Differential Regulation of Granuloma Size and Hepatic Fibrosis in Schistosome Infections

Granuloma size is the variable most frequently used to evaluate the immunopathogenesis of schistosome infections. However, hepatic fibrosis is at the least an equally relevant variable. Hepatic fibrosis and the size of circumoval granulomas are frequently dissociated in experimental murine Schistosoma mansoni and S. japonicum infections. Virtually nothing is known of the immunoregulation of schistosomal hepatic fibrosis. This review notes many of the studies which have found discrepancies in granuloma volume and hepatic fibrosis, attempts to put them in perspective and to evaluate different methods of calculating changes in collagen synthesis or content

An Experimental Approach to the Pathogenesis of "Pipestem" Fibrosis (Symmers' Fibrosis of the Liver)

Pathogenesis of schistosomal hepatic fibrosis ("pipestem" fibrosis of the liver) was investigated by means of the murine model. Although worm load appears as the main pathogenetic factor, alone it is not sufficient to produce that characteristic lesion. By comparing the findings in animals with heavy and prolonged Schistosoma mansoni infection, which developed or not" pipestem" fibrosis, it was observed that the lesion was more frequent in intact animals than in the splenectomized one. However, the size of the spleen, the number of recovered worms, the number of eggs per gram of liver tissue, the level of serum idiotype and anti-idiotype antibodies, the size and volume of periovular granulomas formed in the liver, all that failed to show statistically significant differences between the two groups. After analysing all these data, other factors, that apparently have been hitherto negleted, rested to explain the findings. Among them, the timing and sequence of the egg-induced intrahepatic vascular changes seemed crucial. The sequential development of intrahepatic portal vein obstruction, followed by the opening of periportal collateral veins and the continous arrival of schistosome eggs going to be lodged into the latter, appeared as essential steps in the pathogenesis of "pipestem" fibrosis

Role of cytokines in the formation and downregulation of hepatic circumoval granulomas and hepatic fibrosis in Schistosoma mansoni-infected mice

Schistosoma mansoni infections are associated with a strong Th2 cytokine response. Treatment of mice with IL-12 or anti-IL-2 or anti-IL-4 before i.v. injection of eggs increased IFN-gamma production and downregulated Th2 responses and pulmonary granuloma size. Conversely, anti-IFN-gamma antibody treatment increased Th2 responses and granuloma size. Similar manipulation produced less dramatic results in infected mice. However, sensitization of mice with eggs + IL-12 before infection augmented the Th1 response and decreased Th2 cytokines, granuloma size and fibrosis. Antisera to IFN-gamma, TNF-alpha or IL-12 during IL-12-egg immunization partly restored granuloma size and fibrosis following infection. Variations in the size of granulomas in acute (8 week) infections may be influenced primarily by the number and state of activation of T cells. In chronic (12-16 week) infections immunologic downmodulation proceeded normally in mice without functional CD8+ cells and in IFN-gamma KO mice but not in B cell KO (muMT) mice or in mice deficient in FcR expression in spite of the fact that these mice downregulated their T cell and cytokine responses. It is evident that the participation of cytokines in granuloma formation and regulation is complicated and that the mechanisms controlling both these phenomena are likely to involve both T cells and antibody/FcR interactions.

Myofibroblasts in Schistosomal Portal Fibrosis of Man

Myofibroblasts, cells with intermediate features between smooth muscle cells and fibroblasts, have been described as an important cellular component of schistosomal portal fibrosis. The origin, distribution and fate of myofibroblasts were investigated by means of light, fluorescent, immunoenzymatic and ultrastructural techniques in wedge liver biopsies from 68 patients with the hepatosplenic form of schistosomiasis. Results demonstrated that the presence of myofibroblasts varied considerably from case to case and was always related to smooth muscle cell dispersion, which occurred around medium-sized damaged portal vein branches. By sequential observation of several cases, it was evident that myofibroblasts derived by differentiation of vascular smooth muscle and gradually tended to disappear, some of them further differentiating into fibroblasts. Thus, in schistosomal pipestem fibrosis myofibroblasts appear as transient cells, focally accumulated around damaged portal vein branches, and do not seem to have by themselves any important participation in the pathogenesis of hepatosplenic schistosomiasis.

Effect of interferon-alpha on experimental septal fibrosis of the liver - study with a new model

Interferon-alpha is used in antiviral therapy in humans, mainly for viral hepatitis B and C. An anti-fibrotic effect of interferon has been postulated even in the absence of anti-viral response, which suggests that interferon directly inhibits fibrogenesis. Rats infected with the helminth Capillaria hepatica regularly develop diffuse septal fibrosis of the liver, which terminates in cirrhosis 40 days after inoculation. The aim of this study was to test the anti-fibrotic effect of interferon in this experimental model. Evaluation of fibrosis was made by three separate methods: semi-quantitative histology, computerized morphometry and hydroxyproline measurements. Treatment with interferon-alpha proved to inhibit the development of fibrosis in this model, especially when doses of 500,000 and 800,000 IU were used for 60 days. Besides confirming the anti-fibrotic potential of interferon-alpha on a non-viral new experimental model of hepatic fibrosis, a clear-cut dose-dependent effect was observed.

Worm load and septal fibrosis of the liver in Capillaria hepatica-infected rats

Inocula, varying from 15 to 1,000 embryonated Capillaria hepatica eggs, were administered to young adult rats by gastric tube, in an attempt to investigate the influence of worm load in the production of septal fibrosis of the liver. Low doses of 15, 30 or 50 eggs were sufficient to produce septal fibrosis, but it appeared with variable degrees of intensity and always with focal distribution. Septal fibrosis became diffuse, progressive with time, and already well developed 40 days after infection, when 100 eggs or more were administered. However, higher inocula (200, 500 and 1,000 eggs) did not intensify septal fibrosis, although the number of parasitic focal lesions proportionally augmented.

Intestinal fibrovascular nodules caused by Schistosoma mansoni infection in Calomys callosus Rengger, 1830 (Rodentia: Cricetidae): a model of concomitant fibrosis and angiogenesis

Human schistosomiasis develops extensive and dense fibrosis in portal space, together with congested new blood vessels. This study demonstrates that Calomys callosus infected with Schistosoma mansoni also develops fibrovascular lesions, which are found in intestinal subserosa. Animals were percutaneously infected with 70 cercariae and necropsied at 42, 45, 55, 80, 90 and 160 days after infection. Intestinal sections were stained for brightfield, polarization microscopy, confocal laser scanning, transmission and scanning electron microscopies. Immunohistological analysis was also performed and some nodules were aseptically collected for cell culture. Numerous intestinal nodules, appearing from 55 up to 160 days after infection, were localized at the interface between external muscular layer and intestinal serosa, consisting of fibrovascular tissue forming a shell about central granuloma(s). Intranodular new vessels were derived from the vasculature of the external vascular layer and were positive for laminin, chondroitin-sulfate, smooth muscle alpha-actin and FVIII-RA. Fibroblastic cells and extracellular matrix components (collagens I, III and VI, fibronectin and tenascin) comprised the stroma. Intermixed with the fibroblasts and vessels there were variable number of eosinophils, macrophages and haemorrhagic foci. In conclusion, the nodules constitute an excellent and accessible model to study fibrogenesis and angiogenesis, dependent on S. mansoni eggs. The fibrogenic activity is fibroblastic and not myofibroblastic-dependent. The angiogenesis is so prominent that causes haemorrhagic ascites.

Host nutritional status as a contributory factor to the remodeling of schistosomal hepatic fibrosis

Weaning Swiss mice were percutaneously infected with 30 cercariae of Schistosoma mansoni and submitted to a shifting either from a deficient to a balanced diet or vice-versa, for 24 weeks. The nutritional status was weekly evaluated by measurements of growth curves and food intake. Hepatic fibrosis and periovular granulomas were studied by histological, morphometric and biochemical methods. All mice fed on a deficient diet failed to develop periportal "pipestem" fibrosis after chronic infection. An unexpected finding was the absence of pipestem fibrosis in mice on normal diet, probably related to the sample size. The lower values for nutritional parameters were mainly due to the deficient diet, rather than to infection. Liver/body weight ratio was higher in "early undernutrition" group, after shifting to the balanced diet. Volume density and numerical density of egg granulomas reached lowest values in undernourished animals. The amount of collagen was reduced in undernourished mice, attaining higher concentrations in well-fed controls and in "late undernutrition" (balanced diet shifted to a deficient one), where collagen deposition appeared increased in granulomas. That finding suggested interference with collagen degradation and resorption in "late" undernourished animals. Thus, host nutritional status plays a role in connective tissue changes of hepatic schistosomiasis in mice.

Experimental hepatic fibrosis due to Capillaria hepatica infection (differential features presented by rats and mice)

Rats and mice are among the most susceptible hosts for the helminth Capillaria hepatica. More information on the similarities and differences between the hepatic pathology presented by these two parasite hosts are needed, since they may represent good models for the study of hepatic fibrosis. Early changes are similar for both hosts and are represented by necro-inflammatory lesions around dead parasites and their eggs and diffuse and intense reactive hepatitis. Although worms remain alive longer in mice than in rats, hepatic changes are more rapidly and deeply modulated in the former, even leading to almost complete disappearance of fibrosis. As for the rats, the modulation of the focal lesions is followed by the formation of septal fibrosis, a process where fine and long fibrous septa appear connecting portal spaces and central veins in such a way as to form a final morphologic picture of cirrhosis. Hepatic functional changes usually present good correlations with the morphologic findings at the different phases of the infection evolution. Therefore C. hepatica infection in rats and mice represent two different models of hepatic fibrosis and these differences, if properly known and understood, can be explored to answer different questions regarding several aspects of hepatic fibrosis

Infection of Calomys callosus (Rodentia Cricetidae) with strains of different Trypanosoma cruzi biodemes: pathogenicity, histotropism, and fibrosis induction

The influence of different Trypanosoma cruzi biodemes on the evolution of the infection and on the histopathological lesions of the heart and skeletal muscles, during the experimental infection of Calomys callosus, was investigated. Three groups of C. callosus were infected, respectively, with parasite strains representative of three different Biodemes: Type I (Y strain), Type II (21 SF strain), and Type III (Colombian strain). For each group, normal C. callosus were also used as controls. Marked differences have been detected in the responses of C. callosus to the infection with the three strains in this model. The strains Types I and II (Y and 21 SF) determined moderate lesions, mostly in the myocardium, with low parasitism, a rapid course, and total regression of the lesions by the 60th day of infection. Differently, Type III strain (Colombian), was more pathogenic for C. callosus and induced necrotic-inflammatory lesions in skeletal muscles and myocardium, in correspondence to intracellular parasitism. Proliferation of fibroblasts and amorphous matrix deposits, followed by interstitial fibrosis were present. Progressive regression of the inflammatory changes and collagen deposits occurred spontaneously. The progression and regression of both inflammation and fibrosis induced by the Colombian strain were further submitted to quantitative evaluation by morphometry. Results of the morphometric studies presented good correlation with the histopathological findings. The results confirm the importance of the different biodemes in the determination of tissue lesions and the peculiarities of response of C. callosus to infection with T. cruzi.

Immunological tolerance to pig-serum partially inhibits the formation of septal fibrosis of the liver in Capillaria hepatica-infected rats

Systhematized septal fibrosis of the liver can be induced in rats either by repeated intraperitoneal injections of pig-serum or by Capillaria hepatica infection. The relationship between these two etiological factors, as far as hepatic fibrosis is concerned, is not known, and present investigation attempts to investigate it. C. hepatica-induced septal fibrosis of the liver was considerably inhibited in rats previously rendered tolerant to pig-serum. Pig-serum-tolerant rats developed antibodies against pig-serum when infected with C. hepatica, but this did not happen when the infection occurred in normal rats. On the other hand, anti-C. hepatica antibodies failed to recognize any epitope in pig-serum, by Western blot. However, no evidence of an immunological cross reactivity was found, at least at the humoral level. Alternatively, cell-mediated mechanisms may be involved, and further investigations are warranted.

Malnutrition and hepatic fibrosis in murine schistosomiasis

In this paper, four different approaches attempting to reproduce the schistosomal liver fibrosis in undernourished mice are reported: shifting from a deficient to a balanced diet and vice-versa, repeated infections, influence of the genetic background, and immunological response. Infections were performed with 30 cercariae of Schistosoma mansoni and lasted at least four months. Undernourished mice were unable to reproduce the picture of "pipestem" fibrosis, except the C57 BL/10 inbred strain, four out of 21 mice developing the liver lesion. A link of this histological finding to the type of parasite strain can not be discarded at the moment. Repeated infections increased collagen deposition mainly in well nourished animals (seven out of 16 Swiss mice developed "pipestem"-like fibrosis). In undernourished infected Swiss mice the serum levels of soluble egg antigen specific antibodies IgG1, IgG2a, IgG2b, and IgG3 were two to four times lower than those detected for well nourished controls. The decreased humoral immune response coupled to the morphological, morphometric, and biochemical results reinforce the influence of the host nutritional status on the connective tissue changes of hepatic schistosomiasis.

Capillaria hepatica in rats: focal parasitic hepatic lesions and septal fibrosis run independent courses

Capillaria hepatica causes two main lesions in the liver of rats: multifocal chronic inflammation, directly related to the presence of disintegrating parasites and their eggs, and a process of systematized septal fibrosis. The comparative behavior of these two lesions was investigated in rats experimentally infected with 600 embryonated eggs, following either corticosteroid treatment or specific antigenic stimulation, in an attempt to understand the relationship between these two lesions, and the pathogenesis of septal fibrosis. The two treatments differently modified the morphological aspects of the focal parasitic-related lesions, but did not interfere with the presentation of diffuse septal fibrosis, although a mild decrease in the degree of fibrosis occurred in corticoid-treated animals. These findings indicate that although the two lesions are C. hepatica induced, they are under different pathogenetic control, the induction of septal fibrosis being triggered during early infection to follow an independent pathway.

Remodeling of hepatic vascular changes after specific chemotherapy of schistosomal periportal fibrosis

Hepatosplenic schistosomiasis was the first human disease in which the possibility of extensive long standing hepatic fibrosis being degraded and removed has been demonstrated. When such changes occurred, the main signs of portal hypertension (splenomegaly, esophageal varices) progressively disappeared, implying that a profound vascular remodeling was concomitantly occurring. Hepatic vascular alterations associated with advanced schistosomiasis have already been investigated. Obstruction of the intrahepatic portal vein branches, plus marked angiogenesis and compensatory hyperplasia and hypertrophy of the arterial tree are the main changes present. However, there are no data revealing how these vascular changes behave during the process of fibrosis regression. Here the mouse model of pipestem fibrosis was used in an investigation about these vascular alterations during the course of the infection, and also after treatment and cure of the disease. Animals representing the two polar hepatic forms of the infection were included: (1) "isolated granulomas" characterized by isolated periovular granulomas sparsely distributed throughout the hepatica parenchyma; and (2) 'pipestem fibrosis' with periovular granulomas and fibrosis being concentrated within portal spaces, before and after treatment, were studied by means of histological and vascular injection-corrosion techniques. Instances of widespread portal vein obstruction of several types were commonly found in the livers of the untreated animals. These obstructive lesions were soon repaired, and completely disappeared four months following specific treatment of schistosomiasis. Treatment was accomplished by the simultaneous administration of praziquantel and oxamniquine. The most impressive results were revealed by the technique of injection of colored masses into the portal system, followed by corrosion in strong acid. The vascular lesions of non-treated pipestem fibrosis were represented in the plastic casts by considerable diminution of the fine peripheral portal vein radicles, plus dilatation of periportal collaterals. Four months after treatment, this last picture appeared replaced by tufts of newly interwoven vessels formed along the main portal vein branches, disclosing a strong angiomatoid reparative change. Understanding about the cellular elements at play during fibro-vascular repairing changes of hepatic schistosomiais represents a matter of considerable scientific and conceptual importance. At present time one may only speculate about the participation of some type of natural stem-cell capable of restoring the diseased liver back to normal once the cause of the disorder has been eliminated.

Disagreement between ultrasound and magnetic resonance imaging in the identification of schistosomal periportal fibrosis

Abdominal ultrasound (US) has been widely used in the evaluation of patients with schistosomiasis mansoni. It represents an important indirect method of diagnosis and classification of the disease, and it has also been used as a tool in the evaluation of therapeutic response and regression of fibrosis. We describe the case of a man in whom US showed solid evidence of schistosomal periportal fibrosis and magnetic resonance imaging revealed that periportal signal alteration corresponded to adipose tissue which entered the liver togheter with the portal vein.