Identificação e quantificação das isoformas da heparanase em plasma de pacientes com carcinomas gatrointestinais

• Autor(a) principal: Origassa, Clarice Silvia Taemi [UNIFESP]
• Data de Publicação: 2009
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Repositório Institucional da UNIFESP
• Texto Completo: http://repositorio.unifesp.br/handle/11600/9702
• topic: Carcinoma; Heparam sulfato; Plasma; Heparanases; Carcinoma; Heparitin sulfate; Plasma

Heparanase-1 (HPA1) is an endo-β-glucuronidase that degrades intradisaccharides glycosidic linkage between hexosamine and glucuronic acid into heparan sulfate chains, present on extracellular matrix and surface proteoglycans. HPA1 gene is located at 4q21.3 chromossome. Oligosaccharides generated by HPA1 are involved in cell proliferation, angiogenesis and cell diferentiation related with tumor development and metastasis. Heparanase-2 (HPA2) is encoded by 10q23-24 chromossome. There are three HPA2 isoforms containing 480, 534 and 592 aminoacids. All HPA2 isoforms have shown that they are intracellular, membrane associated proteins, containing C-terminal domain to the cytoplasm and do not present enzymatic activity. The objective of the present study is to identify and quantify HPA1 (isoforms 50 kDa and 65 kDa), and HPA2 isoforms expression in the plasma of gastrointestinal carcinoma patients, compared with individuals that do not present neoplasia (control group). Plasmatic proteins were identified by polyacrilamide gel electrophoresis (10% SDS-PAGE) and transferred to Hybond-CE membrane, incubated with primary anti-HPA1 H-80 or anti-HPA2 C-17 and developed using secondary antibody conjugated with HRP IgG peroxidase. Heparanases isoforms were quantified by densitometry (Scion Image) and confirmed by real time RT-PCR. Statistic analysis were performed using SPSS 13.0 program (SPSS, Chicago, IL). The results have shown that both HPA1 isoforms (HPA1 50 kDa and HPA1 65 kDa) and HPA2 isoforms were significantly increased in gastrointestinal carcinoma patients plasma, compared with control group. Real time analysis of HPA1 and HPA2 expression in the mononuclear fraction of blood demonstrated an increase HPA1 and HPA2 expression in gastrointestinal carcinoma patients that corroborates with plasma analysis. The obtained results demonstrated that both HPA1 and HPA2 have a fundamental role in gastrointestinal carcinogenesis and, therefore, to understand their physiological function it could help the development of possible new antitumor therapies.