O sistema imune inato intestinal em pacientes com estrongiloidíase / The innate intestinal immune system in patients with strongyloidiasis

Foram estudados fragmentos da mucosa duodenal obtidos de nove pacientes apresentando sintomas leves de estrogiloidiase, nove com sintomas moderados, sete com sintomas graves e sete indivíduos aparentemente normais. A muramidase(lisozima) foi imunocitoquimicamente demonstrada em cortes contracorados pela técnica do PAS. Havia aparente aumento progressivo na secreçao de muramidase pela célula de Paneth aaacompanhado o agravamento dos sintomas, näo obstante o fato de que a sua populaçao permanecesse constante. Decréscimo progressivo no número de células caliciformes foi observado enquanto, concomitantemente, haavia aumento na populaçao de células intermediarias. Esses resultados foram interpretados como a indicaçao da participaçao do sistema imune inato intestinal no estabelecimento da interaçao Strongyloides stercoralis/hospedeiro, através do aumento da secreçao da enzima mucolítica muramidase

Absence of lysozyme (muramidase) in the intestinal Paneth cells of newborn infants with necrotising enterocolitis.

AIM: To determine immunocytochemically whether preterm and newborn infants with necrotising enterocolitis (NEC) show differences in numbers of lysozyme positive Paneth cells compared with normal controls, and to relate the findings to the possibility that lysozyme deficiency may facilitate the bacterial infections thought to be associated with this condition. METHODS: Tissues from 10 infants with NEC and from 11 matched controls were sectioned and stained immunocytochemically for lysozyme. Differences in the numbers of Paneth cells and degree of lysozyme positivity in the tissues were assessed. RESULTS: Tissues from NEC patients showed no, or very few, lysozyme positive Paneth cells, whereas controls showed strong positive staining. CONCLUSIONS: A deficiency or developmental defect in Paneth cells, resulting in an absence of lysozyme, may render the intestine more susceptible to bacterial infection, allowing organisms to adhere and translocate across the mucosa. Such enhancement of infection may contribute to the pathogenesis of NEC.

Intra-abdominal sepsis: an immunocytochemical study of the small intestine mucosa.

AIM: To investigate immunocytochemical changes in intestinal tissues from patients with intra-abdominal sepsis, and to relate the changes to the possibility of enhanced bacterial adhesion and translocation. METHODS: Tissues from 17 patients suffering from intra-abdominal sepsis and from controls were sectioned and stained immunocytochemically for IgA, IgM, secretory component, J chain, and HLA-DR. Differences in the distribution and characteristics of positively staining cells between the patient groups were assessed. RESULTS: Patients with intra-abdominal sepsis had noticeable reductions in numbers of IgA and IgM plasma cells, reduced J chain staining, and had little immunoglobulin on the surfaces of enterocytes. In contrast, HLA-DR positive cells were increased in the sepsis compared with the control group. The plasma cells present showed cytological changes suggestive of apoptosis. CONCLUSIONS: Stress associated with sepsis and its immediate causes might result in increased plasma glucocorticoid levels that bring about apoptosis of mucosal plasma cells (or their precursors). The consequent reduction in expression of IgA and IgM may favour bacterial adhesion to the enterocytes and facilitate bacterial translocation into the tissues.

Immunocytochemistry of mucosal changes in patients infected with the intestinal nematode Strongyloides stercoralis.

AIM: To investigate the immunopathological changes in duodenal tissues induced by strongyloidiasis and to relate these to degrees of clinical severity. METHODS: Tissues taken from 21 patients showing mild, moderate or severe symptoms of strongyloidiasis, and from non-infected controls, were sectioned and stained immunocytochemically for IgA, secretory component (SC) and HLA-DR. Immunopathology was assessed by changes in numbers, intensity and distribution of stained cells. RESULTS: Parasitised individuals showed villous atrophy and crypt hyperplasia. There was notable infiltration of the lamina propria by IgA positive plasma cells and of the epithelium by intraepithelial lymphocytes. Infection was also associated with increased expression of SC and decreased expression of HLA-DR in epithelial cells. Changes in all parameters correlated with degree of clinical severity. CONCLUSIONS: Profound mucosal changes are induced by strongyloidiasis. Some are analogous to those seen in coeliac disease, but others seem quite unusual. It is likely that these changes are functionally related to the immunopathophysiological consequences of infection seen in patients with severe disease.

Immunocytochemical demonstration that human duodenal Brunner's glands may participate in intestinal defence.

The immunocytochemical demonstration of IgA and IgM in some secretory units of human Brunner's glands, associated with the presence of secretory component in all secretory cells, indicates the possibility that these glands assist the function of the intestinal crypts in transporting immunoglobulins into the gut lumen. In addition, the presence of muramidase (lysozyme) in the cells of the secretory units suggests that Brunner's glands continuously secrete bactericidal enzyme, thus reinforcing the function of the Paneth cells as contributors to nonspecific defence (innate immunity) in the intestinal tract.

Immunocytochemical study of the ontogeny of the marsupial Didelphis albiventris immune system.

Marsupials have considerable merits as models for studying the developmental dynamics of the mammalian immune system, but until recently there has been a conspicuous lack of specific immune probes to facilitate such studies. To begin a precise study of the ontogeny of the marsupial Didelphis albiventris we have used cross-reactive polyclonal antibodies raised against evolutionarily highly conserved peptides which form part of the antigen specific receptor complexes of human differentiated lymphocytes. Moreover, because of antigen receptor conservation, the antibodies also recognise specifically the immunocompetent T and B lymphocytes of other species including those in the organs of the opossum. Use of the antipeptide antibodies together with other cross-reacting antibodies has allowed us to study the cellular immunology of T and B cells and antigen presenting cells (APC) during the development of thymus, skin, lymph nodes and spleen in the Brazilian white-belly opossum. The molecular nature and identity of the T cell antigens detected in opossum tissues were confirmed by immunoblotting. These findings indicate that it is now possible to exploit these antibody probes for comparative mammalian studies, and indeed to investigate interesting features of the opossum, such as reaction of the immature immune system of the pouch young to antigenic stimulation.

Immunocytochemical study of the ontogeny of Peyer's patches in the Brazilian marsupial Didelphis albiventris.

A detailed ontogenetic immunocytochemical study is reported on gut-associated lymphoid development in the Brazilian marsupial Didelphis albiventris. This employed antibody probes raised to evolutionarily conserved peptides which have been shown to detect HLA-DR-like (class II MHC) antigens and T and B cell markers in a wide range of animal species. Cells with macrophage and dendritic morphology expressing class II MHC and a few cells expressing the T cell marker CD3 were found in the lamina propria of duodenal villi in early (approximately 24 mm crown-rump length) latent opossum. Cells with B cell markers were not detected until lactent animals reached > 60 mm. Development of Peyer's patches (PP) was seen first in the duodenum in 45-60 mm lactent animals, progressing to well developed PP in the duodenum and ileum in lactent animals > 80 mm. These PP, like those in weanling and juvenile animals, consisted of follicles with a network of class II MHC positive dendritic cells and round cells lacking T and B markers, but lacking well defined mantle zones. B cells were present mainly in the lymphatic sinuses, with CD3 T cells present between follicles in the PP and intraepithelially in the villi. The study reveals the sequential development of class II MHC positive dendritic cells, T cells and B cells in the intestinal ontogeny of the opossum PP. These features occurred initially exclusively in the duodenum and subsequently in the ileum, paralleling the physiological maturation of the gut in eutheria.

Morphometry of the microvilli of the enterocyte in the fetus of Didelphis albiventris before and after treatment with hydrocortisone hemisuccinate.

Intestinal closure is the blocking of biopolymers transferred across the small intestine epithelium that coincides with gut maturation. Sixty-one fetuses of D. albiventris, 45 to 55 mm in Crown-Rump length, were studied. Forty-one fetuses were injected subcutaneously with hydrocortisone hemisuccinate (0.4 mg/15.0 g of body weight) daily for 7 days in order to produce intestinal closure (TA). The control group (CA), consisted of twenty fetuses. Seven days after injection several fragments from the duodenum and ileum were prepared for transmission electron microscopic examination. Microvillus lengths and widths were determined from micrographs (final magnification: 100,000 x). The volume of the microvillus was calculated as the volume of the cylinder. Quantitative differences of the microvilli between the treated and control groups taken from duodenum and ileum material were noted. The distribution of the microvilli in the apical part of the enterocytes was more irregular in controls than in the TA group. Quantitatively, microvillus length in the duodenum was more than 50 per cent greater in TA than in CA and more than 30 per cent greater in TA than in CA in the ileum. Consequently, microvilli volume in the duodenum in TA was about 65 per cent greater than in CA and about 33 per cent greater than in CA in the ileum. Microvilli width presented little change and no statistically significant differences were found between TA and CA in either the duodenum or the ileum. These changes in the microvilli of the treated animals most probably indicate improvement in the absorptive function of the epithelium.

Immunocytochemical study of Peyer's patches follicular-associated epithelium in the marsupial, Didelphis albiventris.

The lack of probes defining leukocyte subpopulations has restricted ontogenetic studies of the opossum gut. We report for the first time the organization of the gut cellular immune components using species cross-reactive antibodies. Mouse monoclonal antibodies against human HLA-DR were used together with immunocytochemistry to demonstrate MHC class II-like antigens in the opossum Peyer's patches (PP). Positive staining was obtained in the M cell and enterocytes comprising the follicular-associated epithelium (FAE). Rabbit polyclonal antibody against human CD3 stained opossum thymocytes and T-cell dependent areas of spleen, lymph node, and PP interfollicular zones, but failed to stain intraepithelial lymphocytes in the FAE. In contrast rabbit polyclonal antibody against human IgA stained B-cell immunocytes and plasma cells present in the M-cell lateral invaginations. It is surmised that B-cell activation could occur in the opossum M-cell niches by thymus independent antigens, bypassing T-helper-cell function.

Effects of hydrocortisone acetate treatment on the small intestine of the lactent marsupial Didelphis albiventris.

When hydrocortisone acetate was administered subcutaneously to the lactent Brazilian white belly opossum, it induced precocious intestinal maturation. Histological features of the small intestine were reminiscent of those seen in human protein-caloric malnutrition. There were also indications that the glucocorticoid acted as an inducer of the differentiation of Paneth, goblet and granulomucous cells within the small intestinal epithelium. In the lactent opossum the small intestine is lined by an epithelium containing cells which exhibit intense endocytic activity (Krause et al. 1977; Santiago et al. 1983; H.B. Coutinho, Beck--unpublished) verified that subcutaneous injections of hydrocortisone acetate could determine intestinal closure in lactent opossums. Krause et al. and Santiago et al. have reported that in Didelphis virginiana and Didelphis albiventris, respectively, the goblet cells and Paneth cells differentiate lately in the development of pouch opossum young. Freitas et al. (1982) mentioned that the plasma cortisol level was very low in lactent white belly opossum, but preceding weaning there was an abrupt increase of the plasma cortisol reaching levels comparable to that obtained in adult animals. These facts suggested the need for investigations to clarify if cortisol could act as an inductive agent for the differentiation of goblet cells and Paneth cells in the lactent D. albiventris.

Histological and ultrastructural studies of the marsupial Didelphis albiventris Peyer's patches

Differing from the studied Eutheria the white belly opossum Peyer"s patches do not present a conspicous dome. M cells are located in the inmer layer of bilaminal invaginations formed at the bottom of the villi. A great variation in the morphology of M cells was observed. The enterocytes located at the epithelial inner layer may present endocytic vesicles, and the microvilli are shorter tha the microvilli of enterocytes lining the small intestine. As these morphological aspects have been described to exist in the enterocytes of the lancet opossum small intstine it was surmised that the opossum Peyer's patches special epithelium could represent the persistence in adult animals of a cellular pattern established before the intestinal maturation had occurred

Histological and ultrastructural studies of the marsupial Didelphis albiventris Peyer's patches.

Differing from the studied Eutheria the white belly opossum Peyer's patches do not present a conspicuous dome. M cells are located in the inner layer of bilaminal formed at the bottom of the villi. A great variation in the morphology of M cells was observed. The enterocytes located at the epithelial inner layer may present endocytic vesicles, and the microvilli are shorter than the microvilli of enterocytes lining the small intestine. As these morphological aspects have been described to exist in the enterocytes of the lactent opossum small intestine it was surmised that the opossum Peyer's patches special epithelium could represent the persistence in adult animals of a cellular pattern established before the intestinal maturation had occurred.

Secretory activity and endocrine regulation of male accessory glands in the blood-sucking bug Panstrongylus megistus (Hemiptera: Reduviidae).

The epithelial cells of Panstrongylus megistus male accessory glands (MAG) present ultrastructural characteristics of a secretory cell. Their secretory products are accumulated in the lumen of the four MAG lobes. During the first 8 days of adult life a strong secretion activity occurs, accumulating enough material to produce the first spermatophore. Cerebral neurosecretions as well as juvenile hormone are both involved in MAG secretory activity regulation. Juvenile hormone seems to be the responsible for the stimulation of most protein synthesis in male accessory glands. Cerebral neurosecretion seems to be necessary to stimulate juvenile hormone production and release by the corpus allatum. Furthermore, neurosecretion is required for some polypeptides synthesis by MAG. Although topic application of precocene II to adult males does not reproduce the same effects on MAG as does allatectomy, this compound causes strong reduction on male reproductive capacity.

Immunocytochemical demonstration of insulin in the mesonephros and metanephros of the Brazilian opossum Didelphis albiventris).

The presence of insulin the brush border and apical pole of the cells lining the pouch opossum mesonephric and metanephric proximal tubules was demonstrated by immunofluorescence and immunoperoxidase techniques. Positive result for insulin was also observed in the tufts of capillaries of some metanephric corpuscles.

Demonstration of insulin in the pancreas of the Didelphis albiventris (opossum) by immunocytochemical techniques.

The exocrine secretion of insulin by the suckling opossum was demonstrated by indirect immunofluorescence and immunoperoxidase techniques. In weanling and adult marsupial insulin is present in the pancreatic islets and in scattered cells located in the exocrine secretory units. Positive reaction was also observed in the brush border and in some isolated cells of the proximal segment of the suckling opossum small intestine.

Study of the cellular turnover in the small intestine of the lactent Didelphis albiventris.

A 1% trypan blue aqueous solution, buffered to pH 7.3, orally administrated was used as a tracer in an attempt to establish the cellular turnover in the small intestine epithelium of the lactent marsupial Didelphis albiventris. The complex trypan blue-milk albumin was captured by the cells lining the proximal, medium and distal segment of small intestine, differing from the rodents which are unable to absorb the dye by endocytosis at the level of the proximal small intestine. A complete cellular turnover was observed in the proximal small intestine 7 d after treatment, whilst in the medium and distal segments 8 d were required for the turnover to be completed.