Tumor Necrosis Factor alpha and Interleukin-1 beta Modulate Calcium and Nitric Oxide Signaling in Mechanically Stimulated Osteocytes

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Er : YAG laser: Antimicrobial effects in the root canals of dogs' teeth with pulp necrosis and chronic periapical lesions

Objective: Our goal in this study was to evaluate the antimicrobial effect of Er:YAG laser applied after biomechanical preparation of the root canals of dog's teeth with apical periodontitis. Background Data: Various in vitro studies have reported effective bacterial reduction in infected root canals using Er:YAG laser. However, there is no in vivo research to support these results. Methods: Forty root canals of dogs' premolar teeth with pulp necrosis and chronic periapical lesions were used. An initial microbiological sample was taken, and after biomechanical preparation was carried out, a second microbiological sample was taken. The teeth were divided into two groups: Group I-biomechanical preparation was taken of root canals without Er:YAG laser application; Group II-biomechanical preparation was taken of root canals with Er:YAG laser application using 140-mj input, 63-mJ output/15 Hz. After coronal sealing, the root canals were left empty for 7 days at which time a third microbiological sample was taken. The collected material was removed from the root canal with a #40 K file and placed in transport media. It was serially diluted and seeded on culture dishes selective for anaerobes, aerobes, and total streptococci. Colony-forming units per milliliter (CFU/mL) were counted. Results: Groups I and II showed an increase of CFU/mL for all microorganisms 7 days after treatment, being statistically significant for anaerobes in Group I and for anaerobes and total streptococci in Group II. When comparing CFU/mL of Groups I and II, there was a statistically significant increase after 7 d for total streptococci in Group II. Conclusion: Er:YAG laser applied after biomechanical preparation did not reduce microorganisms in the root canal system.

Chronic heart failure-induced skeletal muscle atrophy, necrosis, and changes in myogenic regulatiory factors

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Development of the insulin secretion mechanism in fetal and neonatal rat pancreatic B-cells: response to glucose, K+, theophylline, and carbamylcholine

We studied the development of the insulin secretion mechanism in the pancreas of fetal (19- and 21-day-old), neonatal (3-day-old), and adult (90-day-old) rats in response to stimulation with 8.3 or 16.7 mM glucose, 30 mM K+, 5 mM theophylline (Theo) and 200 µM carbamylcholine (Cch). No effect of glucose or high K+ was observed on the pancreas from 19-day-old fetuses, whereas Theo and Cch significantly increased insulin secretion at this age (82 and 127% above basal levels, respectively). High K+ also failed to alter the insulin secretion in the pancreas from 21-day-old fetuses, whereas 8.3 mM and 16.7 mM glucose significantly stimulated insulin release by 41 and 54% above basal levels, respectively. Similar results were obtained with Theo and Cch. A more marked effect of glucose on insulin secretion was observed in the pancreas of 3-day-old rats, reaching 84 and 179% above basal levels with 8.3 mM and 16.7 mM glucose, respectively. At this age, both Theo and Cch increased insulin secretion to close to two-times basal levels. In islets from adult rats, 8.3 mM and 16.7 mM glucose, Theo, and Cch increased the insulin release by 104, 193, 318 and 396% above basal levels, respectively. These data indicate that pancreatic B-cells from 19-day-old fetuses were already sensitive to stimuli that use either cAMP or IP3 and DAG as second messengers, but insensitive to stimuli such as glucose and high K+ that induce membrane depolarization. The greater effect of glucose on insulin secretion during the neonatal period indicates that this period is crucial for the maturation of the glucose-sensing mechanism in B-cells.

Detection and partial characterization of a carlavirus causing stem necrosis of soybean in Brazil

No ano agrícola de 2000/2001, ocorreu um surto de nanismo e necrose da haste em soja (Glycine max) plantada em duas áreas do Brasil Central e na safra seguinte, esta anomalia foi constatada em outras regiões produtoras, mesmo distantes mais de 2.000 km de onde fora inicialmente constatada. Estudos envolvendo ensaios de transmissão (enxertia, mecânica e insetos vetores), microscopia eletrônica, purificação, sorologia e ensaios moleculares indicaram que a enfermidade foi causada por um carlavirus transmitido por mosca branca, possivelmente relacionado ao Cowpea mild mottle virus (CpMMV).

Myocyte necrosis is the basis for fibrosis inrenovascular hypertensive rats

The pathogenesis of fibrosis and the functional features of pressure overload myocardial hypertrophy are still controversial. The objectives of the present study were to evaluate the function and morphology of the hypertrophied myocardium in renovascular hypertensive (RHT) rats. Male Wistar rats were sacrificed at week 4 (RHT4) and 8 (RHT8) after unilateral renal ischemia (Goldblatt II hypertension model). Normotensive rats were used as controls. Myocardial function was analyzed in isolated papillary muscle preparations, morphological features were defined by light microscopy, and myocardial hydroxyproline concentration (HOP) was determined by spectrophotometry. Renal artery clipping resulted in elevated systolic arterial pressure (RHT4: 178 ± 19 mmHg and RHT8: 194 ± 24 mmHg, P<0.05 vs control: 123 ± 7 mmHg). Myocardial hypertrophy was observed in both renovascular hypertensive groups. The myocardial HOP concentration was increased in the RHT8 group (control: 2.93 ± 0.38 µg/mg; RHT4: 3.02 ± 0.40 µg/mg; RHT8: 3.44 ± 0.45 µg/mg of dry tissue, P<0.05 vs control and RHT4 groups). The morphological study demonstrated myocyte necrosis, vascular damage and cellular inflammatory response throughout the experimental period. The increased cellularity was more intense in the adventitia of the arterioles. As a consequence of myocyte necrosis, there was an early, local, conjunctive stroma collapse with disarray and thickening of the argyrophilic interstitial fibers, followed by scarring. The functional data showed an increased passive myocardial stiffness in the RHT4 group. We conclude that renovascular hypertension induces myocyte and arteriole necrosis. Reparative fibrosis occurred as a consequence of the inflammatory response to necrosis. The mechanical behavior of the isolated papillary muscle was normal, except for an early increased myocardial passive stiffness

Ischemia and reperfusion in skin flaps: effects of mannitol and vitamin C in reducing necrosis area in a rat experimental model

OBJETIVO: Neste trabalho foi padronizado modelo experimental de isquemia e reperfusão em retalho cutâneo em ratos no qual estudou-se possibilidade de uma solução antioxidante, composta por Ringer lactato, vitamina C e manitol de reduzir a área de necrose. MÉTODOS: O modelo consistiu de levantamento de retalho cutâneo axial de 6,0 x 3,0cm, submetido à isquemia de 8 horas e reperfusão de 7 dias. Os animais foram divididos em quatro grupos: grupos S1, S2 (10 animais cada), C e T (14 animais cada). Nos grupos S1 e S2 todos os procedimentos dos demais grupos foram efetuados, exceto a isquemia e reperfusão: S1 recebeu apenas Ringer lactato e S2 a solução antioxidante. Os grupos C e T foram submetidos à isquemia. O grupo C recebeu somente Ringer lactato e o grupo T a solução antioxidante. No 7(0) dia de pós-operatório as áreas de necrose e pele viável do retalho foram delineadas em decalque de acetato, os quais foram por sua vez analisados em sistema computadorizado KS-300 (Carl Zeiss). RESULTADOS: A análise estatística mostrou que não houve diferenças significativas entre o grupo tratado e controle quanto à área de necrose. CONCLUSÃO: Concluiu-se que o modelo experimental é consistente, determinando área de necrose limitada e uniforme nos animais não tratados e que as drogas usadas, nessa posologia e modo de aplicação, não foram efetivas em diminuir a área de necrose no modelo experimental em questão.

LOW-PROTEIN DIET IMPAIRS GLUCOSE-INDUCED INSULIN-SECRETION FROM AND CA-45 UPTAKE BY PANCREATIC RAT ISLETS

Glucose-induced insulin secretion rom and Ca-45 uptake by isolated pancreatic islets, derived from rats fed with normal (NPD) or low protein diet (LPD), were studied. Insulin secretion from both types of islets in response to increasing concentrations of glucose followed an S-shaped pattern. However, basal secretion observed at substimulatory concentrations of glucose (0-5.6 mM), as well as maximal release, obtained at 16.7 mM or higher glucose concentrations were significantly reduced in islets from LPD. Furthermore, in LPD rat islets, the dose-response curve to glucose was clearly shifted to the right compared with NPD islets, with the half-maximal response occurring at 8.5 and 14.4 mM glucose for NPD and LPD islets, respectively. In islets from NPD rats, the Ca-45 content, after 5 or 90 min in the presence of 8.3 mM glucose, was higher than that observed for islets kept at 2.8 mM glucose and increased further at 16.7 mM glucose. After 5 min of incubation, the Ca-45 uptake by LPD islets in the presence of 8.3 mM glucose was slightly higher than basal values (2.8 mM glucose); however, no further increase in the Ca-45 uptake was noticed at 16.7 mM glucose. In LPD islets a significant increase in Ca-45 uptake over basal values was registered only at 16.7 mM glucose, after 90 min of incubation. These data indicate that the poor secretary response to glucose observed in islets from LPD rats may be related to a defect in the ability of glucose to increase Ca2+ uptake and/or to reduce Ca2+ efflux from beta-cells.

AN ULTRASTRUCTURAL-STUDY ON THE ENDOCRINE PANCREAS OF CAIMAN-LATIROSTRIS, WITH SPECIAL REFERENCE TO PANCREATIC MOTILIN CELLS

Pancreatic endocrine cells of Caiman latirostris were investigated by electron microscopy using conventional and immunocytochemical methods. Ultrastructurally, four types of endocrine cells were classified according to the morphology of their secretory granules. Three types of endocrine cells were identified as either glucagon, insulin or somatostatin cells by the presence of such characteristic granules well established in mammals. The remaining endocrine cell type could not be classified by its ultrastructural features alone.Immunocytochemical observations confirmed the ultrastructural classification of glucagon, insulin and somatostatin cells. In addition, endocrine cells immunoreactive for either pancreatic polypeptide (PP) or motilin were identified. Morphometric analysis of PP- and motilin-immunoreactive granules demonstrated that they were the most polymorphous and smallest granules among the pancreatic endocrine cell granules. Although both PP and motilin granules closely resemble each other, motilin granules were smaller in size and more spherical in shape than PP granules.

IMMUNOHISTOCHEMICAL CHARACTERIZATION OF PROBABLE INTRAVASCULAR HEMATOPOIESIS IN THE VASA RECTAE OF THE RENAL MEDULLA IN ACUTE TUBULAR-NECROSIS

Immunohistochemistry was applied to identify the nature of the nucleated cells that accumulate in the vasa rectae of the corticomedullary junction in acute tubular necrosis. In all 6 cases studied, there were intravascular cells that reacted with monoclonal antibodies to erythroblast, macrophages, myeloid cells, T and B lymphocytes and rave megakaryocytes. The findings are consistent with the occurrence of intravascular haematopoiesis in the renal medulla in acute tubular necrosis.

HIGH DIETARY CARBOHYDRATE AND PANCREATIC LESION

The ability of high dietary carbohydrate to induce acute pancreatitis was investigated in groups of 16,2 1-day and 15-month old rats fed different carbohydrate diets for 30 days. Significantly increased levels of serum amylase (2-fold), phospholipids (50%),phosphorus (2-fold), and lipoperoxides (8-fold) were observed in 15-month old rats fed a high-carbohydrate diet, compared to rats fed a diet with normal carbohydrate levels, indicating peroxidation of membrane lipids which caused final cell death and pancreatic lesion. Serum Cu-Zn superoxide dismutase activity was not altered. Daily administration of bovine Cu-Zn superoxide dismutase conjugated with polyediylene glycol prevented the serum level alterations and pancreatic lesions, indicating that the superoxide radical has a role in dietary carbohydrate-induced acute pancreatitis. No biochemical changes were observed in rats in which treatment was initiated on the 21st day of life indicating that this is an age-related lesion.

EARLY ACTIVATION OF SPLENIC MACROPHAGES BY TUMOR-NECROSIS-FACTOR-ALPHA IS IMPORTANT IN DETERMINING THE OUTCOME OF EXPERIMENTAL HISTOPLASMOSIS IN MICE

Experimental infection of animals with Histoplasma capsulatum caused a massive macrophage infiltration into the spleen and induced the production of tumor necrosis factor alpha (TNF-alpha) locally. The cytokine was also produced in vitro by peritoneal exudate macrophages exposed to a large inoculum of yeast cells. Depletion of the cytokine by injection of polyclonal sheep anti-TNF-alpha antibody was detrimental to sublethally infected mice. Fungous burdens in the spleens of TNF-alpha-depleted mice were higher than they were in the infected control mice at days 2, 7, and 9 after infection, and the antibody-treated animals succumbed to the infection. Histopathological study of spleen sections revealed that splenic macrophages were not able to control proliferation of intracellular yeasts as a result of TNF-alpha depletion. It seems that TNF-alpha plays a role in early activation of splenic macrophages which is important in controlling the outcome of an infection.

Differential regulation of the release of tumor necrosis factor-alpha and of eicosanoids by mast cells in rat airways after antigen challenge

Background: Rat trachea display a differential topographical distribution of connective tissue mast cells (CTMC) and mucosal mast cells (MMC) that may imply regional differences in the release of allergic mediators such as tumor necrosis factor-alpha (TNF-alpha) and eicosanoids.Aim: To evaluate the role of CTMC and MMC for release of TNF-alpha and eicosanoids after allergenic challenge in distinct segments of rat trachea.Materials and methods: Proximal trachea ( PT) and distal trachea (DT) from ovalbumin (OVA)-sensitized rats, treated or not with compound 48/80 ( 48/80) or dexamethasone, were incubated in culture medium. After OVA challenge, aliquots were collected to study release of TNF-alpha and eicosanoids.Results: Release of TNF-alpha by PT upon OVA challenge peaked at 90 min and decayed at 6 and 24 h. Release from DT peaked at 30-90 min and decayed 6 and 24 h later. When CTMC were depleted with 48/80, OVA challenge exacerbated the TNF-alpha release by PT at all time intervals, while DT exacerbated TNF-alpha levels 6 and 24 h later only. Dexamethasone reduced TNF-alpha production after 90 min of OVA challenge in PT and at 3 and 6h in DT. OVA challenge increased prostaglandin D-2 in DT and leukotriene B-4 in both segments but did not modify prostaglandin E-2 and leukotriene C-4 release.Conclusion: OVA challenge induces TNF-alpha release from MMC, which is negatively regulated by CTMC. The profile of TNF-alpha and eicosanoids depends on the time after OVA challenge and of the tracheal segment considered.

TOXIC EFFECTS OF WATER EUTROPHICATION ON PANCREATIC, HEPATIC AND OSTEOGENIC TISSUES OF RATS

Pollution, industrial solvents, concentrations of metals and other environmental agents are widely related to biochemicals values which are used in disease diagnosis of environmental toxicity. A rat bioassay validated for the identification of toxic effects of eutrophication revealed increased serum activities of amylase, alanine transaminase (BLT) and alkaline phosphatase (ALP) in rats that received algae, filtered water and nickel or cadmium from drinking water. Serum Cu-Zn superoxide dismutase activity decreased from its basal level of 40.8 +/- 2.3 to 26.4 U/mg protein, at 7 days of algae and at 48 hr of nickel and cadmium water ingestion. The observation that lipoperoxide concentration was not altered in rats treated with filtered water, while amylase, ALT and ALP were increased in these rats and in those treated with nickel or cadmium, indicated that pancreatic, hepatic and osteogenic lesions by eutrophication were not related to superoxide radicals, and might be due to a novel toxic environmental agent found in filtered and non-filtered algae water.