathogenesis is at least partially of ischemic origin, not due to vascular obstruction, but due to arteriolar sphincter dysfunction. Intense vasodilation can lead to intravascular pressure loss and the blood supply distribution in the borderline regions of irrigation. Thus, sites such as the ventricular apex (between the anterior interventricular and posterior interventricular arteries) and the infero-lateral region of the left ventricle (between the right coronary and circumflex arteries) that often show thinned and fibrotic myocardial wall may be experiencing frequent ischemia episodes (Figure 5).

Figure 5 – Schematic representation of ischemic areas in double irrigation bordering territories due to vessel dilatation episodes that could explain the localized fibrosis of the conduction bundle, LV and RV tip, and LV posterior wall.

This vasodilation would be associated with the vasodilating substances presence, produced not only by the parasite, but also by the inflammatory cells. Myocardial perfusion alterations have also been observed in Chagas disease patients with chest pain without coronary obstruction, and the imbalance participation in the parasympathetic nervous system control is suggested. This alteration in myocardial perfusion may explain the greater fibrosis observed with the use of nuclear magnetic resonance imaging in Chagas disease patients with heart failure or ventricular arrhythmia, in the same locations observed at necropsy: left ventricular apex and infero-lateral regions.

The role of microparticles in Chagas disease: new advances and challenges

Small membrane-coated vesicles have been implicated in communication between human cells, including apoptotic bodies, microvesicles (also called microparticles), and exosomes. Microparticles (MPs), usually between 100 and 1000 μm, are generated from cells by sprouting from the outer cell membrane when the cell is exposed to pro-thrombotic and pro-inflammatory stimuli, cell differentiation, and senescence. MPs contain bioactive molecules that include a variety of adhesion structures, cytokines, cell receptors, microRNA, etc.

Exosomes are extracellular vesicles that range in size from 30 to 100nm, are considered to be vesicles for cellular material disposal, but also seem to play a role in intercellular communication, involving both normal cell physiology and pathological conditions. Exosomes have been found to contain microDNA, messenger RNA, microRNA, etc. and are functional in the recipient cells. In addition, it is now known that the proteins and RNA present in exosomes are highly regulated by various disease-inducing stimuli. Exosomes can induce deleterious effects on cardiac remodeling, are in greater numbers in heart failure, but exosomes are also associated with cardioprotection. On the other hand, microvesicles can also be produced and released by microorganisms. As already described, we find microorganisms in chagasic patients, mainly mycoplasma, chlamydia, and archea. The study suggested that the form with heart failure (HF) is related to higher complement activation, fibrosis and lymphocytic myocarditis due to the pathogenic archaea presence, interaction between microorganisms and higher virulence. In FI (acronym in portuguese), mycoplasma and chlamydia are not in symbiosis and, together with electron-dense non-pathogenic archaea, would lead to less complement activation, less myosin degradation without an “autoimmune” reaction.

Studies in our lab have shown that sera from chagasic patients with FI (acronym in portuguese) have a significant increase in the number of protective exosomes, possibly removing abnormal proteins such as metalloproteases from archaea. On the other hand, in the chronic form with HF, the protective exosomes are decreased in number, but there are archaeal-derived exosomes that would be related to the cardiac complications development, either by releasing metalloprotease, increasing inflammation and causing microcirculation vasodilation, but also decreasing the protective exosomes formation.

FINAL CONSIDERATIONSChagas’ cardiopathy seems to be a result of a complex Trypanosoma cruzi infection evolution. The intensity of the injuries are partly the inflammatory and ischemic lesions result. An imbalanced host immune response against T. cruzi or its products, associated with beta adrenergic system hyperactivity, can induce severe myocarditis with fibrosis. The wider fibrosis areas may be the result of low blood perfusion ischemia due to the microcirculation dilation as a result of exacerbated inflammation that also affects non-parasitized myocardial fibers. Our hypothesis is that this inflammation is due to the presence of microorganisms other than T.cruzi (mycoplasmas, chlamydia and archaea), and perhaps brought to the myocardium by it. The archaea increase inflammation by presenting antigens to CD8 + T lymphocytes, which can lead to microcirculation vasodilation. The symbiosis between these microorganisms leads to the production of circulating infectious microparticles, participating in the cardiac decompensation pathogenesis. Thus, therapeutic action in Chagas disease should include a simultaneous elimination of these different microorganisms and not just T. cruzi.

The cardiac form of Chagas disease – history

Anis Rassi

Medical School of the Federal University of Goiás

Email: anisrassi@arh.com.br

Simone Petraglia Kropf

Oswaldo Cruz House/Fiocruz

Email: simonek@coc.fiocruz.br

Carlos Chagas, in his first works about the new human trypanosomiasis discovered by him in 1909 “history”, pointed out the importance of the cardiac alterations derived from the Trypanosoma cruzi pathogenic action, in the acute phase and, above all, in the infection chronic phase. The histopathological data produced by Gaspar Vianna, also a researcher at the Manguinhos Institute, were fundamental for these alterations characterization. In the autopsies he performed, Vianna located parasites in large numbers inside the acute cases cardiac cells, with an intense inflammatory process around them, after the parasitized cell rupture.

When presenting, in 1910, the first detailed systematization of the disease clinical picture for another segment text, Chagas designated the cardiac form as one of the chronic infection forms, to be expressed by arrhythmias caused by myocardial lesions by T. cruzi. This phenomenon, he stated, was of “striking frequency and certainly never observed” in other heart diseases. Its peculiar signs were, according to Chagas, certain excitability disturbances, such as extrasystoles and, to a lesser extent, disturbances in the conduction of the stimulus, such as complete heart block, published in the article “Nova entidade morbida do homem (in portuguese)” in Brasil Médico in 1910.

In 1911, the heart form relevance was again highlighted. Despite the emphasis on thyroid elements that had been marking his formulations – and that led him to refer to the new trypanosomiasis as “parasitic thyroiditis”, Chagas noted that, in many cases, the heart rhythm irregularities were “the most easily appreciable phenomenon, which first strikes the observer’s attention, standing out as a capital element in the data supplied by physical semiotics”, as published by Chagas in 1911 in the Oswaldo Cruz Institute Memories (Memórias do Instituto Oswaldo Cruz, in portuguese)

Chagas stressed the prognosis seriousness of this clinical form, which, in an unusual frequency, led to sudden death by asystole in young individuals who often appeared to be in good health. This aspect would be progressively valued as an element of specificity in the definition of the new nosological entity and as its medical and social impact evidence, due to the vitality loss it caused among workers in full productive age in rural areas affected by trypanosomiasis.

Carlos Chagas is recognized as a pioneer in the electrocardiography introduction in Brazil, with the first reference in his works to the electrical recording of heartbeats dating from 1912. At the time, the electrocardiograph – invented in 1901 by Dutchman William Einthoven – was a string galvanometer, difficult to handle, especially with regard to the devices stability for obtaining the tracings. In addition, the record was made on photographic paper.

In 1916, due to questioning of his theses on the endocrine and neurological disorders attributed to T. cruzi infection (particularly the conception of the endemic goiter chagasic etiology), Chagas undertook an important revision of his studies. From then on, the emphasis on the thyroid aspects would recede and the cardiac aspects would be progressively strengthened. In the nationalist debate context intensified with World War I, the so-called sanitarist movement – of which Chagas was a leader, alongside Miguel Pereira, Belisário Penna and Monteiro Lobato – projected onto the public scene the chagasic endemic disease social importance. The damage to rural labor caused by the cardiac form was one of the elements justifying the demands for state intervention in favor of rural sanitation in Brazil.

In 1922, in partnership with Eurico Villela, Chagas published an extensive work on the American trypanosomiasis cardiac form, where the main physical signs of the organ progressive exhaustion (such as signs of heart failure and increase in heart volume, for example), the subjective symptoms reported by patients (such as “avexume” or “baticum”), the pathological processes evolution, and experiments with drugs to treat arrhythmias were presented in detail. According to the authors, the cardiac form should be seen as “the clinical feature par excellence of American trypanosomiasis”; the arrhythmias characteristic of this cardiopathy, they stated, constituted sufficiently individualized clinical elements to ensure the nosological entity differential diagnosis, as well as to “evaluate the disease endemic index”, as described by Chagas and Villela.

In the conference he gave in 1923 – at the famous controversy end fought at the National Academy of Medicine around his works – Chagas stated that, even if the statements regarding goiter and his formulations other aspects were refuted, one could not doubt the “inescapable signs” that substantiated American trypanosomiasis as a real and specific clinical entity. Again, he pointed to the cardiac form as “the most American trypanosomiasis interesting and characteristic aspect” to base the disease clinical specificity and epidemiological importance.

In the following years, and until his death in 1934, Chagas would continue his studies on this aspect of the disease that bears his name. An importance of this research path was his own son Evandro trajectory, who dedicated himself to the chagasic cardiopathy clinical and electrocardiographic study, especially in the acute phase.

The idea that the cardiac form study was the preferred way to overcome the doubts surrounding the American trypanosomiasis clinical definition and geographical extent was expressed in a review by the English parasitologist Warrington Yorke in 1937.

“There seems, on the whole, to be a prima facie case that American trypanosomiasis may actually be responsible for a good deal of the heart disease which is apparently so common in certain endemic areas in Brazil, Uruguay and the Argentine, and the cause of so many early deaths. […] If this should eventually prove to be the case, then American trypanosomiasis will indeed assume a pathological significance of the first magnitude. The subject is obviously one which urgently requires much further work”.

This guideline was precisely one of the work done in the 1940s dimensions at the Centro de Estudos e Profilaxia da Moléstia de Chagas, a post of the Oswaldo Cruz Institute created in 1943 in the Bambuí city, Minas Gerais. The advances in this area were basically due to the research conducted by Emmanuel Dias, Chagas’ disciple in Manguinhos and the center director, Genard Nóbrega, from the Evandro Chagas Hospital, and Francisco Laranja, a cardiologist with vast clinical and electrocardiographic experience. At the time, this was a field that was undergoing significant technical improvements, especially due to the American Frank Wilson contribution, who established the multiple precordial leads as a procedure to give greater accuracy to the heart electrical disturbances examination. Cardiology itself was undergoing an important process of institutionalization as a distinct specialty in the medical field. In 1943, the Brazilian Society of Cardiology was founded.

By performing systematic examinations of chagasic infection possible carriers in Bambuí and western Minas Gerais neighboring regions, Laranja, Dias, and Nóbrega came to establish a detailed electrocardiographic picture of chronic chagasic heart disease, individualizing it as a clinical entity before other heart diseases and ensuring the means for it to be recognized not only by specialists, but by clinicians in general.

In 1945, this electrocardiographic study first results were presented, done in a diverse universe in age range terms and infection time. Of the 183 individuals classified as chronic cases (by serologic test or xenodiagnosis), 90 (49.2%) had electrocardiograms showing myocardial damage. This work, one of the contribution main references of the Bambuí post to the Chagas disease clinical study, consolidated the notion that the chronic cardiac form constituted “the true disease clinical expression”. Even the indeterminate form, in which about 50% of the chronically ill were found, was then defined in reference to heart disease, since, according to these authors, the asymptomatic individuals presented themselves as “potential cardiacs” who, at any moment, could be surprised by the effects of the parasite’s action on their heart, as described by Dias, Laranja, and Nóbrega.

As the most indicative signs of chronic Chagas’ disease, Dias, Laranja, and Nóbrega pointed out (i) the stimulus conduction disturbances and, besides the (ii) auriculo-ventricular blocks (especially total A-V block), they highlighted the importance of (iii) right bundle branch block (RBBB), whose detection was made possible thanks to modern electrocardiographic techniques. Besides these, another sign that would be pointed out, through the Bambuí studies, as “suggestive evidence” for the chronic chagasic cardiopathy diagnosis were the (iv) ventricular extrasystoles, already indicated by Chagas as a chronic American trypanosomiasis characteristic arrhythmia.

As the number of cases identified by the Bambuí post increased, the knowledge about the Chagas disease cardiac form was consolidated. In 1948, in another great review work, based on the analysis of more than 600 disease cases, Laranja, Dias and Nóbrega provided new data on this cardiopathy specific electrocardiographic picture – “one of the most varied and curious found in cardiopathology” -, pointing out the related clinical manifestations.

Surveys among individuals not previously selected reinforced the arguments about this clinical entity specificity, showing that the electrocardiographic pattern of the cases studied in Bambuí was found in other population groups in endemic areas. In 1947, Dias, Laranja, and the physician José Pellegrino, from Minas Gerais, examined 312 individuals among the “Rede Mineira de Viação” workers and their families, proving, through electrocardiograms, the high prevalence of the chronic chagasic cardiopathy peculiar signs among those who had a positive serological reaction for T. cruzi infection. This study, published in 1948, was pioneering in using the electrocardiographic method as a criterion for epidemiological investigation. The survey conducted by Pellegrino and Borrotchin in 1948 among patients at the “Santa Casa de Misericórdia” Hospital in Belo Horizonte also confirmed the Chagas’ infection importance as a cardiovascular problems etiological factor in areas with a triatomines high frequency.

Besides the epidemiological argument, another important argument regarding the chagasic heart disease specificity was Pellegrino’s reproduction, in dogs experimentally inoculated with T. cruzi, of a chronic heart disease with the same electrocardiographic, radiological, and clinical features found in humans. At the same time, the advances produced, at the time, in serological diagnosis methods, especially in the complement fixation reaction (Guerreiro and Machado reaction), facilitated the chronic infection cases recognition, corroborating the data from clinical research.

As a result of this research set, Laranja, Dias and Nóbrega stated, in 1948, that the experience acquired in Bambuí allowed them the conviction that American trypanosomiasis in its chronic phase “finds clinical expression essentially in a well-defined cardiopathy in its anatomopathological, clinical, radiological and electrocardiographic characteristics, allowing them secure individualization”. Summarizing the Bambuí post contribution, Laranja considered it the beginning of a new phase in the Chagas disease “eventful history”, in which the chronic chagasic heart disease recognition as a “indisputable reality clinical entity” provided the “widespread skepticism” overcoming that had been imposed on the subject after the discovery initial phase and the first studies by Chagas and his collaborators, as described by Laranja in 1949. The work of Laranja, Dias Nóbrega and Miranda would be internationally disseminated, in 1956, in the prestigious American journal Circulation, in an article that would be one of the most cited in the literature on Chagas disease.

In the 1950s, an important contribution to the studies on chagasic heart disease was the work of the Austrian pathologist Fritz Köberle, hired in 1953 by the recently created Ribeirão Preto Medical School. His research has contributed to explain, in part, the pathogenesis of the morphological and functional changes specific to chronic Chagas’ disease. According to him, such alterations had as a determining factor a denervation process caused by lesions in the heart’s autonomic nervous system, similar to what he found in other organs, mainly the esophagus and the colon.

The second half of the 20th century was very prodigious in the chronic chagasic cardiopathy diagnostic and therapeutic advance, which, due to its potential for morbidity and mortality, represents a serious public health problem, and has three syndromes as manifestations: (i) the arrhythmic syndrome, (ii) heart failure, and (iii) thromboembolic syndrome, which present themselves alone or in association, and whose gradation depends on the disease evolution stage. In general, the heart failure and thromboembolism syndromes behave like those of other myocardiopathies, but the arrhythmic one, because of its frequency, variety, quality, and severity, makes chronic Chagas’ disease an unusual model for clinical, therapeutic, electrocardiographic, electrophysiological, and prognostic investigation. Tiredness and dyspnea at increasingly lower efforts, palpitation, pre-syncope, and syncope are symptoms that are present in patients with Chagas’ disease, and it is important to point out that there are cases of heart disease (especially in its initial stage) without any symptoms; in these cases, the diagnosis is made by finding electrocardiographic alterations suggestive of the etiology (such as right bundle branch block, left anterior hemiblock, ventricular extrasystole, and 1st, 2nd, and 3rd degree atrioventricular block) and subject to confirmation by performing diagnostic serologic tests. Thromboembolism occurs in both the large circulation (brain, limbs, kidneys, spleen, etc.) and the small circulation (lungs).

This advance came about thanks to the laboratory methods introduction, the devices invention, and the pharmaceutical products synthesis to better and more thoroughly examine and treat its different manifestations. There have been so many of them, and of such value, that they have made hitherto relatively new resources obsolete.

The electrocardiograph itself received several improvements, such as direct inscription on heat-sensitive paper, weight and size reduction – allowing it to be portable -, battery operation – of great use in population surveys in rural areas -, automatics, etc.

The dynamic electrocardiography (Holter) and the exercise test (initially used in order to study ischemic heart disease) have become, since the 1970s, unusual pieces to quantify and qualify the tachycardic and/or bradycardic arrhythmias, symptomatic and, especially, the asymptomatic ones, guiding the treatment with accuracy. Such methods are also of great value in the antiarrhythmic drugs therapeutic evaluation and in the possible pro-arrhythmic effect detection of the same, in the implanted artificial cardiac pacemaker functioning evaluation, in the medical-worker evaluation and in the identification of patients at higher sudden cardiac death risk.

Similarly, the echocardiogram that, from the primitive M-mode, was soon improved to two-dimensional and, currently, color dopler, providing cardiac morphodynamics data not supplantable by any other noninvasive investigation method. Echocardiography has become more valuable than radiological examination of the heart by allowing the myocardial contractility study, by determining more accurately the cavities diameters, by reporting the ventricular ejection fraction value, and by detecting ventricular aneurysm and intracardiac thrombosis. The radiological examination left the pulmonary vascular stasis diagnosis.

The serological laboratory diagnosis of the Chagas disease chronic phase has been modified. The Guerreiro-Machado reaction, introduced in 1913, was replaced by simplified methods, such as indirect hemagglutination, indirect immunoflorescence and ELISA, all with sensitivity and specificity around 98%. In parasitological diagnosis, xenodiagnosis has been modified to increase its sensitivity, and today it is better standardized and can be performed by artificial or indirect technique. At the same time, hemocultures have also undergone considerable advances, allowing for even greater diagnostic sensitivity than xenodiagnosis. It must be emphasized that both xenodiagnosis and hemocultivation are, fundamentally, tests performed in research laboratories on patients undergoing specific treatment to evaluate the treatment results.

Another method, this semi-invasive, for the chronic chagasic cardiopathy investigation, which has been in use since 1980, is represented by the diagnostic and therapeutic intracardiac electrophysiological study, performed through catheterization, to study the specific conduction system, whose main indications consist in the sinus node function evaluation, atrial and intra-ventricular blocks study, differential diagnosis between paroxysmal ventricular tachycardia and paroxysmal supraventricular tachycardia with aberrant conduction, malignant tachyarrhythmias reproduction, arrhythmogenic focus location and ablation of the same transcatheter using direct electrical current.

From a drug treatment point of view it was also model progress. Mercurial diuretics gave way, starting in the 1960s, to the more potent and less toxic furosemide and thiazides. In recent years, angiotensin-converting enzyme inhibitors, spironolactone, platelet anti-adhesives, anti-coagulants, and also beta-blockers, have been incorporated into the heart failure and thromboembolism clinical treatment. Here, the new antiarrhythmics introduction, especially amiodarone, from the 1970s on, in the treatment of ventricular arrhythmias, to significantly improve the chronic Chagasic cardiopathy prognosis, is also noteworthy.

For bradyarrhythmias, sadly assisted until then by cardiologists and culminating with sudden death, due to the clinical treatment inefficiency, since the 1960s there has been the use of implantable cardiac pacemakers (initially epimyocardial, requiring thoracotomy, and later endocardial via transvenous), which has undergone successive improvements over the years. So many improvements have been made that, in the specialists’ jargon, it is said that, before, the pacemaker patient had to adapt to it, whereas, today, it is the pacemaker that adapts to the needs of those who have it implanted; moreover, its generator is small (3 cm in diameter and 0.5 cm thick) and light (10 g). For ventricular tachyarrhythmias (sustained ventricular tachycardia and ventricular fibrillation) another electrical resource was invented, and used from the 1990s on. This is the implantable cardioverter-defibrillator, which, according to its programming, delivers a shock, practically unnoticed by its bearer, in the face of paroxysmal tachycardia and, necessarily, during ventricular fibrillation, when the patient is unconscious (syncope). It also, primitively, like the pacemaker, was of large proportions and heavy. Today, however, their dimensions and weight have diminished a lot, to the point of allowing transvenous implantation, and more, they also have a pacemaker function, to treat eventual bradyarrhythmia that, with relative frequency, is associated with tachyarrhythmia in chronic chagasic cardiopathy.

For chronic Chagas’ disease terminal cases, especially irreducible heart failure, heart transplantation is the solution. In its history there are two periods: (i) the first, from 1967 (when it was performed by Christian Barnard) to 1980, and (ii) the second, after 1980, period in which new immunodepressant drugs were incorporated into the treatment, significantly improving the prognosis. The first heart transplantation in chronic chagasic patients was performed in 1985, and up to the present hundreds of cases have been performed in 57 accredited hospitals, with results as good or even better than those performed for heart disease of other etiology. In the context, T. cruzi has to be considered, in view of the immunodepressants use, which can cause the infection reactivation. There is no consensus on the trypanosomicide prophylactic use before or after surgery. We are of the opinion, on an empirical basis, that it should be performed (preoperatively if the patient’s general conditions allow) or, otherwise, in the immediate postoperative period, using benznidazole (5 mg/Kg/day for 60 days).

Another resource for the severe chronic chagasic heart disease treatment was recently adopted by a group of researchers from Bahia, led by Ricardo Ribeiro dos Santos, which consisted in the stem cells from the patient’s own bone marrow implantation, based on literature data. As the initial results of this study were promising, the Ministry of Health provided a multicenter, national, double-blind, placebo-controlled trial, already underway, to ascertain the method real value. It will include 300 patients with chronic Chagas’ disease (NYHA functional class III and IV), half of whom will be treated with bone marrow stem cells in intracoronary injection and the other half with placebo. We look forward to the result.

All these diagnostic and therapeutic advances have gradually allowed the optimization of the care given to people with Chagas’ disease, changing the chronic Chagas’ disease prognosis in the second half of the 20th century in favor of a better quality and greater quantity of life.

In parallel to all the semiotechnical and therapeutic achievements observed in the second half of the 20th century, there was a great increase in the study of chronic chagasic cardiopathy under all its aspects, making it an obligatory and frequent subject in medical congresses and related meetings, as well as the research centers formation in several centers, such as: Ribeirão Preto (SP), Goiânia (GO), Belo Horizonte (MG), Uberaba (MG), São José do Rio Preto (SP), São Paulo (SP), Salvador (BA), Rio de Janeiro (RJ), Brasília (DF), Recife (PE), Uberlândia (MG), Campinas (SP).

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Heart Disease – Functional Assessment

The Cardiac Form of Chagas Disease – Current Reality and Functional Assessment

Manoel Otávio da Costa Rocha

Department of Internal Medicine, School of Medicine, Federal University of Minas Gerais

Email: rochamoc@medicina.ufmg.br

Márcio Vinicius Lins Barros

Cardiology Sector, Clinical Hospital, Federal University of Minas Gerais

Email: marciovlbarros@uolcom.br

Vitor Tadeu Vaz Tostes

Intensive Care Center, Clinical Hospital, Federal University of Minas Gerais

Lucas Lodi Junqueira

Cardiology Sector, Clinical Hospital, Federal University of Minas Gerais

Antônio Luiz Pinho Ribeiro

Department of Internal Medicine, School of Medicine, Federal University of Minas Gerais

Email: tom@hc.ufmg.br

Infectious myocardiopathies, also called infectious myocarditis, can be caused by a range of agents (viruses, rickettsiae, bacteria, fungi, protozoa, and helminths). The most common myocarditis worldwide is that caused by Trypanosoma cruzi, the protozoan parasite that causes Chagas disease, endemic in rural areas of South and Central America. Chagas disease remains a major health problem in Latin American countries, where it is estimated that there are approximately 20 million infected patients, 25-35% of whom will develop cardiovascular changes. It is also known that left ventricular dysfunction represents the greatest mortality predictor in Chagas disease, and that asymptomatic dysfunction is at least as frequent as symptomatic dysfunction.

The initial infection is usually asymptomatic, but manifestations may occur in the acute phase, including cardiac involvement (acute myocarditis), but there are few reports of studies using complementary exams in this phase.The Chagas disease chronic phase can be subdivided into the indeterminate chronic form and the so-called determined chronic forms (cardiac, digestive, cardio-digestive, and nervous). About 50% of infected patients in endemic areas are in the indeterminate chronic form, and although these patients main characteristic is the absence of significant clinical, electrocardiographic, and radiological abnormalities, morphofunctional cardiac alterations have been observed when more sophisticated complementary methods are used, such as ergometry, dynamic electrocardiography, non-invasive autonomic tests, radioisotopic ventriculography, and echocardiography, as reviewed by Ribeiro and Rocha. Although several longitudinal studies have demonstrated the good prognosis of patients in the indeterminate chronic form, about 2-5% of these patients progress to one of the disease other chronic forms, usually with a benign pattern, eventually progressing to severe and potentially lethal forms. It is estimated that within 5 to 10 years, about 30% of these patients evolve to chronic heart disease. Moreover, sudden death has been reported in Chagas patients as the first disease clinical manifestation. Therefore, the identification of myocardial damage early markers in Chagas disease is important in risk stratification, so that individualized conducts can be established, improving the quality of life and longevity in these patients, as proposed by Rocha and collaborators (Figure 1).

Figure 1

Methods for non-invasive functional assessment of Chagas’ cardiopathy

The non-invasive techniques of complementary cardiovascular propaedeutics are available in our country, in most of the medium-sized cities, and have been incorporated into the approach to patients with cardiomyopathy in reference outpatient clinics, research centers and, in some special cases, by the Unified Health System and Social Security. They constitute techniques of great importance in the evolutive, therapeutic, medical and prognostic clinical evaluation.

The information relevance, which can be obtained by using exercise testing, conventional and dynamic electrocardiography (Holter), echocardiography, myocardial scintigraphy, and magnetic resonance imaging, makes it necessary to disclose the main methods fundamental characteristics, the parameters that can be analyzed, and their indication in the infectious cardiomyopathies evaluation, such as chronic chagasic cardiopathy. On the other hand, its more frequent use becomes a concrete need, when it is intended to adequately measure a significant number of patients with heart disease, especially those who are symptomatic, with suspected functional impairment, or who have a work activity that requires too much or continuous physical effort.

Conventional Electrocardiography

The electrocardiogram is a more sensitive and specific method in the diagnosis of myocardial involvement in Chagas disease than other evaluation easily accessible means, such as the anamnesis, physical examination and chest X-ray. However, the method sensitivity in detecting myocardial damage is not high. The electrocardiographic alterations absence is not a reliable indicator of the cardiac involvement absence. When studied by more sophisticated propaedeutic methods, a variable proportion of patients with normal electrocardiograms show heart structural or functional alterations. Additionally, between 20 and 50% of these patients will develop electrocardiographic changes suggestive of Chagas’ disease when followed for about 10 years. Regardless of these considerations, however, the medium-term prognosis of the chagasic patient with normal electrocardiogram is excellent: in a longitudinal study with seven-year follow-up in a rural community, Maguire and collaborators found no differences with respect to mortality between seropositive and seronegative individuals with normal ECG.

Although there are some electrocardiographic alterations that are more suggestive that the cardiac involvement is, in a given case, secondary to the chagasic etiology, almost all the existing electrocardiographic abnormalities can be found, with abnormalities predominance of the formation and conduction of the heart rhythm, as described by Rosembaum and Alvarez. The combination of different electrocardiographic alterations in the same tracing can occur, and is more frequent in those with more advanced heart disease and worse prognosis. In the hypertensive heart disease presence or other etiologies, electrocardiographic alterations characteristic of these conditions may overlap with those typical of chagasic heart disease. Electrocardiographic alterations suggestive of myocardial damage by T. cruzi occur also in other pathological processes, so that the electrocardiogram is not, at all, a method with high specificity in the Chagas’ disease detection.

Complete or incomplete right bundle branch block of the His bundle is the most frequent conduction disturbance in Chagas’ disease, being found in 10 to 50% of infected patients, depending on the casuistry studied. It is frequently associated with blockade of the left bundle branch anterosuperior fascicle of the His bundle (left anterior hemiblock), a Chagasic cardiopathy characteristic combination. Other times, it is associated with left inferior-posterior blockade (left posterior hemiblock), incomplete atrioventricular blocks, ventricular extrasystoles, or other less frequent alterations. For reasons not completely understood, left bundle branch block is at least 10 times less frequent than right bundle branch block in Chagas disease.

The QRS complex duration is directly related to the left ventricle dimensions and, inversely, to the LV systolic function, so that Chagas patients with intraventricular conduction disturbances more often present depression of left ventricular function, as shown by Ribeiro and collaborators. Additionally, the intraventricular blocks presence, especially right bundle branch block, is known to significantly increase the fatal outcome risk among the infected, as described by Maguire and collaborators. There are no data, however, that inform whether the intraventricular blocks presence constitutes an independent prognostic factor for ventricular function in the Chagas’ disease evolution.

Ventricular extrasystole is also a very frequent finding in Chagas’ disease, affecting 6 to 55% of the serologically positive individuals. Characteristically, extrasystoles are frequent, polymorphic, and complex, and occasionally repeated (pairs) or sustained (ventricular tachycardia) forms of ventricular arrhythmia are recorded on short routine tracings. The arrhythmia paroxysmal nature makes conventional electrocardiography not the ideal method for its detection. Thus, when patients with resting electrocardiogram changes and heart failure are studied using dynamic electrocardiography, ventricular extrasystoles are found in 99% of the cases, and in 87% multiform extrasystoles or repeated forms are found, as described by Carrasco and collaborators.

Ventricular arrhythmias are more frequent, complex and sustained in patients with worse left ventricular function. The complex extrasystoles presence on the ECG is related to an increased fatal outcome risk, as described by Maguire and collaborators. Additionally, Casado and collaborators have shown that patients with intraventricular blocks and ventricular arrhythmia on the ECG are those with the greatest left ventricular dilatation and the cardiac contractibility most pronounced depression. Guerrero and collaborators described that in a Chagas patients population with depressed ventricular function on echocardiography, the complex ventricular extrasystole presence is an independent risk predictor of progression to death.

In the follow-up of the ventricular arrhythmias treatment, the electrocardiogram is useful mainly for monitoring the electrophysiological effects of the antiarrhythmic drugs use. PR interval Prolongation, widening, atrioventricular and intraventricular blocks, and QTc interval prolongation can occur. The latter effect is related to prolonged QT syndrome and potentially fatal ventricular arrhythmias, such as “torsade de pointes”-ventricular tachycardia type. The ventricular extrasystoles disappearance on a routine ECG performed after the antiarrhythmics use is not sufficient evidence that the drug was able to suppress the arrhythmia, due to the ventricular arrhythmias paroxysmal nature.

The sinus node involvement by Chagas’ cardiopathy was first postulated by Brazil when he described the absence of sinus node response to physical or pharmacological stimuli in 19% of Chagas’ patients. Later, sinus node structural and functional involvement was demonstrated in a patients high proportion with Chagas disease. However, only a minority show sinus dysfunction manifestations on surface ECG (1 to 16% of patients). The main electrocardiographic manifestations of sinus node dysfunction are (i) sinus bradycardia, especially if the sinus rate is less than 40 bpm, (ii) sinus arrest, (iii) 2nd degree sinoatrial block, and (iv) the surrogate rhythms existence denoting inhibition of the normal sinus pacemaker: the junctional and accelerated idioventricular rhythms.

The conduction system involvement extends to the atrioventricular node, the His bundle, and its branches. Although AV node functional changes on invasive electrophysiological study are present in a large number of heart disease patients, most atrioventricular blocks (AVB) occur due to injury distal to the His bundle trunk. BAVs of (i) first degree, (ii) second degree (types I, II, 2:1 or advanced), and (iii) third degree or complete BAVs can occur. First and second-lobe AVBs can be associated, among other electrocardiographic alterations, with intraventricular conduction disorders. When concomitant to right bundle branch complete block associated with left hemiblock, they usually denote advanced and diffuse lesion of the conduction system and, probably, a greater probability of progression to complete block. In CHB, the ventricles are depolarized by subsidiary pacemakers, usually located distal to the His bundle division, generating slow idioventricular rhythms (frequency lower than 40 bpm) and enlarged QRS complexes.

Atrial fibrillation is the most common supraventricular arrhythmia among patients with Chagas’ disease, being found in 4 to 12% of electrocardiographic tracings. Most of the time, atrial fibrillation presents in a chronic form, being associated with pronounced myocardial damage, diffuse involvement of the conduction system, ventricular arrhythmias and, consequently, a dismal prognosis, as initially described by Rosenbaum and Alvarez and Dias and Kloetzel, being negatively related to survival, other variables independently, in a multiple regression model, as shown by Espinosa and collaborators. Supraventricular extrasystoles are less frequent and less important than ventricular extrasystoles, occurring between 1.5 and 12% of the Chagas’ patients.

Other significant changes found in chagasic heart disease include (i) electrically inactive zones (simulating acute myocardial infarction), (ii) low peripheral voltage, and (iii) primary T-wave changes, although known to be frequent, these changes prevalence varies widely among different series, in part due to the different diagnostic criteria use. Electrically inactive zones and repolarization changes were related to worse left ventricular function, with Salles and collaborators reporting a threefold increased risk of death and a sixfold increased risk of sudden death in the abnormal T-wave axis presence, even after correction for covariates such as left ventricular ejection fraction. Increased QT interval dispersion was also associated with left ventricular dysfunction and increased risk of death by Salles and collaborators.

Ergometry

Chagas disease patients without clinical, radiological and electrocardiographic alterations may present abnormal parameters in their ergometric evaluation. Studies in patients with normal electrocardiogram and chest radiological study have revealed abnormal pressure and chronotropic responses, as well as ventricular arrhythmias during exercise, as described by Gallo Jr. and collaborators, Marins and collaborators and Ribeiro and collaborators.

The exercise stress test allows the effort capacity quantification of individuals, in addition to providing information about the heart rhythm behavior during physical activity. Undoubtedly, it is a fundamental importance method in the working capacity evaluation of patients with Chagas’ disease, and should be used as a parameter to establish criteria for work admission and retirement, especially in those who present cardiac impairment clinical or electrocardiographic evidence. Its use is recommended for those individuals who intend to exercise high-risk activities, such as heavy physical labor or professions that put the lives of others at risk, such as drivers, pilots, etc, as proposed by Rocha.

Although several studies have shown that alterations such as abnormal pressure and chronotropic responses, low body oxygen consumption rates, and ventricular arrhythmias related to effort are prevalent in the evaluation of patients with Chagas’ cardiopathy, as shown by Faria and Molina and collaborators, no direct correlation was found between the degree of ventricular dysfunction and such abnormalities, as published by Tostes. It is suggested that the organic response to effort in patients with Chagas’ disease is influenced by other factors besides left ventricular function. Autonomic denervation and structural and functional lesions in the conduction and stimulus generation system may be related to the abnormal hemodynamic responses to effort.

Exertion can cause supraventricular and ventricular cardiac arrhythmias, both in patients with heart disease and in individuals with normal cardiovascular systems. The increase in adrenergic tone and the parasympathetic tone modulation determine electrophysiological alterations that favor the establishment of important arrhythmogenic mechanisms, such as: (i) increased automaticity, (ii) late potentials, and (iii) reentry circuits, as shown by Podrid and collaborators.

Chagas’ cardiopathies, as they present with fibrosis intermingled focal areas with intact myofibrils, possess a vast anatomic substrate for heart rhythm disorders, being particularly susceptible to arrhythmogenic mechanisms triggered by effort, as shown by Rassi and collaborators. Ventricular arrhythmias are among the most prevalent abnormalities in the patients ergometric evaluation with Chagas disease and most studies, including those of Faria and Molina and collaborators, associate the complex ventricular arrhythmias finding during effort with the presence of ventricular dysfunction or ventricular arrhythmia in the resting electrocardiographic tracing. The presence of ventricular tachycardia on exertion is a death risk independent predictor in Chagas’ cardiopathy, as shown by Paola and collaborators.

Ergometry can be used as a therapeutic evaluation method both in patients with heart failure and in those undergoing antiarrhythmic therapy.

It is known that functional capacity has great prognostic importance in people with Chagas disease. Mady et al. demonstrated a survival rate of 97% in functional class II patients, 58% in functional class III patients, and 16% in functional class IV patients, categorized according to the criteria of the New York Heart Association.

Ambulatory Electrocardiography (Holter)

In Chagas’ disease, ambulatory ECG has been used primarily for the cardiac arrhythmias evaluation for (i) diagnostic, (ii)prognostic, and (iii) therapeutic purposes.

One of the ambulatory ECG main uses is the cardiac arrhythmias diagnosis in patients with unexplained cardiovascular symptoms, especially those attributed to arrhythmias: palpitations, dizziness, and syncope. While the method`s great value in many of these patients, there are several limitations to its use for this purpose. In order to attribute a symptom to a particular rhythm change, it is necessary to make the temporal correlation between the presenting symptom, noted in the diary by the patient with or without the use of the recorder’s event marker, and the significant arrhythmias presence on the simultaneous electrocardiographic tracing.

Since the symptoms are usually occasional, in most examinations patients do not show the manifestation during recording. Among those who present symptoms during the examination, at least half do not show simultaneous electrocardiographic changes. Thus, in only about a quarter of symptomatic patients will the method reveal an arrhythmia causing the manifestation. On the other hand, the arrhythmia absence on the electrocardiographic tracing in a patient who presented symptoms during the recording helps in its exclusion as the symptom cause in question.

Among patients who, although symptomatic, have no symptoms during recording, silent arrhythmias are found in 4 to 30% of cases. The diagnostic value of these silent arrhythmias presence in these patients is not known. It is possible that the threshold of these symptoms perception varies, and that in some situations the arrhythmic event causes symptoms and in others it is silent. However, some silent arrhythmias may have prognostic value and indicate the need for therapeutic measures, such as sustained ventricular tachycardias and complete atriventricular blocks with slow ventricular outflows.

There are no papers estimating the diagnostic value of ambulatory ECG specifically in symptomatic patients with Chagas disease, although our personal experience confirms the above statements. However, the method can be used for early myocardial damage diagnosis in patients with Chagas’ disease without apparent heart disease, revealing increased ventricular ectopy frequency when compared to normal controls, according to studies by Almeida and collaborators, Marins and collaborators, Rassi Jr. and collaborators, Ribeiro and collaborators.

The ambulatory ECG use in the prognostic evaluation has established value in patients after acute myocardial infarction and with hypertrophic cardiomyopathy, in which the frequent detection, polymorphic and repeated ventricular extrasystoles, such as pairs and ventricular tachycardia episodes, are related to increased mortality. In Chagas’ disease, the presence of complex and/or sustained ventricular arrhythmias is more frequent in those patients with more pronounced myocardial damage, as shown by Carrasco and collaborators and is an independent marker of increased death risk in those with ventricular function depression, as described by Guerrero and collaborators.

However, since the anti-arrhythmic treatment value is not known in these situations, the method does not have universal indication in asymptomatic Chagas’ disease patients, since there are other prognostic indicators, clinical or through simpler complementary methods, of easier access and lower cost.

In patients undergoing therapeutic interventions, such as the antiarrhythmic drugs use for supraventricular or ventricular arrhythmia, the ambulatory ECG can be used to control therapeutic efficacy, provided that the aforementioned phenomenon of spontaneous variability is considered. In the ventricular arrhythmia treatment, treatment is considered to be effective when there is a 60-80% reduction in the frequency of ventricular extrasystoles and complete repetitive forms suppression. In the patient with a cardiac pacemaker, indicated for the treatment of atrioventricular block or sinus node disease, special recorders can be used for the pacemaker dysfunction detection and in the intrinsic rhythm response evaluation to effort and to the patient’s usual daily stress.

Autonomic tests

The involvement existence of the autonomic nervous system (ANS) in Chagas disease was postulated already by Carlos Chagas, in 1913, and it is attributed to Möckenberg, in 1924 (apud Köberle 1958), the first lesions description in cardiac autonomic ganglia and nerve fibers during experimental infection in dogs. But it was Fritz Köberle, in fundamental anatomopathological studies carried out in the 1950s and 1960s, who demonstrated the existence of autonomic nervous system in Chagas disease important involvement especially the parasympathetic nervous system. In the late 1960s, studies by the group of Amorim, Marin-Neto, Gallo Junior, Manço and collaborators, in Ribeirão Preto, confirmed the functional involvement existence of the ANS in chagasic cardiopathy. Since then, much has been written about the autonomic dysfunction possible role in Chagas disease. The sections below summarize the literature current state.

Most studies evaluating autonomic function indicate that the dysfunction is predominantly parasympathetic. Thus, chagasic heart disease patients, even without heart failure, frequently present an abnormal response to pharmacological or reflex vagal inhibition, manifested by an absence of sinus tachycardia after administration of atropine or during phase two of the Valsalva maneuver. These patients also fail to develop bradycardia when subjected to reflex vagal stimulation by physiological (phase four of the Valsalva maneuver, facial immersion in water) or pharmacological methods, intravenous infusion of metaraminol and phenylephrine, according to Amorim and collaborators, Manço and collaborators and Amorim and collaborators. Even patients with normal response to atropine and Valsalva maneuver presented abnormal response when submitted to baroreflex sensitivity evaluation, as shown by Junqueira and collaborators. It was also demonstrated that a reduction in respiratory sinus arrhythmia, a vagal activity index, occurred during deep breathing, with standardized tidal volume and respiratory rate, as described by Marin-Neto and collaborators and Ribeiro and collaborators. In 24-hour Holter tracings, a significant decrease in spectral power of night and morning chronotropic variability was observed, both of the low and high frequency components, shown by Emdim and collaborators, in addition to heart rate variability vagal indices reduction in the time domain and the three-dimensional return map, a new nonlinear method, as used in Chagas disease by Ribeiro and collaborators.

All these data show that many Chagas disease carriers are deprived of the tonic inhibitory vagal action on the sinus node, present in normal individuals, besides not presenting a fast vagus-dependent bradycardizing mechanism, responsible for the fast reflex modulation to transient elevations in blood pressure, found in physiological and pathological conditions, as described by Amorim and Marin-Neto.

The existence of sympathetic nervous system, cardiac, and vascular involvement, however, is still a matter of controversy. The Cordoba school authors, as reviewed by Iosa, found, based on autonomic functional tests, sympathetic involvement significant evidence, with a lower increase in circulating norepinephrine compared to patients with heart failure of other etiologies and no increase in the low-frequency spectral component in HRV analysis during active postural test and isometric effort, suggesting sympathetic involvement. Using the passive body tilt test, Marin-Neto and collaborators also found cardiac sympathetic system impairment evidence. However, the authors from the Ribeirão Preto group did not find significant changes in the sympathetic response to effort, as shown by Gallo and collaborators, nor in the vascular autonomic control, having not documented significant changes in the pressure response to maneuvers such as isometric effort or the passive tilt test, described by Marin-Neto and collaborators. Thus, the sympathetic dysfunction importance and severity is still subject to controversy.

When studying patients with Chagas disease various forms, it is recognized that, in most cases, autonomic indices gradually change as the heart disease worsens. Thus, the alterations in autonomic functional parameters found in the indeterminate form or in the heart disease absence are almost always less intense than those found in patients with evident heart disease, being more pronounced in those with heart failure and cardiodigestive forms (link to another segment), as described by Iosa and Marin-Neto and collaborators. However, studies from our group have shown that there is significant vagal dysfunction even in patients without apparent heart disease, also indicating that dysautonomia is independent of deterioration in left ventricular function, as published by Ribeiro and collaborators and Oliveira.

Although the autonomic dysfunction mechanism has not been clarified, evidence derived from anatomopathological correlations, described by Amorim and collaborators and experimental correlations, shown by Junqueira and collaborators indicate that, in a substantial fraction of the cases, the vagal control alteration over the heart is related to the intracardiac parasympathetic autonomic nervous system morphological lesions presence. On the other hand, Iosa and collaborators attribute the autonomic nervous system functional alterations to an adrenergic and muscarinic receptors progressive blockade, followed by denervation, related to autoimmune mechanisms, mainly the presence of antibodies against autonomic receptors, as reviewed by Borda and Sterin-Borda, and of anti-ganglioside antibodies, as shown by Iosa.

Reduced vagal autonomic influence on the heart, detected through autonomic testing or through reduced heart rate variability, is a known risk factor for cardiac death and, in particular, sudden death in other clinical conditions, such as post-infarction patients, described by Kleiger and collaborators. In Chagas disease there is also evidence of these variables independent prognostic importance, shown by Veloso. Additionally, autonomic dysfunction could be implicated in other pathophysiological processes in Chagas’ disease, such as in the coronary microangiopathy genesis, one of the most important Chagas’ disease physiopathological manifestations. Further studies are needed to clarify what is the real role of autonomic dysfunction in the pathophysiology and natural history of Chagas disease.

Echocardiogram

Echocardiography (link to box) has become an essential complementary examination not only for pathological changes precise localization, but also for determining the cardiac involvement evolutionary stage and severity, providing excellent indices for therapeutic and prognostic guidance, as reviewed by Rocha and collaborators. Echocardiography, combined with Doppler techniques, allows for a morphofunctional approach to the heart in a non-invasive and innocuous way. Besides having a relatively low cost, it presents a diagnostic reliability high degree, being, therefore, a propedeutic element of high value in the approach to the chagasic patient.

Initial studies using M-mode echocardiography demonstrated changes in the various Chagas disease clinical forms. Monti and collaborators detected alterations in 40% of the patients with subclinical Chagas’ disease and in 100% of the cases in symptomatic patients, and Finaret and collaborators, analyzing 152 patients with asymptomatic chronic Chagas’ infection, without alterations on ECG evidence, chest X-ray, and ergometry, showed echocardiographic abnormalities in 21.3% of the cases, with left ventricular cavity enlargement being the most commonly found finding. In our midst, Friedman was the first to publish about echocardiography in Chagas disease. However, M-mode echocardiography has several limitations in evaluating the heart. By determining only the visualization of interventricular septum and posterior wall segments, the other segments contractility analysis is not possible, including the cardiac apex study, an important segment in the Chagas’ disease evaluation, due to the involvement frequency and peculiarities of this region in this pathology. In addition, right cavities analysis, valvulopathies quantitative evaluation, intracavitary thrombi detection, diastolic function (DF) determination and pulmonary arterial pressure, high importance data in global cardiac evaluation, cannot be measured by this technique. The introduction of two-dimensional (box) and Doppler (box) techniques has added Chagas disease patients important information in the evaluation.

One of the most interesting aspects related to the cardiac involvement in Chagas disease is the myocardial contractility segmental involvement, showing, in this disease, aspects similar to those of coronary artery disease. The predominantly involved segments are the left ventricle inferior-posterior wall and the apex. Segmental involvement can be found in about 20-30% of patients in the indeterminate form and almost universally in those with advanced heart disease, and can be seen in about 25% in patients under 30 years old, as shown by Acquatella and collaborators, Ortiz and collaborators, Câmara and Xavier-Salles. The apical aneurysm represents a characteristic lesion in Chagas disease, being found in about 16 to 46% of Chagas disease patients and having an important prognostic impact. Two-dimensional echocardiography allows the identification of almost all apical aneurysms, including the smaller ones. It is important to note that apical dyssynergia in Chagas disease is not related to involvement of the left ventricular anteroseptal wall, as often occurs in ischemic heart disease.

Several studies reflect the study importance of left ventricular contractility in the patients with Chagas’ disease prognostic evaluation, regardless of their clinical stage, and the performance of at least one echocardiographic study in a chagasic patient can bring valuable information regarding the risk stratification of this individual, as shown by Mady, Bestetti and Muccillo, Rodriguez-Salas and collaborators, and Xavier-salles. Sérgio Xavier Salles, after cohort echocardiographic follow-up of 604 patients over a period of nine years, demonstrated that the left ventricular systolic diameter by echocardiography evaluation was one of the variables that best explained the death evolution from cardiac causes in patients with Chagas disease. Mady and collaborators and Bestetti and Muccillo demonstrated a worse prognosis in patients with Chagas’ disease who presented with ejection fraction decreased by two-dimensional echocardiography. Maciel and collaborators demonstrated greater myocardial damage in patients with Chagas’ disease who presented contractile abnormalities compared with those who presented only electrocardiographic alterations. Almeida Filho and collaborators demonstrated ventricular function deterioration in 69.2% of the patients who presented with segmental contractile abnormalities and normal global function, while they observed worsening in only 22.2% of the patients who did not present with contractile abnormalities at the clinical follow-up beginning.

The diastolic function parameters evaluation in Chagas’ disease has demonstrated an early abnormality of the left ventricular relaxation in patients with Chagas’ disease, as described by Caeiro and collaborators, Martinez Filho and collaborators and Sousa and collaborators. Chagas’ cardiopathy can lead to impairment of diastole both phases, initially determining alterations in ventricular relaxation and, progressively, disorders related to complacency. The diastolic function Doppler analysis was especially analyzed in Cunha study, in which although signs of altered relaxation were demonstrated in the Chagas disease patients group, throughout the period studied, in no cases were pseudonormal and restrictive flow patterns detected, which are related to more severe disturbances of relaxation and ventricular compliance. The characterization of the various patterns of ventricular filling in chronic Chagas disease using Doppler echocardiography was studied by Barros and collaborators who demonstrated that 73.6% of patients with left ventricular ejection fraction below 50% presented with some diastolic dysfunction degree, while patients with normal ejection fraction, ventricular diastolic diameter and parietal motility score presented with normal diastolic function in 92.6%, 93.8% and 94.7% of cases, respectively.

The fibrosis (link to another segment) Chagas’ myocarditis characteristic contributes to the heart progressive stiffening, determining various patterns of diastolic dysfunction during the heart disease evolution. The evidence of elevated ventricular filling pressures by flow Doppler analysis (pseudonormal and restrictive patterns) correlates with the left ventricular systolic dysfunction presence in almost all the patients studied. Therefore, in chronic Chagas’ disease, the presence of ventricular filling patterns with increased diastolic pressures signs seems to be related to the myocardial fibrosis degree, with subsequent contractile impairment of muscle mass and consequent systolic dysfunction. These patients follow-up becomes relevant due to the possibility that the diastolic dysfunction represents an important prognostic marker in the Chagas disease evolution. Nunes demonstrated in a longitudinal follow-up of 93 patients over three years that the restrictive filling pattern on Doppler, indicating diastolic dysfunction, was a significant cardiac mortality predictor in patients with chagasic dilated cardiomyopathy. A relevant aspect to be observed was the presence identification of regional diastolic dysfunction in Chagas disease. This concept, recently introduced with the tissue Doppler use, has demonstrated that patients with Chagas disease can present early heterogeneity in longitudinal segmental relaxation, particularly involving the interventricular septum and the left ventricle posteroinferior wall, demonstrated by Barros and collaborators.

Severe myocardial dysfunction also favors the intracavitary thrombi formation and thromboembolic phenomena, which constitute the third cause of death in Chagas disease. It is noteworthy the apical lesion great importance as a frequent thrombi site, with variable prevalence depending on the ventricular dysfunction degree, as described by Oliveira and collaborators and Nunes. Two-dimensional echocardiography is the procedure of choice for mural thrombus detection, being used in several studies on dilated cardiomyopathy as the “gold standard” for identification of the systemic emboli incidence and development, shown by Combellas and collaborators.

The ventricular function analysis using two-dimensional echocardiography, either in the assessment of motility and/or wall thickening, or in the volumes and ejection fraction determination, requiring the delineation of the muscle/cavity interface, involves subjective character and subject to inter observations- and intraobserver variations, and the experience required in the myocardial motility demands subjective quantification a long time of experience with the method. Therefore, it is important to use techniques that can quantitatively analyze cardiac dynamics, especially when this organ evaluation in longitudinal studies is required. Among these techniques, tissue Doppler represents a new method that allows quantitative and regional analysis of the heart systolic and diastolic function, and studies have shown its utility in the quantitative evaluation of segmental contractile abnormalities and early regional diastolic dysfunction in both ventricular cavities in Chagas disease, and may be a useful tool in these patients risk stratification, as shown by Barros and collaborators.

Myocardial scintigraphy

Several studies have demonstrated the myocardial (box) scintigraphy applicability in the organ-function involvement evaluation in Chagas’ disease.

Scintigraphy with thallium-201, associated with exercise testing, allows one to verify the transient ischemia zones existence (induced by effort and that disappear with rest) or definitive ischemia (compatible with the presence of necrosis and/or myocardial fibrosis). Apical radioisotopic hypocaptation can be demonstrated in Chagas disease carriers even without apparent heart disease, Thom and collaborators showed, allowing tip lesion early diagnosis. Apical radioisotopic uptake can also be observed in normal individuals, and should therefore be interpreted with caution.

Radioisotopic ventriculography provides reliable information on global left ventricular function and parietal motion abnormalities in patients with Chagas’ heart disease. Arreaza and collaborators found a good correlation between ejection fraction measured by cineventriculography and radioventriculography. When comparing the two techniques regarding the parietal contractility analysis, they verified agreement of the results in 77% of the cases, even greater when the analysis referred to the infero-apical region. Fifty-six percent of the patients with abnormal left ventricular wall motion had regional changes, and the involvement severity progressed from the asymptomatic patients group to those with heart failure. The changes in ejection fraction were in proportion to the number of parietal regions affected by the disease, being normal in asymptomatic patients and markedly depressed in patients with heart failure.

Chagas’ myocardiopathy presents diffuse contractility changes only in the disease advanced stages, with great dilatation and heart failure. In the less advanced forms, there is a segmental changes predominance in both ventricles. The segmental hypokinesia seems to relate to the pathological findings of ventricular wall thinning, with fibrosis and myocytolysis in systematized foci. The ventricular apex is the most frequently affected segment and the one that presents the most intense degrees of hypokinesia, which is also related to the anatomopathological data known in Chagas disease, as described by Kuschnir and collaborators.

The use of Tc-99m labeled 2-methoxy-isobutyl-isonitrile (Rp 30) allows simultaneous information of myocardial function and perfusion to be obtained. Patients with early Chagas’ disease clinical forms showed segmental abnormalities in Tc-99 isonitrile uptake as a marker of viable myocardium. These changes were more pronounced in patients with cardiomegaly, as shown by Castro and collaborators.

A significant Chagas disease patients number complain of atypical chest pain, sometimes resembling angina pectoris. Marin-Neto and collaborators performed a scintigraphic and cineagiocardiographic study in 23 Chagas disease patients who complained of precordial pain, in order to evaluate an ischemic cause possibility for this abnormality. They consider that myocardial ischemia, possibly microvascular in nature, may contribute to the genesis of the symptom. They believe that the definite uptake defects found in regions of the ventricular wall with pronounced changes in motion probably correspond to necrosis or fibrosis areas evident at patients necropsy in Chagas disease various stages. Also in his series, most of these perfusion defects involved the apical region, which is known to be the preferred site of aneurysmal lesions in Chagas disease.

In the causal analysis of segmental hypokinesia and ventricular aneurysms found in patients with positive epidemiology for Chagas disease, one should consider this etiologic possibility and include it in the differential diagnosis with coronary artery disease.

The radioisotopic techniques use acquires special relevance in patients with chronic pulmonary diseases and with thoracic conformation abnormalities, in whom echocardiographic study may be difficult or even impossible. Performing a comparative analysis between these two propaedeutic methods, Arreaza and collaborators consider that two-dimensional echocardiography and scintigraphic study complement each other, with the former seeming to allow apical lesion earlier detection, while the latter seems to provide more reliable and reproducible information regarding ventricular function. It is noteworthy that radioisotope scintigraphy allows the both ventricles morphofunctional study.

Nuclear medicine techniques can still be very useful in clarifying the disease pathophysiology. Simões and collaborators have studied the relations between thallium-201 myocardial perfusion and sympathetic innervation alterations, using I-123 meta-iodobenzylguanidine, and found a marked topographic association between perfusion and innervation defects and regional contractile abnormalities. Thus, the occurrence of early sympathetic innervation changes may causally relate to regional perfusion and contractility changes, both of which are precursors to global left ventricular dysfunction.

Thus, nuclear cardiology comprises easy-to-perform non-invasive propaedeutic methods that provide valuable information for the evaluation, diagnosis and treatment follow-up of patients with Chagas’ disease. Its judicious use, as with the other non-invasive techniques studied herein, must be considered in the selected cases complementary diagnosis in which more precise information on the heart morphology and function is sought, although it is not a substitute for the traditional cardiological diagnostic techniques.

Brain natriuretic peptide (BNP)

This cardiac hormone is a reliable indicator of left ventricular dysfunction, as described by McDonagh and collaborators. Recently, Ribeiro and collaborators demonstrated that this alteration also applies to patients with Chagas’ disease presenting with left ventricular dysfunction, revealing a high negative predictive value, and can therefore be used as a screening method, especially if there are alterations on ECG or chest X-ray. Therefore, with a normal BNP concentration, the probability of global left ventricular dysfunction is very low. In those with elevated BNP levels, on the other hand, echocardiographic investigation is needed to confirm the dysfunction.

Ergometric assessment

Ergometry is a widely used technique in cardiology, having been developed from the principle that cardiovascular system functional limitations, not demonstrable at rest, can be exposed by effort. It is a fundamental method, with well-defined diagnostic and prognostic value in coronary insufficiency, but also useful in the approach to patients with cardiomyopathies and valve diseases, allowing accurate determination of functional capacity and objective therapeutic response assessment. It is also applicable to the cardiac arrhythmias diagnosis and treatment control, to the apparently healthy individuals evaluation, and to the exercises prescription in rehabilitation programs, as described by Gibbons and collaborators.

Isotonic exercise is the preferred exercise for ergometric evaluation, since it determines a gradual increase in cardiac output, proportional to body oxygen consumption, as described by Chaitman, and the workload can be accurately measured. The physiological organic response to isotonic exercise involves cardiocirculatory adjustments that aim to maintain adequate blood flow to the skeletal muscles, without compromising cerebral and coronary perfusion. Immediately before exertion, reflex neurogenic mechanisms determine increased adrenergic tone and reduced parasympathetic tone. These changes become more marked as exercise is performed, determining an increase in cardiac output, resulting from the increase in systolic volume, an increase in heart rate, and a reduction in peripheral vascular resistance, shown by Ellestad and Duarte.

Ergometry is considered a safe technique when performed by experienced and well-trained physicians. The respect for contraindications and interruption criteria are fundamental for the patients’ safety, as well as the availability of a defibrillator and complete cardiopulmonary resuscitation material in the examination room. Among the absolute contraindications to the ergometric evaluation are: acute myocardial infarction (in the first two days), high-risk unstable angina, malignant cardiac arrhythmias causing symptoms or hemodynamic repercussion, symptomatic severe aortic stenosis, decompensated heart failure, thromboembolism or acute pulmonary infarction, acute myocarditis or pericarditis, acute aortic dissection. Relative contraindications are: left main coronary artery stenosis, moderate valve stenosis, hydro-electrolyte disturbance, severe arterial hypertension (systolic BP > 200 mmHg and/or diastolic BP > 110 mmHg), tachyarrhythmias or bradyarrhythmias, hypertrophic cardiomyopathy and other forms of flow obstruction, mental or physical alteration that prevents adequate exercise, high-grade atrioventricular block, as described in the studies by Detrano and Froelicher and Gibbons and collaborators.

The exercise stress test evaluates clinical, hemodynamic, and electrocardiographic parameters. Symptoms and signs that appear during the effort may have remarkable diagnostic value and should be valued and noted by the physician performing the test.

The physiological blood pressure response to effort implies a progressive rise in systolic blood pressure until a plateau is reached, while the diastolic pressure remains stable or fluctuates around 10 mmHg. Failure to raise the systolic blood pressure, as well as its reduction below resting levels during effort, may reflect inadequate cardiac output elevation due to left ventricular dysfunction, left ventricular outflow tract obstruction, or excessive reduction in peripheral vascular resistance.

The heart rate increases progressively with exercise. Hypovolemic, anemic, anxious, and unfit patients may have an exaggerated chronotropic response in the early phases of exercise, although the most important abnormality is chronotropic incompetence, i.e., inadequate heart rate elevation, less than 95% of the confidence interval limit for age and sex. Chronotropic incompetence may indicate sinus node dysfunction, left ventricular failure, myocardial ischemia, and the use of drugs with a negative chronotropic effect, as shown by Froelicher.

The double product, or stress-time index, is obtained by multiplying the maximum systolic blood pressure and heart rate reached during the test and provides a myocardial oxygen consumption estimate, and can be used a cardiovascular function index, as studied by Duarte and Froelicher.

The analysis of the electrocardiographic record obtained during stress has as fundamental parameters the behavior of the ST segment and T wave and the study of cardiac rhythm disturbances. Strain-induced ST-segment and T-wave abnormalities may indicate myocardial ischemia.

The functional capacity or maximal exertion capacity, despite suffering interference from environmental conditions and factors such as training, motivation, and the examinee familiarity with the test, is one of the most important procedure variables, having great prognostic value. Effort capacity is measured through body oxygen consumption (VO2), which reflects the oxygen amount that is removed from the air while performing exercise. The VO2 and carbon dioxide production (VCO2) determination during exertion is made by a specific procedure, called ergoespirometry, which evaluates the O2 amount consumed (resulting from the difference between the inspired O2 constant in the atmosphere and the expired CO2 amount) and that can be captured and analyzed by a sensitive apparatus, described by Alfieri and Duarte, which makes the procedure costly and complex. However, in the conventional exercise test, one can estimate the maximum oxygen consumption for a given effort load. The ergospirometry should be reserved for specific cases, such as in the differentiation of dyspnea induced by cardiogenic or pulmonary cause effort and in the objective evaluation of exercise capacity and therapeutic response in possible candidates for heart transplantation, as described by Ellestad, Froelicher, Detrano, and Gibbons.

Dynamic Electrocardiography

Dynamic electrocardiography, or simply Holter, was originally developed by Norman Holter and adopted as a propaedeutic method in the 1960s. Holter added the time dimension to electrocardiography, allowing the electrocardiographic recording to be made for prolonged periods and during the patients’ usual activities, with immediate impact on the arrhythmias and myocardial ischemia diagnosis. Since then, the method has improved, mainly through automation and miniaturization, and has been incorporated into clinical practice, with definition of both its applications and technical and operational limitations. New techniques have emerged with the introduction of the computers use in cardiology, such as the heart rate variability (HRV) study, which allows the cardiac autonomic control study, as well as different forms of recording and new analysis resources, for example, for pacemaker patients. Thus, the method is today a very important tool in the management with heart disease patients, as demonstrated by Crawford and collaborators.

Autonomic function assessment

The importance of the autonomic nervous system (ANS) involvement in cardiovascular diseases has been the subject of intense research in recent decades. The cardiovascular response fast and precise regulation to environmental changes and physiological stimuli, such as effort and emotion, is predominantly accomplished through the balance between vagal and sympathetic activity. Although these two ANS divisions are usually antagonistic, it is known that, in specific situations, they can act independently or even synergistically, making it difficult to specifically evaluate the contribution of the sympathetic and parasympathetic components.

The ANS control functional evaluation over the heart can be done through autonomic tests, in which the physiological reflex response to the application of a quantifiable physiological or pharmacological stimulus, such as breathing, exercise, and the atropine and phenylephrine injection, is observed. Alternatively, information about cardiac autonomic control can be obtained by observing the intrinsic heart rate variation, both in short recordings of two to five minutes at rest and in prolonged tracings of 24 hours during normal activities. Heart rate variability (HRV) analysis assumes that, under normal conditions, heart rate changes in response to various stimuli, such as exercise and mental stress, or even in resting conditions, fluctuating around an average. Such variability is predominantly related to continuous changes in the sympathetic-vagal balance, in response to cardiovascular control mechanisms. HRV can be studied by mathematical techniques that address the statistical characteristics of this variation (time domain), that decompose the different rhythms involved (frequency domain) or by non-linear methods, which use advanced mathematical methods to describe the heart rate variability behavior, as described by “Task force” and Lombardi.

Statistical methods provide simple, easily calculated indices that evaluate the heartbeat intervals dispersion around the mean (such as the SDNN, or standard deviation of normal heart intervals) or compare the adjacent cycles duration (such as the RMSSD, which is the absolute values average of successive differences, or the PNN50, the successive normal heart intervals percentage with a variation greater than 50 ms). While SDNN is the product of all autonomic (mainly parasympathetic) and neurohumoral influences on HRV, RMSSD and PNN50 are vagal influence direct results on the heart. In experimental models, withdrawal of vagal tone decreases the fibrillatory threshold and predisposes to sudden death. The prognostic value of reduced HRV time domain indices has been validated in several retrospective and prospective studies, especially after acute myocardial infarction and in heart failure.

Frequency domain analysis, through HRV spectral analysis, allows the study of the different autonomic nervous system divisions. In short duration recordings, it is recognized that high frequency variability (between 0.15 and 0.40 Hz) is related almost exclusively to vagus and respiratory sinus arrhythmia. The variability concentrated between 0.04 and 0.15 Hz, of low frequency, related to the baroreflex, has sympathetic and/or vagal origin, while the low / high frequency ratio would be an indicator of sympathetic-vagal balance. Despite the HRV spectral analysis theoretical advantages and pathophysiological potential, there are no clinical studies demonstrating its advantage over conventional time domain indexes.

Among the newer techniques, the most promising is the heart rate turbulence study, Bathel and collaborators and Ribeiro and collaborators describe the method that evaluates the changes in heart rate caused by ventricular extrasystoles. After an extrasystole, there is usually a compensatory pause and a subsequent forced contraction, activating the baroreflex and heart rate oscillations, a phenomenon known as heart rate turbulence. This physiological oscillation is reduced in a pathological conditions number, such as in Chagas disease and after myocardial infarction, a situation in which it has high prognostic value.

Echocardiogram

Echocardiography currently represents a valuable noninvasive diagnostic method whose application in cardiology is widely established. It is particularly useful in the infectious heart diseases study, by bringing the possibility to diagnose and monitor cardiac involvement, allowing better understanding of the disease natural history and pathophysiological mechanisms involved. The three modalities of the method – M Mode, Bidimensional and Doppler – complement each other, making it possible to evaluate anatomical, functional and heart pathology hemodynamic aspects. Echocardiography allows the cavity dilation identification, wall thickness, global and segmental contractility changes, tip lesion presence (in Chagas’ myocardiopathy) and intracavitary thrombi, in addition to estimating functional changes resulting from cardiac involvement.

The echocardiography ability to provide important information with minimal discomfort or risk, without the contrast or radiation use, associated with the method wide availability, makes its use so widespread. However, the indiscriminate use or screening in healthy patients is not justified for two reasons: the cost is not so low, and, as in any test, there may be false-positive results and changes without clinical significance, generating anxiety for the patient, additional investigation, and even unnecessary treatments, as described by Cheitlin and collaborators.

Myocardial scintigraphy

The different techniques applied to image acquisition and processing in nuclear cardiology are now well established, the progressive development result over the past four decades.

Radioisotopic ventriculography and radioisotopic angiography aim at the functional evaluation of the ventricular chambers by labeling circulating RBCs with a radioactive isotope (99mTc). Cardiac images are acquired synchronously with the electrocardiogram. The parameters analyzed are related to heart functional aspects, including global and regional wall motility, ventricular volumes, and the physiological changes that occur in the ventricular cavities throughout the cardiac cycle. The volumetric data allow accurate and highly reproducible calculation of the ventricular ejection fraction. Parameters of the fast and slow filling phase of the left ventricle can also be studied, with important implications for the diastolic function assessment. The role of ventriculography and radioisotopic angiography has been decreasing with the echocardiography techniques increase, but for selected patients, they still have their value, since echocardiography has limitations, such as the fact that the inferior wall evaluation of the left ventricle is difficult, patients with pneumopathies have their examination impaired due to poor image quality, and it is an examiner-dependent method, with interobserver variability of 11% in ejection fraction, as shown by Pennell and Prvulovich.

Myocardial perfusion scintigraphy with tomographic imaging (SPECT) has replaced planar imaging methods, facilitating neighboring regions separation, improving contrast resolution, and allowing better detection of differences in myocardial activity concentrations. In our environment, the main tracers available for myocardial imaging include thallium-201 and technetium-labeled tracers, among them mainly 99mTc-sestamibi and the 99mTc-tetrofosmin.

Myocardial perfusion scintigraphy with ECG-synchronized CT imaging (Gated-SPECT) allows the ventricular function analysis additionally and simultaneously to myocardial perfusion analysis, contributing to more accurate diagnosis and prognostic evaluation, and increasing the specificity of some perfusion study findings. In situations where there is doubt between a persistent perfusional defect and an artifact due to mammary or diaphragmatic attenuation, motility analysis and ventricular wall thickening can contribute to the differentiation of these two causes. When the hypoconcentration is due to an artifact, this wall motility is normal, as is the systolic thickening.

Positron emission tomography (PET) allows you to quantitatively study not only myocardial viability, but also regional perfusion. The most commonly used flow tracers are ammonia (N-13) and rubidium (Rb-82), but water, labeled with oxygen-15, can also be used.

In infectious myocardiopathies, the myocardial inflammation presence can be detected by radiotracers, captured in the myocardium. The most commonly used are 99mTc-pyrophosphate, gallium-67 citrate, and 99mTc ou 111In- antimyosin antibodies. When myocardial necrosis occurs, the myocyte membrane is lost, exposing the intracellular myosin heavy chains. The 111In-antimyosin, therefore, detects these necrosis areas and can be used in the diagnosis of acute myocardial infarction, myocarditis and post-transplant cardiac rejection, shown by Wackers and collaborators. Other tracers, such as thallium-201 and technetium-labeled tracers allow to check perfusion changes, while 123I-metaiodobenzylguanidine (123I-MIBG) assesses the myocardium sympathetic autonomic innervation. To check the functional status of the heart chambers, isotopic angiocardiography or ventriculography is used to measure ejection fraction, regional myocardial motility, and ventricular volumes.

Cardiac enzymes

Elevated cardiac enzymes reflect myocardial necrosis. Some studies suggest that troponins I and T are more sensitive than creatine phosphokinase MB fraction (CK-MB), both in myocarditis and in other occasions where there is myocardial necrosis, such as in acute myocardial infarction, described by Smith and Lauer. Usually the increase occurs in the first month, suggesting that most of the damage occurs early. Troponin elevation is not related to the histological myocarditis severity.

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Digestive form

The digestive form of Chagas disease: History, clinical picture and current situation

Joffre Marcondes de Rezende

Medical School of the Federal University of Goiás

Email: jmrezende@cultura.com.br

Since the Chagas disease discovery in 1909, it has been considered one of the possible choking sickness causes, an endemic disease in Brazil, whose symptomatology is the same as the esophageal achalasia universal occurrence. It is characterized by the esophageal loss peristalsis and lower sphincter lack of relaxation during swallowing, which causes difficulty in swallowing food, which is largely retained in the esophagus, causing this organ progressive dilation. The similarity of the two conditions has led many authors to believe that they are one and the same morbid entity. What distinguished them was the unusual frequency of choking sickness in certain regions of Brazil, in contrast to the achalasia rarity anywhere in the world. To have an idea of the choking sickness incidence in Brazil, it is enough to say that the casuistry reported in works published by only five authors, between 1939 and 1993, reached the sum of 7,435 cases, reviewed by Rezende in 1993.

Several hypotheses have been put forward to explain this fact, admitting that the environmental factors action, of toxic, infectious or food nature would be contributing to the disease endemic character. Thought was given to (i) the toxic action of a cassava consumed variety by the rural population, revealed by Paranhos’ studies in 1913; (ii) malaria and other infectious diseases, studied by Parisi in 1925; (iii) malnutrition in general or the vitamin B1 in the diet specific lack, studied by Etzel.

In the book Brazil and Brazilians, published in 1857, Kidder and Fletcher, two American missionaries, based on information from a physician, whose identity has only recently been revealed to be Dr. Joseph Cooper Reinhardt, who practiced in Limeira, in the São Paulo state, described the evil of choking as “a new disease”, which raged in the country interior, from “Limeira to Goiás”.

Arthur Neiva and Belisário Penna, in their celebrated scientific trip through the interior of Brazil in 1912, described in great detail the choking disease from a clinical and epidemiological point of view. They even cogitated an infectious cause for the endemic disease, as can be inferred from their experiment injecting blood from a sick man into a wild boar, but they did not relate it to Chagas disease.

The first reference documenting the chagasic etiology possibility of choking sickness is from Carlos Chagas himself in his work on the trypanosomiasis acute form, published in 1916. Suspecting an etiological relationship between the two endemics, he expressed himself thus:

is choking sickness an additional element of Brazilian trypanosomiasis and does this dysphagia of the acute forms translate to the syndrome initial phase?”. And, further on, he concluded. “…new research is needed to authorize, irrefutably, the inclusion of choking sickness in the Trypanosoma cruzi multiform symptomatology infection“.

It is noteworthy that Chagas referred to choking sickness as a syndrome and not as a disease.

For unexplained reasons, Chagas’ disease scholars at the time were either unaware of Chagas’ words or did not feel motivated to undertake the new research that was needed.

Years went by, and only in the 1930s did the first studies on the pathology of choking disease appear, carried out by researchers from São Paulo. Moacyr Amorim, Alípio Correa Netto, and Eduardo Etzel shed new light to clarify the problem. They demonstrated the existence, in choking sickness, of Auerbach’s myenteric plexus degenerative lesions, not only in the esophagus wall but also in the entire digestive tract. This finding allowed the unification into one entity of choking malady and colonic dilatation, commonly found in the same patient.

The colon dilation was called megacolon; by analogy, considering that the esophagus is also dilated, the denomination megaesophagus, proposed by von Hacker in 1907 for cases of idiopathic achalasia, was adopted for choking malady. Since then, the two conditions have been considered the same disorder manifestations.

The enteric nervous system degenerative lesions were attributed to the lack of vitamin B1 in the diet, according to the theory defended by Eduardo Etzel in 1935, a theory that was well accepted by the national and international scientific community, lasting until the 1950s.

On the other hand, choking sickness has always been an object of investigation by physicians working in the interior, both from a clinical and epidemiological point of view. It was a frequent observation that the same patient had megaesophagus and Chagas’ disease. Epidemiological data indicated that choking sickness was a rural endemic; the city dwellers affected by the disease, with very few exceptions, had lived on farms or in small villages.

In Goiânia, Joffre Rezende found that the disease was mainly found in low-income families living in precarious houses built of “pau-a-pique” with barreadas walls, where the hematophagous triatomines colonized, and that the cases largest number of megaesophagus seen in hospitals in the city came from municipalities in the state interior with a natural infection high rate triatomines by T. cruzi, according to a survey conducted by the National Department of Rural Endemics.

Everything indicated that, like the heart disease, the megaesophagus was a Chagas disease consequence.

Another argument supporting the chagasic etiology hypothesis of endemic megaesophagus lay in the high rate of Guerreiro and Machado’s complement fixation reaction positivity in patients with megaesophagus or megacolon, far above that found in any unselected group of the population from endemic areas. The first research in this sense was done by Eurico Vilela in 1930, in the city of Belo Horizonte. In 186 people investigated, 53 (28.5%) had a positive Guerreiro e Machado reaction, while in 13 patients with megaesophagus, eight were positive (61.5%). Also in Belo Horizonte, Melo Alvarenga performed the Guerreiro e Machado reaction in 16 megaesophagus cases, finding it positive in 8 (50%). The low percentages obtained were due to the low sensitivity of the antigen used. With its improvement, in the 40’s, Pedreira de Freitas obtained 91.2% in 80 megaesophagus and megacolon cases, and Laranja, Dias and Nóbrega, 97.0% in 81 cases. Several other authors have recorded a positivity high rate of the Guerreiro e Machado reaction in patients with megaesophagus and megacolon.

Despite all the clinical, epidemiological and serological evidence, there were still opponents to the chagasic etiology of megas. They were based on the following theoretical arguments of apparent logic, namely: (i) megaesophagus does not occur in all Chagas disease endemic regions, such as Venezuela and Central American countries; (ii) the chagasic patients vast majority do not present megaesophagus or megacolon; (iii) the high positivity of the serological reaction can be explained by the two endemic areas superposition, as occurred with Chagas disease and endemic goiter; (iv) no parasites were found in the esophagus or colon wall of autopsied cases; (v) megaesophagus is the same as the esophagus achalasia, found where there is no Chagas disease; (vi) megacolon may be due to other causes and also occurs in non-chagasic regions.

In the Medical Congresses of the “Triângulo Mineiro” and Central Brazil, held from 1947 on, the subject was much debated and the local doctors, based on their experience with the problem, always defended the chagasic etiology of megas against all these arguments.

The importance given in these Congresses to Chagas disease and to endemic megaesophagus can be appreciated in the article by Porto and Porto, entitled “História do Megaesôfago nos Congressos Médicos do Brasil Central”. In this article, the authors refer, in chronological sequence, to the most relevant contributions and end with the following words:

In these 13 Medical Congresses of the “Triângulo Mineiro” and Central Brazil held from 1947 to 1965 the megaesophagus theme was constant. This constancy had the appearance of insubordination. It seemed to be a protest attitude against the professors… who kept silent for a long time about a common evil widespread in Brazil and of secular knowledge even among healers and healers”.

Besides the theme megaesophagus, the ”Triângulo Mineiro” and Central Brazil Congresses also influenced the Chagas disease seriousness recognition as a public health problem and the awareness of the country’s health authorities on the urgent need to start a prophylaxis campaign for this endemic disease. In this sense, the participation of Emmanuel Dias and other illustrious chagologists, among them Amilcar Viana Martins, José Pelegrino, Humberto Ferreira, Pedreira de Freitas, was decisive.

For definitive chagasic etiology acceptance of megaesophagus and megacolon, however, anatomopathological proof was lacking.

Fritz Köberle was the one to prove it. Austrian by birth, Köberle came to Brazil in late 1953, hired to set up and run, for a period of three years, the Faculty of Medicine of Ribeirão Preto Department of Pathology at the University of São Paulo. At the end of his contract, he decided to stay in Brazil, became a naturalized Brazilian citizen, revalidated his medical degree, and took a competitive examination for full professor, becoming a full professor at the school until his retirement in 1976. From the very beginning he was interested in the choking sickness problem and its possible relation to Chagas disease. After reviewing the existing literature on the subject, he became convinced that trypanosomiasis was the megaesophagus and megacolon endemic in Brazil real cause. On the one hand, the convincing clinical, epidemiological, and etiological relationship serological data between the two endemics; on the other hand, the degenerative lesions anatomopathological findings of Auerbach’s myenteric plexus in autopsied megaesophagus cases, lesions found not only in the dilated segments, but throughout the digestive tract.

The myenteric plexus main function in the digestive tract wall is to coordinate motility in its different segments. It remained to be proven that those lesions were primitive and the dilatation secondary to motor changes resulting from the denervation produced by the T. cruzi infection.

Köberle developed his research on Chagas’ patients autopsies, on naturally infected animals, and on experimental infection in laboratory animals. In the infection acute phase there is the muscular wall parasitism of the digestive tract and an inflammatory process involving the myenteric plexus. In the chronic phase it was observed that the denervation is irregular, with variable distribution and intensity. He then had the idea of performing a quantitative esophageal neurons study in autopsies of non-chagasic individuals and in chagasic individuals with and without megaesophagus. In the neurons counting, which was done in the lower third of the esophagus, the reduction in the neurons number proved to be very variable. Denervation was a constant in Chagas patients, but less intense in those cases of apparently normal esophagus. He concluded, after comparing cases with and without megaesophagus, that the Chagas’ esophagopathy evolution into a typical megaesophagus only occurs when denervation reaches a threshold, which was estimated at 90%.

Quantitative studies have also been done in relation to the colon, which have shown that denervation is not restricted to the dilated segment, commonly rectum and sigmoid, as was believed, but extends to the entire colon. The critical level of reduction in the neurons number for the megacolon onset has been estimated at 55%.

Next, his collaborators continued the quantitative studies in relation to other digestive tract segments: stomach, duodenum, small intestine, cecal appendix, and colon, demonstrating in all of them a significant reduction in the neurons number. When studying chronic Chagas’ disease, Köberle also found a reduction in the nerve cells number in the heart.

From the works published series by Köberle on the digestive tract pathology in Chagas disease, the Preliminary Note in collaboration with Stephen Nador and those of greater relevance that followed between 1955 and 1968 should be mentioned as a reference.

Initially, Köberle had admitted that neuronal destruction resulted from a neurotoxin released by the parasites after amastigote pseudocysts rupture; he later recognized it to be an immunological mechanism.

In view of the findings that configured an entirely new aspect in the T. cruzi infection pathology, Köberle established a Chagas disease new view, conceptualizing it as a disease of the peripheral autonomic nervous system.

This concept has proven to be very fertile and has led to numerous research studies that have shown multiple disorders in different body sectors in Chagas disease patients.

Later, other authors not only confirmed Köberle’s studies, but also verified the denervation existence in other organs.

Faced with the chagasic etiology proof of endemic megaesophagus and megacolon and of the intrinsic denervation in the entire digestive tract, it was necessary to review the Chagas disease clinical forms classification, considering the large number existence of megaesophagus and megacolon cases without heart disease, previously included in the undetermined form and considered only as potential cardiac.

Joffre Rezende, in 1956, proposed the digestive form denomination to characterize the manifestations resulting from “lesions of the digestive tract with the consequent alterations of motility, a denomination that was well accepted by the scientific community. In 1959, Joffre Rezende expanded the digestive form concept to encompass, in addition to motor disorders, the digestive tract secretory and absorptive alterations, already known or that would be described in the future, regardless of the esophagus or colon dilatation presence or absence.

Many Chagas disease pathology new aspects and its manifestations in the digestive tract have been disclosed by the Revista Goiana de Medicina. In the 35-year period, (1955 to 1990), among 564 titles of original articles published, 201 (35.6%) refer to Chagas disease, and of these, 85 (42.3%) concern the digestive form in its different aspects, from pathology to the megaesophagus and megacolon surgical treatment.

As a result of the knowledge acquired, the Chagas disease classification into the following clinical forms is currently accepted: (i) indeterminate, (ii) cardiac, (iii) digestive, and (iv) mixed or associated (cardiac + digestive). The digestive form has been reported with a widely varying prevalence depending on the geographical region. In Brazil, the existing medical literature records a cases higher number in the central region of the country, comprising part of Minas Gerais, Goiás, São Paulo, Bahia, and southern Piauí states. Its occurrence is exceptional in countries situated above the equatorial line, such as Venezuela and the Central American countries where Chagas disease is well studied and recognized as a heart disease cause.

In Brazil, the digestive form prevalence has been estimated based on the esophagopathy diagnosis in radiological surveys done in chagasic populations in endemic areas, by means of 35 and 70 mm abbreviation. In seven of these surveys, totaling 2,073 cases, the prevalence ranged from 7.1 to 18.8, with a mean of 8.8, as described by Rezende and Luquetti.

Chagas’ esophagopathy has been found to be a digestive form good indicator, given that many of the changes found in other digestive system parts occur in association with the megaesophagus.

At radiological exam, depending on the disease evolution, the esophagus can present different morphological aspects that were classified into four groups by Joffre Rezende and collaborators. The first two groups comprise the megaesophagus compensated phase, in which there is esophageal muscle wall greater contractile activity, while the last two correspond to the decompensated phase in which motor activity is minimal or nonexistent.

The motor changes recorded on manometry are very variable in the compensated phase and follow a uniform pattern in the decompensated phase, as demonstrated by Henrique Pinotti, Renato Godoy, and Joffre Rezende.

In the digestive form, besides the megaesophagus and megacolon, motor and/or functional alterations in the stomach, duodenum, jejuno-ilus, extrahepatic biliary tracts, salivary glands, and pancreas have been described by several authors, according to the literature review recently done by Joffre Rezende and Helio Moreira.

The existence of a Chagas’ gastropathy had previously been suspected by Calil Porto based solely on clinical observation. Gastric changes are found in about 20% of patients presenting with megaesophagus. On radiological examination, the gastric volume is extremely variable and an air chamber absence in the stomach is characteristic in patients with advanced megaesophagus. Hypersensitivity of the gastric wall muscles to cholinergic pharmacological stimulation, as well as motility and secretion disturbances can be detected by different methods in patients with the digestive form. In such cases, gastric emptying is accelerated for liquids and delayed for solids, and there is gastric body less adaptive relaxation to stomach distension. Recently, a study by Rezende Filho, using electrogastrography, demonstrated a stomach altered electrical rhythm, with gastric dysrhythmia.

Gastric secretion studies revealed chlorhydropeptic hyposecretion, both basal and under different stimuli: histamine, histalog, insulin, pentagastrin, and calcium ion infusion. When stimulation with a cholinergic substance, such as methacholine or betanechol, is associated with one of these tests, an increase in the secretion of both hydrochloric acid and pepsin is obtained, which shows that the hyposecretion is mainly determined by the stomach intrinsic denervation and not by a reduction in the secretory cells number. In parallel, fasting and postprandial hypergastrinemia are noted.

Besides these motor and secretory alterations, the chronic gastritis presence in different degrees of intensity is frequent, whose etiopathogenic factors seem to be multiple, possibly including duodenogastric biliary reflux and Helicobacter pylori infection.

In cases with marked difficulty in gastric emptying, previously labeled “pylorus achalasia,” pyloric muscle hypertrophy is found. In these cases, pyloroplasty is indicated as a complement to cardiomyotomy in the megaesophagus surgical treatment.

After the esophagus and the colon, the duodenum is the segment that is most often dilated. Megaduodenum is almost always associated with other visceromegalies. The dilatation can be located only in the bulb (megabulb), in the second and third portions, or it can involve the entire duodenal arch. Even in cases where there is no dilatation, dyskinesia and hyperreactivity to cholinergic stimulation due to denervation are common. The symptoms eventually caused by megaduodene may be confused with dyspepsia of gastric origin of the dysmotility type.

In the small intestine, histopathological studies showed less marked denervation of the enteric nervous system than in the esophagus and colon. Jejunum or ileum dilatation to the point of forming megajejunum and megaioleum is a rare occurrence, with only a few cases reported.

The repercussions of Chagas enteropathy are not very evident from a clinical point of view, but can be detected in targeted investigations. Motor changes have been proven in both radiological and manometric studies. The interdigestive migratory motor complex shows abnormalities in patients with other digestive form manifestations. Possibly as a result of this fact there is an increased bacterial flora growth, which resembles, in certain cases, that found in stagnant loop syndrome.

Studies on intestinal absorption in patients with the digestive form have revealed an accelerated glucose and other sugars absorption. As a result, the oral glucose tolerance test may show abnormal glucose curves, with transient hyperglycemia in the first hour. In addition to the hyperabsorption of glucose, there was mild lipid hypoabsorption, which does not alter the fecal fat excretion. Both of the changes described are attributed, in part, to gastric emptying disturbances.

Colonic dilatation most often occurs in the distal segment, comprising the rectum and sigmoid colon.

The gallbladder also suffers from intrinsic denervation in the digestive form of Chagas disease, with filling and emptying motor alterations. Abnormalities were also recorded by manometry on the Oddi`s sphincter. Cholecystomegaly, however, is infrequent, as is choledochal dilatation. There are data in the literature that suggest a higher incidence of cholelithiasis in Chagas disease patients with megaesophagus and/or megacolon.

The salivary glands, notably the parotid glands, are hypertrophied in patients with megaesophagus, which is common in any obstructive esophagopathy as a esophagosalivary reflex consequence, which produces hypersalivation. In Chagas patients, however, there is greater sensitivity of the salivary glands to the mechanical stimulation of chewing and to pharmacological stimulation by pilocarpine. Moreover, parotids hypersalivation and hypertrophy persist in esophagectomized patients, which demonstrates that it is not only the esophagossalivary complex but the impairment of these glands innervation in Chagas disease.In relation to the exocrine pancreas, its functional capacity is preserved in relation to the direct stimulus on the gland. There may, however, be a secretory deficiency due to indirect stimulation, resulting from alterations in the duodenojejunal hormones release.

Figure 1 – Distribution of 150 cases of megaesophagus in the Goiás state in 1956. Its occurrence geographical area coincided with the region most infested by T. cruzi infected triatomines.
Figure 2 – Neurons number in a 1 mm segment of the esophagus lower third in normal people, in chagasic patients without megaesophagus, and in chagasic patients with megaesophagus, (Köberle, 1961).
Figure 3 – Histological section of the esophagus muscular wall in a Chagas mega esophagus case, showing inflammatory process in the myenteric plexus region and neuronal depopulation. A single degenerating neuron is seen (arrow).
Figure 4 – Chagas disease digestive form Pathophysiology.
Figure 5 – Megaesophagus radiological classification into four groups, according to the evolution of the condition.
Figure 6 – A – Megaesophagus and mega-stomach. B – Megaesophagus and megaduoden. C – Megabulb and megajejunum D – Megacolon.

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Nervous form

Andréa Alice da Silva

Pathology Department, Medical School, Fluminense Federal University

Email: aasilva@id.uff.br

Edition:

Joseli Lannes

Laboratory of Interactions Biology, Oswaldo Cruz Institute/Fiocruz

Email: lannes@ioc.fiocruz.br, joselilannes@gmail.com

In the last two decades the central nervous system (CNS) has come to be considered an immunospecialized site protected from inflammation and infection by the blood-brain barrier (Ransohoff & Brown, 2012; Obermeier et al., 2016, Manglani & McGavern, 2018), as it has the ability to respond to infection in a “quieter” manner than most tissues. Frequently, the immune and inflammatory response, with interaction between immune cells and CNS resident cells, participate in the infectious agents control that often result in tissue damage. However, the CNS can harbor infectious agents in their latent form, such as the intracellular protozoan Toxoplasma gondii and various viruses in an apparently silent manner. Since the discovery of American trypanosomiasis in 1909, later named Chagas disease, it has been known that Trypanosoma cruzi affects the CNS (Chagas, 1909). At that time, Carlos Chagas, Gaspar Vianna and Margarino Torres, among others, demonstrated the T. cruzi presence in CNS cells, mainly in the disease acute phase. It was proposed that, in an attempt to control T. cruzi, mononuclear cells would be recruited to the extravascular space, contributing to the acute meningoencephalitis formation. Interestingly, Carlos Chagas himself defined the acute malignant form, referring to the focal inflammatory reactions presence in the CNS that would lead the individual to a morbid condition. According to a report by Gaspar Vianna (1911), the histopathological alterations present in the CNS of a 3-month-old child who died of the “schizotripane acute form” were inflammation multiple foci with varied distribution throughout the brain tissue, and the parasite presence was observed in several glial cells. Corroborating these data, Deolindo Couto et al. (1964) reviewed the nervous manifestations studies concluding that in the Chagas disease acute phase the involvement of nervous tissue and leptomeninges is “indubitable”. Aluizio Prata (2001), in his turn, called attention to premature newborns cases, resulting from vertical transmission, who developed neurological symptoms. In addition, the researcher reports that 5-10% of children in the acute untreated phase die at this stage due to severe heart failure or severe encephalomyelitis.

The use of various experimental models with different strains and inocula of T. cruzi and animals species (mice, rats, dogs, monkeys) has allowed advancement in the neurological changes study during T. cruzi infection. The high pathogenicity of some parasite strains, especially those from armadillo led to the neurotropism hypothesis described by Villela (1925). However, Amaral et al. (1975) using albino mice infected with T. cruzi different strains suggest that the difference in the parasitism intensity is also due to the host characteristics and not exclusively to the T. cruzi strain tropism. Thus, some hosts are particularly susceptible, while others have shown some refractoriness to infection or immunopathological changes in the CNS, these characteristics being influenced by (i) age, (ii) genetic factors, n nutritional conditions, (iv) individual immune status, among other factors (Kolodny 1939, Culberston et al., 1942, Hauschka et al., 1950, Roggero et al., 2002, Andrade et al., 2002, Carvalho et al., 2003, Marinho et al., 2004).

Similar to what was described by Gaspar Vianna (1911), the most studied murine models infected with parasite different strains present encephalitis in the acute phase, characterized mainly by the presence of (i) cerebellar edema, (ii)inflammatory infiltrates, and (iii) T. cruzi amastigotes forms, as reported by Alencar and Elejalde (1960) and reviewed by Pittella (1991). The parasite presence is found in the brain and spinal cord, as well as meningoencephalomyelitis lesions characteristic, with perivascular inflammatory foci rich in macrophages and scattered throughout the white and gray matter, but with no preferential area (Villela & Torres, 1926, Queiroz, 1975).Seeking to understand the CNS alterations pathophysiology during T. cruzi infection, we have been developing studies with several mice and parasite strains. In our studies characterizing the immunopathological changes developed in C3H/He mice infected with the Colombian strain, we observed that meningoencephalitis is related to the parasitemia peak, and is constituted by the CD8+ T cells, and macrophages predominance surrounded by a fine network of one of the main extracellular matrix components, fibronectin (Silva et al., 1999). Considering the three-dimensionality of the nervous tissue, inflammatory infiltrates are not associated with the T. cruzi antigens presence. Seeking to understand the molecular mechanisms involved in the inflammatory cells entry into the CNS and Chagas encephalitis formation, we have shown that cell adhesion molecules, mainly VCAM-1 and its receptor the integrin VLA-4, play an essential role in susceptibility to meningoencephalitis (Roffê et al., 2003). The inflammatory cells present in the peripheral blood of acute-phase animals express receptors for CC-chemokines, such as the CCR5 molecule, one of its ligands being CCL5/RANTES expressed in the CNS during T. cruzi infection. However, unlike what we previously showed for Chagas’ myocarditis (Marino et al., 2004), the inflammatory cells migration to the CNS is independent of CCR5 (Silva et al., 2007). However, CC-chemokines present in the CNS may be playing a role in the inflammatory cells activation and not necessarily in their entry into the injury site. Roffê et al. (2007) showed that in mice experimental infection with T. cruzi strain Y there is an increase in peripheral blood of endothelin (ET), a potent vasoconstrictor produced by endothelial cells that can act as a pro-inflammatory factor. To verify ET-1 participation in the CNS, Rachid et al.(2010) used ET-1 receptor antagonists and proposed that ETA and ETB contribute to parasite control in the infection early phase after entry into the CNS, but do not act on parasite replication in a late phase. Further, they noted that ET receptor antagonists fail to reduce neuroinflammation, including with regard to CCR5 and interferon gamma (IFN-g) expression.

Figure 1: Central nervous system sections of mice infected with Trypanosoma cruzi: (a) various parasite amastigotes forms – green, arrow; (b) astrocyte characterized by the GFAP filaments presence (red); (c) astrocytes infected by T. cruzi, (d) cells expressing MHC-II are infected by T. cruzi; (e) macrophages (F4/80+ cells) are infected by T. cruzi. 1000X magnification; Immunofluorescence; Authors: Andrea Alice da Silva and Joseli Lannes, LBI/IOC, Fiocruz.

Regarding nerve cells, both in human infection and in experimental infection in dogs, cats and mice, neurons are rarely infected by T. cruzi (Pires, 1978, Pittella et al 1990, 1991). However, Chagas (1934) and Queiroz (1975) showed that pyramidal cells are targets for infection by T. cruzi. Also, parasite amastigote forms are observed at inflammatory sites or within macrophages, glial cells (astrocytes and microgytes) and endothelial cells in the infection acute phase, as first described by Chagas (1934) and confirmed by other authors (Queiroz, 1975, Pittella, 1990, Da Mata et al., 2000, Guarner et al., 2001, Silva, 2006, Morocoima et al., 2012). Corroborating the above data, Vargas-Zambrano et al. (2013) showed that the human CRL-1718 strains astrocytes when infected in vitro with T. cruzi have increased MHC-I expression density and the frequency of cells expressing MHC-II molecules and producing the chemokines CCL2 and CXCL8, suggesting that infected astrocytes may regulate the immune response within brain tissue. Further, they suggest that astrocytes infection occurs in a low-density lipoprotein (rLDL) receptor-independent manner. Recently, in an acute and chronic mouse model of infection we showed that astrocytes are the main cells infected by T. cruzi in the CNS. Also, showing that primary cultured murine and human astrocytes of the U-87MG strain are infected in vitro by T. cruzi, allowing the trypomastigote-amastigote-tripomastigote cycle development, which is, seemingly surprisingly, favored by the presence of inflammatory cytokines, tumor necrosis factor (TNF) and IFN- (Silva et al., 2015, Silva et al., 2017), as best described below.The T. cruzi presence in the cerebrospinal fluid has been demonstrated in immunocompetent individuals (11/08) in the acute phase, without neurological changes (Kohl et al., 1982). This study further suggests cerebrospinal fluid as a possible T. cruzi route entry into the CNS. To date, few studies have focused on clarifying the parasite’s route of entry and even the molecules involved in the glial cells infection.

Figure 2: Sections of central nervous system from Trypanosoma cruzi infected mice, showing the presence of Mac-1+ cells (leukocytes); F4/80+ cells (macrophages) and CD4+ and CD8+ T cells in meninge areas and leptomeninge (left column) and brain parenchyma. Magnification 100X; Immunohistochemistry; Authors: Andrea Alice da Silva and Joseli Lannes, LBI/IOC, Fiocruz.

We cannot fail to report the studies demonstrating the presence of antibodies recognizing CNS components such as neuron/glia, myelin and myelin basic protein (Khoury et al., 1979, Ribeiro-dos-Santos et al., 1979, Spinella et al., 1992 and Al-Sabbagh et al., 1998). We have also shown high circulating concentration of anti-myelin basic protein antibodies and its encephalitogenic fragment formed by amino acids 4-14 (Yasuda, 1991), in parallel with the CNS lesion presence in the acute phase. In the chronic phase, in the inflammatory nervous form absence, we observed the presence of the anti-myelin basic protein antibodies, but not the anti-fragment 4-14 antibody, as reviewed by Silva et al. (2010). Thus, despite the acute chagasic meningoencephalitis intense studies, biological significance clarification of the antibodies presence that recognize nervous tissue is still needed.

The chronic nervous form is still debatable today, despite the recognition of this form by Chagas (1913) as reported in the classic article by Köberle (1967). The studies showed (i) cortical atrophy with or without hydrocephalus, (ii) decreased neuronal population and (iii) glial cells activation such as astrocytes and microgytes, but no histopathological alterations or even parasitism could be evidenced (reviewed by Alencar, 1982). Unlike other models, our study with C3H/He mice infected with the T. cruzi Colombian strains showed presence, although scarce, of the parasite in the chronic phase, confirming what was proposed by Pittella et al. (1990). However, the inflammation absence suggests the inflammatory processes resolution found in the CNS during the experimental infection acute phase, since all animals survived the acute infection. Intriguingly, the mice presented locomotor disturbances that led to partial paralysis of the hind limbs, mainly during the chronic phase, despite the fact that they did not present immunopathological alterations in the CNS (Silva et al., 1999). Interestingly, motor and/or sensory disturbances previously described (Vianna, 1911) were also observed in a study of 46 chronic Chagas disease carriers (Mangone et al., 1994). These patients showed cognitive deficits characterized by reduced (i) information processing speed, (ii) problem-solving capacity and (iii) learning, when compared to non-infected individuals. Also, it was shown that Chagas disease chronic carriers present electroencephalographic (20%) and cognitive (11.4%) alterations in a manner not associated with cardiac alterations, suggesting the brain impairment existence in the Chagas infection chronic phase (Prost et al., 2000). Studies in mice infected by T. cruzi have shown that they present sleep disorders and memory deficit, even in the histopathological alterations absence in the CNS (Arankowsky-Sandoval et al., 2001). Indeed, we have shown that mice of strains with differentiated susceptibility degrees to acute meningoencephalitis C3H/He (susceptible) and C57/BL6 (resistant), when infected with the T. cruzi Colombian strain, show cognitive and behavioral changes in the acute phase, which persisted in the infection chronic phase, such as depression and anxiety (Silva et al, 2010; Vilar-Pereira et al., 2012; Vilar-Pereira et al., 2015), therefore, in a manner independent of histopathological changes. Also, considering the biological and T. cruzi molecular diversity (Zingales, 2018), we show that depression is induced by infection with the Colombian strain (Type I), but not by the Y strain (Type II) of the parasite. Also, this behavioral alteration was blocked by antidepressant (fluoxetine) or antiparasitic (benznidazole) therapy (Vilar-Pereira et al., 2012).

It is known that cardiovascular diseases favor the neurocognitive changes onset (Elderon & Whooley, 2013). However, this does not seem to be the case in chronic Chagas disease carriers, since even part of the carriers of the indeterminate/asymptomatic form present cognitive and behavioral changes (Prost et al., 2000; Lima-costa et al., 2009). We are evaluating whether this is also the case in experimental CD infection, in order to outline mechanistic proposals that explain these alterations existence in chronic Chagas disease. In this context, knowing that the chronic Chagas disease severity is associated with a systemic inflammatory process enriched in TNF, IFN- and other cytokines (Pérez-Fuentes et al., 2003, Perez et al., 2011), we treated chronically infected animals with TNF cytokine blocking antibody (anti-TNF) and TNFR1 modulator (pentoxifylline). These therapies were able to reduce immobility (forced swim and tail suspension tests) of infected animals compared to their respective uninfected and vehicle-treated controls (Vilar-Pereira et al., 2012). This dataset suggests that behavioral changes, such as depression, present in experimental T. cruzi infection may be due to the parasite direct influence or due to the systemic production of pro-inflammatory cytokines, such as TNF, however suggesting that it is not due to this cytokine in situ production in the CNS (Vilar-Pereira et al., 2012), which needs to be further explored.

The CNS study in chagasic infection has gained greater interest in immunocompromised individuals such as patients with neoplasms or undergoing transplants, as well as immunocompromised co-infected individuals who develop acquired immunodeficiency syndrome (AIDS), as these develop severe immunopathological alterations in this tissue (Pacheco et al., 1998, Ramos Jr., 2004, Almeida et al., 2009, Fernandes et al., 2017). Suggested by Ramos Jr. (2004), the reactivation of Chagas’ disease became a clinical event of epidemiological interest as an AIDS condition indicative in America, supported in 2005 by the WHO in a Technical Report (reviewed by Almeida et al., 2011). In this context, the main neurological changes in individuals with chagasic meningoencephalitis reactivation are (i) intracranial hypertension, (ii) meningitis, (iii) paresis, (iv) plegia, (v) consciousness progressive loss, (vi) headaches, and (vii) seizures. In fact, the reactivation manifestation is more frequent when the HIV+ individual has a count ≤ 200 CD4+ lymphocytes/mm3 of circulating blood, and the exact mechanism of CNS infection reactivation during AIDS is not yet clarified. Histopathological changes, found mainly in areas of white matter but absent in the basal core, are necrotic or tumor-like lesions, with numerous parasite amastigotes forms found in glial cells or macrophages located in the meninges perivascular areas. In these cases, the parasite can be detected in the blood and especially in the cerebrospinal fluid (Jardim & Takayanagui 1994), but the significance of this finding has not been clarified to date. We cannot fail to mention that 44% of individuals develop intense myocarditis during reactivation, but meningoencephalitis is the most frequent (75-80% in HIV co-infection and 20% of infected patients who undergo kidney transplantation) (Ferreira, 1997, Prata, 2001). In these cases, the parasite participation is supported by the findings showing that trypanosomicidal treatment with benznidazole reverses the meningoencephalitis picture (Antunes et al., 2000, Pinazo et al., 2013). In an reactivation experimental mode of chagasic infection established in the chronic phase with various immunosuppressants (azathioprine, betamethasone and cyclosporine), Andrade and et al. (1997) showed that these animals showed inflammatory infiltrates with necrosis sites in the CNS. However, parasite nests and/or antigens were not detected in the nervous tissue, although they were found in the heart tissue. The chagasic infection reactivation in chronic immunosuppressed patients, in addition to the high presence of circulating parasites and necrotic and hemorrhagic lesion, shows markedly parasitized glia cells and macrophages (Rocha et al., 1994). Corroborating these findings, we have shown that C3H/He mice infected and subjected to immunosuppressive treatment in the chronic phase exhibit intense inflammatory infiltrates and elevated CNS parasitism associated with re-expression of the vascular adhesion molecule VCAM-1 on blood vessels endothelial cells present in the CNS (Roffê et al., 2003). Recently, it has been shown that the human astrocyte when infected in vitro can concomitantly harbor HIV and T. cruzi, and in this condition the parasite is able to inhibit viral replication without interfering with HIV infectivity. In addition, T. cruzi infection reduces astrocytes apoptosis, via a protective effect of IL-6 cytokine secretion by these coinfected cells (Urquiza et al., 2017). However, many questions still remain unclear, such as what are the reactivation mechanisms in the CNS? would it be via the parasite already installed silently, since the parasite is detected via immunohistochemistry in scarce brain sections (Roffê et al., 2003), or even within astrocytes in vitro (Silva et al., 2015)? Or would it be via a reinfection and new access to the CNS by the parasite? Which molecules would be involved in the parasite entry into glial cells and even in the parasite control in the chronic phase? What would be the role of systemic inflammation and that produced at the brain site?

Seeking to understand the cytokines role in acute chagasic encephalitis reactivation, we used IL-12 (IL-12-/-) or IFN (IFN-/-) deficient mice and their C57BL/6 controls infected with the Colombian strain. After infection, the mice were treated with benznidazole, and upon stopping this treatment, we did not observe inflammation in the C57BL/6 mice heart or CNS. The IL-12-/- animals showed drastic inflammation, accompanied by intense parasitism in the CNS compared to the mild inflammation and parasitism developed by the IFN-/- animals. The differences in reactivation at the CNS site found between IL-12-/- and IFN-/- animals were attributed especially to the IFN presence, since IFN-producing cells are present in the IL-12-/- mice spleen. Thus, the level of IFN deficiency would be the determining factor of the T. cruzi infection reactivation site in immunocompromised hosts (Michailowsky et al., 2001). Recently, we showed that in vivo T. cruzi infected GFAP+ cells (astrocytes) are in close proximity to IFN-+ cells and to Iba-1+ and CD3neg cells suggesting interactions between IFN- producing microglia with T. cruzi infected astrocytes. Importantly, the IFN- presence in the in vitro astrocytes infection favors the T. cruzi proliferation and release, which completes its cycle in these cells, and the TNF and nitric oxide production (Silva et al., 2015). In this context, the TNF presence favors parasite colonization and egression via increased TNF receptor 1 (TNFR1) expression and also the TNF and IL-6 production by infected astrocytes in vitro (Silva et al., 2017). It is likely that T. cruziinfection in the TNF presence, with inflammatory environment maintenance induces the parasite replication perpetuation in glial cells, such as astrocytes, thus favoring behavioral changes in T. cruzi infection (Vilar-Pereira et al., 2012). Corroborating these data, we show that treatment with pentoxifylline, a TNFR1 expression modulator, inhibits astrocyte susceptibility to T. cruzi infection in vitro, while in vivo treatment with anti-TNF antibodies (infliximab) reduces the amastigote nests number within the CNS. Although it is not possible to distinguish the influence of systemic TNF and TNF produced in situ, we propose that this inflammatory circuit induced by the interaction of glial cells with the parasite explains, at least in part, the cognitive changes, such as depression and anxiety, described in the T. cruzi infection chronic phase (Vilar-Pereira et al., 2012, Vilar-Pereira et al., 2015). This proposal is based on the increasing confirmation of the neuroinflammation importance leading to neurodegenerative disorders (Ransohoff et al., 2016). Also considering age, metabolic changes and infections that favor neuronal damage and death (Klein et al., 2017; Kabba et al., 2018), also microglial activation can lead to the secretion of neurotoxic or neuroprotective molecules, influencing the inflammatory process and consequently neuronal death, leading to neurodegenerative processes, such as the depression described in Chagas disease and its experimental models.

In view of the above, today we can confirm that in the Chagas disease acute phase there is parasite intense presence in the brain tissue accompanied by neuroinflammation, but in the chronic phase, regression of this inflammation and parasite load reduction is suggested, even though it persists mainly in astrocytes. Thus, the nervous form of neurodegenerative rather than neuroinflammatory aspect is installed, whose molecular mechanisms need to be unraveled. However, the indication that specific therapeutic (antidepressants) and antiparasitic interventions can reverse these processes, already opens perspectives for a better quality of life of the Chagas disease patient affected by behavioral alterations such as anxiety and depression.

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