Ação intrácrina da angiotensina II em células mesangiais: localização subcelular dos receptores AT1 e AT2 e comunicação mediada por exossomos

• Ano de defesa: 2016
• Autor(a) principal: Novaes, Antonio da Silva [UNIFESP]
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• dARK ID: ark:/48912/0013000025zxd
• Idioma: por
• Instituição de defesa: Universidade Federal de São Paulo (UNIFESP)
• Palavras-chave em Português: at1 receptor; at2 receptor; Exosomes; Mesangial cells; Receptores at1; Receptores at2; Exossomos; Células mesangiais
• Link de acesso: https://sucupira.capes.gov.br/sucupira/public/consultas/coleta/trabalhoConclusao/viewTrabalhoConclusao.jsf?popup=true&id_trabalho=4697551; http://repositorio.unifesp.br/handle/11600/46276

Introduction: Currently is discussed the biological relevance of the intracellular renin angiotensin system (RAS) mediating the intracrine actions of angiotensin II (Ang II). The presence of AT1 and AT2 receptors in the intracellular compartment may mediate the nonclassical effects Ang II, including regulation of gene expression of Ang II target genes such as pro-fibrotic cytokines. Intracrine effects of Ang II can be mediated by peptides transfer to other cells via exosomes. Objectives: 1) to evaluate the intracellular distribution of AT1 and AT2 receptors in human mesangial cells (HMC) and its pathophysiological role under stimulus with high glucose concentration (HG); 2) to verify whether exosomal signaling initiated in HCM is responsive to HG stimulus. Methods: The distribution of AT1 and AT2 like receptors was determined by immunofluorescence in whole cell and in intact isolated nuclei. Molecular weight (MW) of both receptors were confirmed by Western blotting (WB) of nuclear extract proteins. Intracellular synthesis of Ang II was stimulated by HG and the intracrine effect of Ang II was analyzed on fibronectin gene expression in the presence and absence of candesartan which discriminates the intra and extracellular effects of Ang II. The exosomal contents of angiotensinogen and renin were analyzed by Western Blotting. Exosomes obtained from control (C-Ex) and HG exposed HMC (HG-Ex) were used to evaluate whether they are able to modify the function of target control HMC on expressions of fibronectin, angiotensinogen, renin, AT1, AT2 and proliferation. Results: AT1 and AT2 were found in whole cell including nuclear membrane and in the membrane of the isolated nuclei. WB analysis confirmed the MW of both receptors in nuclei. Labelled exogenous Ang II bound to nuclear membrane. HG stimulated cells presented higher levels of fibronectin and this effect was not inhibited by candesartan, suggesting an intracellular effect of Ang II. HG stimulus induced a change in the amount, but not in the size of exosomes. Exosomes derived from control HMC contained angiotensinogen and renin proteins, which expression were increased in the exosomes obtained from HMC stimulated by HG. The expression levels of fibronectin and angiotensinogen were higher in control HCM treated with HG-Ex compared with those treated with C-Ex, indicating that HG-Ex can modify the function of normal HMC. Conclusion: Nuclear AT1 and AT2 receptors are active in cultured HMC. The exosomes derived from HG stimulated HMC modified the function of target control cells, suggesting that the intercellular communication through the exosomes may have pathophysiological implications in the diabetic kidney.