Novas propriedades do SKTI (Inibidor de tripsina de soja): inibição para elastase neutrofílica humana e efeitos no processo de injúria pulmonar aguda

Seeds from legumes including the Glycine max are known to be a rich source of protease inhibitors. The soybean Kunitz trypsin inhibitor (SKTI) has been well characterised and has been found to exhibit many biological activities. However its effects on inflammatory diseases have not been studied to date. In this study, SKTI was purified from a commercial soy fraction, enriched with this inhibitor, using anion exchange chromatography Resource Q column. The purified protein was able to inhibit human neutrophil elastase (HNE) and bovine trypsin. . Purified SKTI inhibited HNE with an IC50 value of 8 µg (0.3 nM). At this concentration SKTI showed neither cytotoxic nor haemolytic effects on human blood cell populations. SKTI showed no deleterious effects on organs, blood cells or the hepatic enzymes alanine amine transferase (ALT) and aspartate amino transferase (AST) in mice model of acute systemic toxicity. Human neutrophils incubated with SKTI released less HNE than control neutrophils when stimulated with PAF or fMLP (83.1% and 70% respectively). These results showed that SKTI affected both pathways of elastase release by PAF and fMLP stimuli, suggesting that SKTI is an antagonist of PAF/fMLP receptors. In an in vivo mouse model of acute lung injury, induced by LPS from E. coli, SKTI significantly suppressed the inflammatory effects caused by elastase in a dose dependent manner. Histological sections stained by hematoxylin/eosin confirmed this reduction in inflammation process. These results showed that SKTI could be used as a potential pharmacological agent for the therapy of many inflammatory diseases

Bioecologia da ictiofauna marinha descartada pelo arrasto camaroeiro em praias da Bacia Potiguar, Brasil

The composition of ichthyofauna discarded by trawling shrimping, their reproductive status and feeding ecology were studied on the beaches of Basin Rio Grande do Norte, Brazil. Fish were collected monthly in the year of 2012. During biometrics, portions of the digestive tract and of gonads were removed, fixed in formalin 10% and Bouin, respectively, for be submitted to histological processing by the techniques of hematoxylin-eosin. Stomach content analyzes were performed using the methods of Frequency of Occurrence and Volumetric and was calculated the repletion index. Throughout the study period were recorded a total of 49 species. The fish assemblages differed between sections monitoring, with the highest abundance, biomass and indices of richness and diversity in sections B, D and C. Already the excerpt A, showed higher values for dominance and equitability. In the cluster analysis according to the faunal similarity was observed the formation of three groups: group I formed by excerpts B and D, group II by excerpt C and group III formed by excerpt A. The assessment of reproductive stage revealed that the fish assemblages discarded by trawling are composed mainly of juveniles. Regarding the feeding ecology, the species Larimus breviceps, Menticirrhus littoralis and Pomadasys corvinaeformis characterized as carnivorous with tendency to carcinofagia. Already Conodon nobilis characterized as carnivorous with tendency to piscivory, but all proved generalist-opportunistic with increase of feeding activity during drought. The dendrogram of grouping of the species developed based on the food items used shows the formation of four groups: Group I consists of species that feed mainly of "gastropod" and "sediment"; group II of "teleost fish"; the group III of "crustacea" and group IV of "echinodermata" and "bivalve". The anatomical and histological characteristics of the digestive tract were consistent with the dietary habits of the analyzed species. In this context, the Costa Branca of Rio Grande do Norte can be considered a feeding site and recruitment for juveniles, which, opportunistically, utilize resources associated with the background

Avaliação da expressão das moléculas HLA de classe I não clássicas HLA-G E HLA-E em espécimes gástricas de pacientes acometidos com a bactéria Helicobacter Pylori

The expression of human leukocyte antigen G (HLA-G) and human leukocyte antigen E (HLA-E) in physiological and pathological processes remains unknown, it is believed that these molecules play a fundamental role in the establishment and maintenance of immune tolerance by inhibiting the functions of immunocompetent cells. In literature we found no published study involving the bacterium Helicobacter pylori (H. pylori) with expression of HLA-G and HLA-E. The objective this study is investigated the expression of this protein in gastric biopsies of patients with the bacterium H. pylori. Sixty-four biopsies of the patients with diagnosis of infection by H. pylori were evaluated to expression of HLA-G and HLA-E. The samples were stratified according to the presence of carcinoma or peptic ulcers. Patients without H. pylori were used to control. To investigate the expression of this protein were used immunohistochemistry technique with monoclonal antibody anti-HLA-G and anti-HLA-E. Other criteria such as analysis of the inflammatory infiltrate (hematoxylin-eosin) and identification of H. pylori (Giemsa) were analyzed. We detected HLA-G and HLA-E molecules in the most samples containing ulcer and gastric carcinoma. In negative control group was not detected the presence of HLA-G and HLA-E. The presence of H. pylori seems modulate the expression of HLA-G and HLA-E, favoring the evolution of infection, giving different degrees of gastric lesion in epithelium of these patients

Novas propriedades do SKTI (Inibidor de tripsina de soja): inibição para elastase neutrofílica humana e efeitos no processo de injúria pulmonar aguda

Seeds from legumes including the Glycine max are known to be a rich source of protease inhibitors. The soybean Kunitz trypsin inhibitor (SKTI) has been well characterised and has been found to exhibit many biological activities. However its effects on inflammatory diseases have not been studied to date. In this study, SKTI was purified from a commercial soy fraction, enriched with this inhibitor, using anion exchange chromatography Resource Q column. The purified protein was able to inhibit human neutrophil elastase (HNE) and bovine trypsin. . Purified SKTI inhibited HNE with an IC50 value of 8 µg (0.3 nM). At this concentration SKTI showed neither cytotoxic nor haemolytic effects on human blood cell populations. SKTI showed no deleterious effects on organs, blood cells or the hepatic enzymes alanine amine transferase (ALT) and aspartate amino transferase (AST) in mice model of acute systemic toxicity. Human neutrophils incubated with SKTI released less HNE than control neutrophils when stimulated with PAF or fMLP (83.1% and 70% respectively). These results showed that SKTI affected both pathways of elastase release by PAF and fMLP stimuli, suggesting that SKTI is an antagonist of PAF/fMLP receptors. In an in vivo mouse model of acute lung injury, induced by LPS from E. coli, SKTI significantly suppressed the inflammatory effects caused by elastase in a dose dependent manner. Histological sections stained by hematoxylin/eosin confirmed this reduction in inflammation process. These results showed that SKTI could be used as a potential pharmacological agent for the therapy of many inflammatory diseases

Prevalence of helicobacter pylori infection in advanced gastric carcinoma

There is substantial evidence that infection with Helicobacter pylori plays a role in the development of gastric cancer and that it is rarely found in gastric biopsy of atrophic gastritis and gastric cancer. On advanced gastric tumors, the bacteria can be lost from the stomach. Aims - To analyze the hypothesis that the prevalence of H.pylori in operated advanced gastric carcinomas and adjacent non-tumor tissues is high, comparing intestinal and diffuse tumors according to Lauren’s classifi cation. Methods - A prospective controlled study enrolled 56 patients from “Hospital Universitário”, Federal University of Rio Grande do Norte, Natal, RN, Brazil, with advanced gastric cancer, treated from February 2000 to March 2003. Immediately after partial gastrectomy, the resected stomach was opened and several mucosal biopsy samples were taken from the gastric tumor and from the adjacent mucosa within 4 cm distance from the tumor margin. Tissue sections were stained with hematoxylin and eosin. Lauren‘s classifi cation for gastric cancer was used, to analyse the prevalence of H. pylori in intestinal or diffuse carcinomas assessed by the urease rapid test, IgG by ELISA and Giemsa staining. H. pylori infected patients were treated with omeprazole, clarithromycin and amoxicillin for 7 days. Follow-up endoscopy and serology were performed 6 months after treatment to determine successful eradication of H. pylori in non-tumor tissue. Thereafter, follow-up endoscopies were scheduled annually. Chi-square and MacNemar tests with 0.05 signifi cance were used. Results - Thirty-four tumors (60.7%) were intestinal-type and 22 (39.3%) diffuse type carcinomas. In adjacent non-tumor gastric mucosa, chronic gastritis were found in 53 cases (94.6%) and atrophic mucosa in 36 patients (64.3%). All the patients with atrophic mucosa were H. pylori positive. When examined by Giemsa and urease test, H. pylori positive rate in tumor tissue of intestinal type carcinomas was higher than that in diffuse carcinomas. In tumor tissues, 34 (60.7%) H. pylori-positive in gastric carcinomas were detected by Giemsa method. H. pylori was observed in 30 of 56 cases (53.5%) in tissues 4 cm adjacent to tumors. This difference was not signifi cant. Eradication of H. pylori in non-tumor tissue of gastric remnant led to a complete negativity on the 12th postoperative month. Conclusions - The data confi rmed the hypothesis of a high prevalence of H. pylori in tumor tissue of gastric advanced carcinomas and in adjacent non-tumor mucosa of operated stomachs. The presence of H. pylori was predominant in the intestinal-type carcinoma

Avaliação bioquímica e hostológica de fígado de ratas wistar diabéticas e tratadas com tamoxifeno

Tamoxifen (TX), a drug used in the treatment of breast cancer, may cause hepatic changes in some patients. The consequences of its use on the liver tissues of rats with or without diabetes mellitus (DM) have not been fully explored. The purpose of this multidisciplinary study was to evaluate the correlation between plasma hepatic enzyme levels and the presence of iron overload in the hepatic tissue of female Wistar rats with or without streptozotocin-induced DM and using TX. Female rats were studied in control groups: C-0 (non-drug users), C-V (sorbitol vehicle only) and C-TX (using TX). DM (diabetic non-drug users) and DM-TX (diabetics using TX) were the test groups. Sixty days after induced DM, blood samples were collected for glucose, alanine aminotransferase (ALT), aspartate aminotransferase (AST) alkaline phosphatase (ALP) and bilirubin measures. Hepatic fragments were processed and stained with hematoxylin and eosin (H&E), Masson s trichrome, Perls. The hepatic iron content was quantified by atomic absorption spectrometry. AST, ALT and ALP levels were significantly elevated in the DM and DM-TX groups, with unchanged bilirubin levels. Liver iron overload using Perls stain and atomic absorption spectrometry were observed exclusively in groups C-TX and DM-TX. There was positive correlation between AST, ALT and ALP levels and microscopic hepatic siderosis intensity in group DM-TX. In conclusion, TX administration is associated with liver siderosis in diabetic and non-diabetic rats. In addition, TX induced liver iron overload with unaltered hepatic function in 2 non-diabetic rats and may be a useful tool for investigating the biological control of iron metabolism

Análise imuno-histoquímica de proteínas relacionadas às respostas Th1, Th2 e Th17 na doença periodontal

Periodontal disease is an inflammatory condition of infectious nature characterized by destruction of protecting and supporting dental tissues. It happens as a response produced by the host when attacked by microorganisms. Several factors are involved in this process. Among them, cytokines are key regulatory molecules in this immune response, playing a role either protective and/or destructive in lesion progression. Thus, this study investigated the immunohistochemical expression of IFN- , GATA-3, IL-17, IL-23, IL-6 and TGF- in gingival tissues of humans, in an attempt to gain a better understanding of the participation of Th1, Th2 and Th17 immune responses in the development of periodontal disease processes. To this end, eighty-two samples of gingival tissues were divided into three groups: Group 1 = 15 (samples of healthy gum tissue as controls), Group 2 = 36 (samples with chronic gingivitis) and Group 3 = 31 (samples with chronic periodontitis). All cases were submitted to morphological analysis from sections stained with hematoxylin and eosin and then subjected to staining by immunohistochemistry using the streptavidin-biotin method. Results showed positive labeling for all proteins. Nonetheless, we observed a greater expression of Th1 cytokines and Th17 cells in group 3. We found statistically significant difference between TGF- expression and the clinical condition of the samples (p=0.02). We conclude that Th1 and Th17 responses may act synergistically in the destructive process of periodontal tissue, overlapping the Th2 response that was also present in these tissues

Análise imuno-histoquímica das proteínas RANK, RANKL e OPG em dentes de ratos reimplantados

The RANK / RANKL / OPG sy stem plays an important role in bone formation and resorption . This finding has been regarded as one of the m ost important advances in the understanding of bone biology with respect to osteoclastogenesis. The aim of this study was to investigate the expression of RANKL / RANK / OPG markers in reimplanted t eeth of rats, and to observe the relationship between the expression of these markers and to oth and bone resorption. Thirty male Wistar rats (Rattus norvegicus albinos) had their maxillary right incisors extrac ted , and were divided into 2 groups according to the period that the extracted teeth were kept in dry air before reimplantation : G1 (n = 15) - 5 minutes , and G2 (n = 15) - 60 minutes . After reimplantation, teeth were analyzed at intervals of 1, 3 and 7 da ys. After these experimental periods, the animals were euthanized. Longitudinal sections with 5μm thick were obtained and stained with Hematoxylin and Eosin for histological analysis , while 3μm thick sections were subjected to immunohistochemical analysis of OPG , RANK and RANKL. The results showed that the RANK / RANKL / OPG system actively participates in both the repair process, as well as tooth and bone resorption . Extr a - alveolar time of 60 minutes before replantation caused minor expressions of RANKL a nd OPG, not influencing the expression of RANK; RANKL immunostaining showed higher in both groups when compared to other biomarkers, participating in all phases of bone and tooth resorption; RANKL was associated to both osteoclastogenesis and c ell ular proliferation , and was expressed in both groups.

Estudo imunoistoquímico do CD34 e podoplanina na doença periodontal

Angiogenesis and lymphangiogenesis are changes that occur due to gingival inflammation caused by microorganisms present in the biofilm, as well as the migration of immune cells and secretion of mediators in the aggressed site. This study aimed to research angiogenesis and lymphangiogenesis in 90 specimens of clinically healthy, with gingivitis and chronic periodontitis gingival tissue biopsies. The histological sections were evaluated by hematoxylin and eosin and the immunohistochemical technique through immunostaining for CD34 and podoplanin. To evaluate the angiogenic and lymphangiogenic indexes we performed a microvessel counting technique. The results showed that there is a correlation between the indexes (p = 0.030), however, we observed that periodontitis showed less lymphatic vessels than clinically healthy gingival tissue (p = 0.016). Podoplanin showed positive staining in the basal layers of the epithelium, and we observed a relationship between immunostaining intensity and the intensity of inflammatory infiltrate, with more intense staining in the presence of severe inflammatory infiltrate (p = 0.033). For this study, we concluded that there are fewer blood vessels in periodontitis compared with clinically healthy gingiva. The signaling present in the inflammatory process and the actual role of gingival blood and lymphatic vasculature are not fully understood, with further studies on angiogenesis and lymphangiogenesis being suggested.

Estudo imunoistoquímico do CD34 e podoplanina na doença periodontal

Angiogenesis and lymphangiogenesis are changes that occur due to gingival inflammation caused by microorganisms present in the biofilm, as well as the migration of immune cells and secretion of mediators in the aggressed site. This study aimed to research angiogenesis and lymphangiogenesis in 90 specimens of clinically healthy, with gingivitis and chronic periodontitis gingival tissue biopsies. The histological sections were evaluated by hematoxylin and eosin and the immunohistochemical technique through immunostaining for CD34 and podoplanin. To evaluate the angiogenic and lymphangiogenic indexes we performed a microvessel counting technique. The results showed that there is a correlation between the indexes (p = 0.030), however, we observed that periodontitis showed less lymphatic vessels than clinically healthy gingival tissue (p = 0.016). Podoplanin showed positive staining in the basal layers of the epithelium, and we observed a relationship between immunostaining intensity and the intensity of inflammatory infiltrate, with more intense staining in the presence of severe inflammatory infiltrate (p = 0.033). For this study, we concluded that there are fewer blood vessels in periodontitis compared with clinically healthy gingiva. The signaling present in the inflammatory process and the actual role of gingival blood and lymphatic vasculature are not fully understood, with further studies on angiogenesis and lymphangiogenesis being suggested.

Prevalence of helicobacter pylori infection in advanced gastric carcinoma

There is substantial evidence that infection with Helicobacter pylori plays a role in the development of gastric cancer and that it is rarely found in gastric biopsy of atrophic gastritis and gastric cancer. On advanced gastric tumors, the bacteria can be lost from the stomach. Aims - To analyze the hypothesis that the prevalence of H.pylori in operated advanced gastric carcinomas and adjacent non-tumor tissues is high, comparing intestinal and diffuse tumors according to Lauren’s classifi cation. Methods - A prospective controlled study enrolled 56 patients from “Hospital Universitário”, Federal University of Rio Grande do Norte, Natal, RN, Brazil, with advanced gastric cancer, treated from February 2000 to March 2003. Immediately after partial gastrectomy, the resected stomach was opened and several mucosal biopsy samples were taken from the gastric tumor and from the adjacent mucosa within 4 cm distance from the tumor margin. Tissue sections were stained with hematoxylin and eosin. Lauren‘s classifi cation for gastric cancer was used, to analyse the prevalence of H. pylori in intestinal or diffuse carcinomas assessed by the urease rapid test, IgG by ELISA and Giemsa staining. H. pylori infected patients were treated with omeprazole, clarithromycin and amoxicillin for 7 days. Follow-up endoscopy and serology were performed 6 months after treatment to determine successful eradication of H. pylori in non-tumor tissue. Thereafter, follow-up endoscopies were scheduled annually. Chi-square and MacNemar tests with 0.05 signifi cance were used. Results - Thirty-four tumors (60.7%) were intestinal-type and 22 (39.3%) diffuse type carcinomas. In adjacent non-tumor gastric mucosa, chronic gastritis were found in 53 cases (94.6%) and atrophic mucosa in 36 patients (64.3%). All the patients with atrophic mucosa were H. pylori positive. When examined by Giemsa and urease test, H. pylori positive rate in tumor tissue of intestinal type carcinomas was higher than that in diffuse carcinomas. In tumor tissues, 34 (60.7%) H. pylori-positive in gastric carcinomas were detected by Giemsa method. H. pylori was observed in 30 of 56 cases (53.5%) in tissues 4 cm adjacent to tumors. This difference was not signifi cant. Eradication of H. pylori in non-tumor tissue of gastric remnant led to a complete negativity on the 12th postoperative month. Conclusions - The data confi rmed the hypothesis of a high prevalence of H. pylori in tumor tissue of gastric advanced carcinomas and in adjacent non-tumor mucosa of operated stomachs. The presence of H. pylori was predominant in the intestinal-type carcinoma