Four whole-istic aspects of schistosome granuloma biology: fractal arrangement, internal regulation, autopoietic component and closure

This paper centers on some whole-istic organizational and functional aspects of hepatic Schistosoma mansoni granuloma, which is an extremely complex system. First, it structurally develops a collagenic topology, originated bidirectionally from an inward and outward assembly of growth units. Inward growth appears to be originated from myofibroblasts derived from small portal vessel around intravascular entrapped eggs, while outward growth arises from hepatic stellate cells. The auto-assembly of the growth units defines the three-dimensional scaffold of the schistosome granulomas. The granuloma surface irregularity and its border presented fractal dimension equal to 1.58. Second, it is internally regulated by intricate networks of immuneneuroendocrine stimuli orchestrated by leptin and leptin receptors, substance P and Vasoactive intestinal peptide. Third, it can reach the population of ± 40,000 cells and presents an autopoietic component evidenced by internal proliferation (Ki-67+ Cells), and by expression of c-Kit+ Cells, leptin and leptin receptor (Ob-R), granulocyte-colony stimulating factor (G-CSF-R), and erythropoietin (Epo-R) receptors. Fourth, the granulomas cells are intimately connected by pan-cadherins, occludin and connexin-43, building a state of closing (granuloma closure). In conclusion, the granuloma is characterized by transitory stages in such a way that its organized structure emerges as a global property which is greater than the sum of actions of its individual cells and extracellular matrix components.

Cross-immunoreactivity between anti-potato apyrase antibodies and mammalian ATP diphosphohydrolases: potential use of the vegetal protein in experimental schistosomiasis

We have previously showed that Schistosoma mansoni ATP-diphosphohydrolase and Solanum tuberosum potato apyrase share epitopes and the vegetable protein has immunostimulatory properties. Here, it was verified the in situ cross-immunoreactivity between mice NTPDases and anti-potato apyrase antibodies produced in rabbits, using confocal microscopy. Liver samples were taken from Swiss Webster mouse 8 weeks after infection with S. mansoni cercariae, and anti-potato apyrase and TRITC-conjugated anti-rabbit IgG antibody were tested on cryostat sections. The results showed that S. mansoni egg ATP diphosphohydrolase isoforms, developed by anti-potato apyrase, are expressed in miracidial and egg structures, and not in granulomatous cells and hepatic structures (hepatocytes, bile ducts, and blood vessels). Therefore, purified potato apyrase when inoculated in rabbit generates polyclonal sera containing anti-apyrase antibodies that are capable of recognizing specifically S. mansoni ATP diphosphohydrolase epitopes, but not proteins from mammalian tissues, suggesting that autoantibodies are not induced during potato apyrase immunization. A phylogenetic tree obtained for the NTPDase family showed that potato apyrase had lower homology with mammalian NTPDases 1-4, 7, and 8. Further analysis of potato apyrase epitopes could implement their potential use in schistosomiasis experimental models.

Indirect effects of oral tolerance to ovalbumin interfere with the immune responses triggered by Schistosoma mansoni eggs

The objective of the present study was to investigate whether the injection of a tolerated protein (indirect effects) affects the formation of granulomas around Schistosoma mansoni eggs trapped in the lungs after intravenous (iv) injection into normal (noninfected) C57BL/6 mice (6 animals per group). To induce oral tolerance to chicken egg ovalbumin a 1/5 dilution of egg white in water was offered ad libitum in a drinking bottle for 3 days. Control mice received water. After 7 days, control and experimental animals were injected iv with 2,000 S. mansoni eggs through a tail vein. In some mice of both groups the iv injection of eggs was immediately followed by intraperitoneal (ip) immunization with 10 æg of dinitrophenylated conjugates of ovalbumin (DNP-Ova) emulsified in complete Freund's adjuvant (CFA) or only CFA; 18 days later, mice were bled and killed by ether inhalation. The lungs were fixed in formalin and embedded in paraffin. Serial sections of 5 æm were stained with Giemsa, Gomori's silver reticulin and Sirius red (pH 10.2). Granuloma diameters were measured in histological sections previously stained with Gomori's reticulin. Anti-DNP and anti-soluble egg antigen (SEA) antibodies were analyzed by ELISA. In mice orally tolerant to ovalbumin the concomitant ip injection of DNP-Ova resulted in significantly lower anti-SEA antibodies (ELISA*: 1395 ± 352 in non-tolerant and 462 ± 146 in tolerant mice) and affected granuloma formation around eggs, significantly decreasing granuloma size (area: 22,260 ± 2478 to 12,993 ± 3242 æmý). Active mechanisms triggered by injection of tolerated antigen (ovalbumin) reduce granuloma formation

Collagen arrangement in hepatic granuloma in mice infected with Schistosoma mansoni: dependence on fiber radiation centers

The collagen structure of isolated and in situ liver granuloma from Swiss Webster mice infected with Schistosoma mansoni was sequentially and three-dimensionally analyzed during different times of infection (early acute, acute, transitional acute-chronic, and chronic phases) by laser scanning confocal microscopy and electron scanning variable vacuum microscopy. The initial granuloma structure is characterized by vascular collagen residues and by anchorage points (or fiber radiation centers), from where collagenous fibers are angularly shed and self-assembled. During the exudative-productive stage, the self-assembly of these fibers minimizes energy and mass through continuous tension and focal compression. The curvature or angles between collagen fibers probably depends on the fibroblastic or myofibroblastic organization of stress fibers. Gradually, the loose unstable lattice of the exudative-productive stage transforms into a highly packed and stable architecture as a result of progressive compactness. The three-dimensional architecture of granulomas provides increased tissue integrity, efficient distribution of soluble compounds and a haptotactic background to the cells

Systemic modulation of peripheral eosinophilia (air pouch model) in Schistosoma mansoni infection

Schistosoma mansoni infection induces in their hosts a marked and sustained eosinophilia, which is influenced or modulated by complex mechanisms, that vary according to the phase of infection. To address this phenomenon, we used the air pouch (AP) model in control and infected Swiss webster mice, analyzing the cellular, tissue response and local expression of adhesion molecules [CD18 (ß2-chain), CD44, ICAM-1 (CD54), L-selectin (CD62L), CD49d (alpha4-chain), LFA1 (CD11a)]. Infected animals were studied at 3 (pre-oviposition phase), 7 (acute phase), and 14 (chronic phase) weeks after infection (5-6 mice/period of infection). Normal mice were age-matched. Results showed that after egg stimulation, compared with matched controls, the infected mice, at each point of infection, showed a lower eosinophil response in the acute (7 weeks) and chronic phase (14 weeks) of infection. However, when the infected mice were in pre-oviposition phase (3 weeks) their eosinophil response surpassed the control ones. In the AP wall of infected mice, a significant decrease in the expression of ICAM-1 and CD44 in fibroblastic-like cells and a reduction in the number of CD18 and CD11 a in migratory cells were observed. The other adhesion molecules were negative or weakly expressed. The results indicated that in the air pouch model, in S. mansoni-infected mice: (1) eosinophil response is strikingly down-regulated, during the acute ovular phase; (2) in the pre-oviposition phase, in contrast, it occurs as up-regulatory modulation of eosinophil responses, in which the mechanisms are completely unknown; (3) in the chronic phase of the infection, the down modulation of eosinophil response is less pronounced; (4) Down-regulation of adhesion molecules, specially of ICAM-1 appear to be associated with the lower eosinophil response.

Coelom-associated lymphomyeloid tissue (milky spots): site of lymphoid and myelomonocytic cell generation

Pleural and peritoneal milky spots (MS) are small morphofunctional structures representing subsidiary foci of coelom-associated lymphomyeloid tissue (CALT). In this paper we studied the cellular composition of CALT in normal and Schistosoma mansoni-infected mice. In the healthy mouse, CALT is mainly composed of IgM (+) B cells and presents lower numbers of CD23 and CD45R (B220) B2 lymphocytes. When activated by the infection, it may show pronounced lymphocytosis, plasmocytogenesis (IgM >IgG>IgA>IgG2a>IgG1) and myelomonocytosis. The lymphocytes were mainly of the B1 type (double positive CD5/IgM), with smaller number of T cells (TCR alpha beta (+), TCR gamma delta (+), CD3 (+) and CD5 (+)) and conventional B2 cells (B220 (+), CD23 (+)). The myeloid compartment was composed of immature and mature cells of monocyte/macrophage, eosinophil, neutrophil and megakaryocytic lineages, especially in the omental milky spots. CALT is also a favorable microenvironment for LFA-1 (+) mast cells. Thus, CALT appears to be a mixed lymphoid organ, with secondary and/or primary lymphoid organ functions, being an important site of B1 cell generation, plasma cell maturation and extramedullar hematopoiesis. CALT operates as an interface between blood and lymphatic circulation and coelomic cavities, because locally or externally produced cells have easy and ready access to the pleural and peritoneal cavities. Furthermore, MS cells can escape into blood and lymphatic vessels, providing lymphocytes to other lymphoid organs and to the mucosa-associated lymphoid tissue.

Calomys callosus: an alternative model to study fibrosis in schistosomiasis mansoni. The pathology of the acute phase

Twenty Calomys callosus, Rengger, 1830 (Rodentia-Cricetidae) were studied in the early stage of the acute schistosomal mansoni infection (42nd day). The same number of Swiss Webster mice were used as a comparative standard. Liver and intestinal sections, fixed in formalin-Millonig and embedded in paraffin, were stained with hematoxilin and eosin, PAS-Alcian Blue, pH = 1.0 and 2.5, Lennert's Giemsa, Picrosirius plus polarization microscopy, Periodic acid methanamine silver, Gomori's silver reticulin and resorcin-fuchsin. Immunohistological study (indirect immunofluorescence and peroxidase labeled extravidin-biotin methods) was done with antibodies specific to pro-collagen III, fibronectin, elastin, condroitin-sulfate, tenascin, alpha smooth muscle actin, vimentin and desmin. The hepatic granulomas were small, reaching only 27 of the volume of the hepatic Swiss Webster granuloma. They were composed mainly by large immature macrophages, often filled by schistosomal pigment, characterizing an exsudative-macrophage granuloma type. The granulomas were situated in the parenchyma and in the portal space. They were often intravascular, poor of extracellular matrix components, except fibronectin and presented, sometimes alpha smooth muscle actin and vimentin positive cells. The C. callosus intestinal granulomas were similar to Swiss Webster, showing predominance of macrophages. Therefore, the C. callosus acquire very well the Schistosoma mansoni infection, without developing strong hepatic acute granulomatous reaction, suggesting lack of histopathological signs of hypersensitivity.

Extramedullary hematopoiesis in murine schistosomiasis mansoni

During Schistosoma mansoni infection, there is morphological evidence of involvement of various hematopoietic growth factors, which cause eosinophil, neutrophil, megakaryocytic and erythroid extramedullary foci in the liver, lymph nodes and omental and mesenteric milky spots. While the eosinophil metaplasia in the periphery of hepatic granulomas roughly reproduced the intensity of the medullary eosinopoiesis, the neutrophil metaplasia, on the contrary, was more intense during the period of neutrophil depression in the bone marrow. This fact suggests that extramedullary hematopoietic foci are locally regulated, and amplify and/or compensate the systemic hematopoietic response during the infection.

Fibrosis patterns of lesions developed by athymic and euthymic mice infected with Paracoccidioides brasiliensis

Athymic and euthymic mice with BALB/c background were used to study the patterns of fibrosis during ip infection with a virulent isolate of Paracoccidioides brasiliensis. Specimens from various organs were collected from the animals at 1,4 and 7 weeks after infection and observed under light microscopy using various histologic staining methods. Lesions from the first week of infection, in both animal groups, presented a predominance of collagen III over I, carboxylated proteoglycans, and a tendency to encapsulation. From 4 weeks onward, the lesions of nu/+mice tended to involute to macrophage-pseudoxanthomatous aggregates or to encapsulation with an increase of collagen I and sulfated proteoglycans. On the contrary, with the evolution of the infection, the nu/nu mice displayed permanently active lesions, rich in reticular fibers and carboxylated proteoglycans, with varied amounts of collagens III and I, without or with minimal encapsulation. However, independent of the type of mice, or of the type of lesions, the minimal P. brasiliensis-ECM unit was formed by a fibrillar cocoon of reticular fibers that encloses an individual yeast or a "family" composed of a mother cell plus one or various peripheral daughter cells, alone or engulfed by macrophages or giant cells. The overal difference of the lesions of nude and normal mice was not in isolated aspects of their components, but in the general architecture of the lesions. Those of nu/+mice were either of involutive or of encapsulated type (slightly active), and those of nu/nu mice were of the sustained-expansive type (very active), without or with minimal encapsulation

Milky spots reactions to schistosomal mansoni infection

Milky spots (MS), considered by the authors as a Coelomatic Lympho-myelopoietic Organ (CLMO), present a strong reactivity during experimental schistosomal mansoni infection, characterized by an increase of lymphocytes, macrophages, plasmocytes, mast cells, neutrophils and expression of eosinophil metaplasia. Intraperitoneal injection of purified Schistosoma mansoni (Sm) eggs provoked a rise in the number and size of MS, which developed the sessile marginal and pedunculated types. The authors conclude that egg antigens are, at least partially, responsible for MS reactivity during Sm infection

Comparative distribution of eosinophils in bone marrow, blood and peritoneal cavity in murine schistosomiasis

Eosinophilia in murine schistosomiasis is very intense and extensive, involving distinct compartments such as bone marrow, blood, peritoneal cavity and tissues. Comparison of the shapes of eosinophil concentration or distribution curves showed a synchronization of the tendencies around 50% between blood and bone marrow, 33 to 64% between bloode and peritoneal cavity, and 33 to 43% between peritoneal cavity and bone marrow. The hepatic eosinophil granulocytopoiesis or metaplasia follows the same pattern as observed in bone marrow. Schistosoma infection can be divided into three distinct phases based on the eosinophilic response: 1) non- or low-productive phase (before 35-40 days of infection), 2) acute productive phase (from 35-40 to 70-90 days), and 3) chronic productive phase (after 70-90 days of infection)

Phillocaulis variegatus - an intermediate host of Angiostrongylus costaricensis in south Brazil

Moluscos coletados em cinco localidades no estado do Rio Grande do Sul (Brasil) foram digeridos e examinados. As lesmas infectadas foram identificadas como Phyllocaulis variegatus e as larvas encontradas foram inoculadas per os em camundongos albinos. Após 50 dias, parasitos com as características de A. costaricensis foram recuperados do sistema arterial mesentérico. Estes resultados estabelecem o papel do P. variegatus como hospedeiro intermediário de A. costaricensis no sul do Brasil, onde diversos casos de angiostrongilíase abdominal têm sido diagnosticados

Pancreatic involvement in murine Schistosomiasis

Pancreatic involvement during murine schistosomiasis is frequent (30 to 80%), heterogeneous, usually mild, but can occasionally be severe, characterized by granulomatous pancreatitis. After infection, pancreatic granulomas appear from day 50 on, with th most severe pancreatitis being demonstrable between days 90 and 100. Mice thus appear to be a useful model for study of the pathogenesis of Schistosoma mansoni-induced pancreatitis

Paracoccidioidomycoses: a sequential histopathologic study of lesions in experimentally infected rats

Foram utilizados ratos albinos, fêmeas, para o estudo histopatológico sequencial da paracoccidioidomicose experimental. Os animais foram inoculados intraperitonealmente com uma cepa de Paracoccidioides brasiliensis na fase leveduriforme e sacrificados, em determinados intervalos, a partir de 1 a 168 dias pós-infecçäo;cada animal recebeu um inóculo de 4 x 10 células em 0,8 ml de salina. Os animais controles receberam salina contendo raspado do meio de cultura. Foram estudados tecidos correspondentes à área de inoculaçäo. Analisou-se pela microscopia óptica o processo inflamatório granulomatoo em todo o seu conjunto, estudando a populaçäo celular, a matriz extracellular e apresença e características do fungo. Os resultados possibilitaram desmembrar a cinética da resposta inflamatória em três fases: 1) neutrofílica ou macrofágica-neutrofílica; 2) pré-granulomatosa; 3) granulomatosa. A síntese de matriz extracelular iniciou=se pela deposiçäo de material fibrinóide, intensificando-se de modo gradativo com depósito de colágeno, de proteoglicanos e glicoproteínas. Os parasitos estavam presentes em todas as fases estudadas. Períodos de reativaçäo da doença eram nitidamente evidenciados através da concomitância de granulomas recém-formados com granulomas mais antigos, indicando que o processo granulomatoso neste modelo näo resolve a doença, nem täao pouco consegue limitar a disseminaçäo do fungo por um período prolongado

Intrinsic vascularization of granuloma in experimental paracoccidioidomycosis

Rats infected with Paracoccidioides brasiliensis exhibited vascularized granulomas during the granulomatous stage, with more intensity on the 49 th day after infection. These data change the classical belief that the granuloma is always an avascular structure. The authors suggest that the granuloma is under the balanced influence of angiogenic or angiostatic factors, with the usual predominance of the latter, with the consequent advantage of limited antigen diffusion to the vascular system

Participation of endothelial cells in murine schistosomiasis

The endothelial cells participate in the morphological events occuring during murine schistosomiasis, taking part in the development of hepatic periovular granuloma. The cells also show an increase in the expression of Factor WIII - related antigen in the portal vessels and hepatic sinusoids during the infection. Endothelial cells are suggested play an important role in the pathogenesis of the disease and in the balance of the coagulant - anticoagulant mechanisms which favor the intravascular survial of the parasites

In vivo kinetics of eosinophils and mast cells in experimental murine Schistosomiasis

During the schistosomiasis infection there is a [quot ]dance of the cells[quot ], varying from site to site and related to the time of infection. 1 - Eosinophil levels exhibit a bimodal pattern, with the first peak related to the egg deposition and maturation and increased Kupfferian hyperplasia; the second peak precedes the death of some adult worms; 2 - The peritoneal eosinophilic levels are inversely proportional to the blood eosinophilic levels; 3 - Eosinopoiesis in the bone marrow begins at day 40, reaching the highest levels at day 50 and coincides with hepatic eosinophilic and neutrophilic metaplasia; 4 - Peritoneal mast cell levels present a bimodal pattern similar to the blood eosinophils, and inverse to the peritoneal eosinophils. They also show a cyclic behaviour within the hepatic and intestinal granulomas. Integral analysis of the events related to the eosinophils in the blood, bone marrow, peritoneal cavity and hepatic and intestinal granulomas allows the detection of two important eosinophilic phases: the first is due to mobilization and redistribution of the marginal pool and the second originates from eosinophilic production in the bone marrow and liver. The productive phase is characterized by an increase in the number of eosinophils and monocyte/macrophages, and a decrease in neutrophils and stabilization of megakariocytes and erithroid lineages.