Importance of syndecan-4 and syndecan -2 in osteoblast cell adhesion and survival mediated by a tissue transglutaminase-fibronectin complex

Tissue transglutaminase (TG2) has been identified as an important extracellular crosslinking enzyme involved in matrix turnover and in bone differentiation. Here we report a novel cell adhesion/survival mechanism in human osteoblasts (HOB) which requires association of FN bound TG2 with the cell surface heparan sulphates in a transamidase independent manner. This novel pathway not only enhances cell adhesion on FN but also mediates cell adhesion and survival in the presence of integrin competing RGD peptides. We investigate the involvement of cell surface receptors and their intracellular signalling molecules to further explore the pathway mediated by this novel TG-FN heterocomplex. We demonstrate by siRNA silencing the crucial importance of the cell surface heparan sulphate proteoglycans syndecan-2 and syndecan-4 in regulating the compensatory effect of TG-FN on osteoblast cell adhesion and actin cytoskeletal formation in the presence of RGD peptides. By use of immunoprecipitation and inhibitory peptides we show that syndecan-4 interacts with TG2 and demonstrate that syndecan-2 and the a5ß1 integrins, but not a4ß1 function as downstream modulators in this pathway. Using function blocking antibodies, we show activation of a5ß1 occurs by an inside out signalling mechanism involving activation and binding of protein kinase PKCa and phosphorylation of focal adhesion kinase (FAK) at Tyr861 and activation of ERK1/2.

Inhibition of Neurotoxic Secretory Phospholipases A(2) Enzymatic, Edematogenic, and Myotoxic Activities by Harpalycin 2, an Isoflavone Isolated from Harpalyce brasiliana Benth

Secretory phospholipases A(2) (sPLA(2)) exert proinflammatory actions through lipid mediators. These enzymes have been found to be elevated in many inflammatory disorders such as rheumatoid arthritis, sepsis, and atherosclerosis. The aim of this study was to evaluate the effect of harpalycin 2 (Har2), an isoflavone isolated from Harpalyce brasiliana Benth., in the enzymatic, edematogenic, and myotoxic activities of sPLA2 from Bothrops pirajai, Crotalus durissus terrificus, Apis mellifera, and Naja naja venoms. Har2 inhibits all sPLA(2) tested. PrTX-III (B. pirajai venom) was inhibited at about 58.7%, Cdt F15 (C. d. terrificus venom) at 78.8%, Apis (from bee venom) at 87.7%, and Naja (N. naja venom) at 88.1%. Edema induced by exogenous sPLA(2) administration performed in mice paws showed significant inhibition by Har2 at the initial step. In addition, Har2 also inhibited the myotoxic activity of these sPLA(2)s. In order to understand how Har2 interacts with these enzymes, docking calculations were made, indicating that the residues His48 and Asp49 in the active site of these enzymes interacted powerfully with Har2 through hydrogen bonds. These data pointed to a possible anti-inflammatory activity of Har2 through sPLA(2) inhibition.

DEFINIÇÃO DE CRITÉRIOS PARA A DELIMITAÇÃO DA ZONA DE AMORTECIMENTO DE UM SETOR DO PARQUE ESTADUAL DA CANTAREIRA SECCIONADO PELA RODOVIA FERNÃO DIAS (BR- 381), SÃO PAULO, BRASIL

A delimitação da zona de amortecimento de um setor do Parque Estadual da Cantareira, seccionado pela rodovia Fernão Dias (BR-381), foi elaborado com base na integração dos estudos de uso da terra e fragilidade ambiental. Também foram utilizados os instrumentos de planejamento territorial e as legislações de cunhos ambiental e urbano incidentes na área de estudo. A Zona de Amortecimento engloba remanescentes florestais expressivos e áreas com reflorestamentos situados, predominantemente, em terrenos com Fragilidade Potencial Alta. De maneira geral, as áreas urbanizadas ou em processo de expansão urbana localizadas nos limites ou próximos ao Parque, foram incluídas na zona de amortecimento com o intuito de evitar o adensamento da ocupação e o parcelamento ainda maior do solo urbano. Quanto aos campos antrópicos/pastagem situados em áreas de preservação ambiental deverão ser promovidas medidas para a recuperação da vegetação nativa. As pedreiras ativas e os aterros sanitários foram mantidos na zona de amortecimento com o propósito de intensificar o controle de suas atividades altamente impactantes ao meio ambiente. Este trabalho procura desenvolver metodologia e estabelecer critérios ambientais e legais que possibilitam delimitar a zona de amortecimento de unidades de conservação, sobretudo aquelas sujeitas a pressões provocadas pela expansão urbana.