抵抗素在膀胱癌发生发展中的作用及机制研究

Bladder cancer is the most common cancer of genitourinary system in China, morbidity and mortality account for the first of genitourinary tumors. The survival rate of patients was affected by the high recurrence, progression and distant metastasis rates. Recently, studies have shown that the adipocytokine family such as leptin, adiponectin，resistin and visfatin associated with tumor genesis and development are closely related. Resistin is an adipocytokine with immunomodulatory function, which play an important role in the regulation of tumor cell apoptosis. Resistin is involved in tumor genesis and development possibly through the activation of PI3K, MAPK or NF-κB pathway. Our previous work have shown that bladder cancer tissue resistin expression was significantly increased, suggested that there is closely related about resistin and bladder cancer genesis. Our team intends to work at the DNA genetic variation, subcellular protein position, the mRNA and protein expression levels to reveal the relationship between resistin and bladder cancer. The expression of resistin is effected by plasmid transfection and RNA interference, research the cell survival rate and signal pathway. Explore the relationship of regulation resistin expression and bladder cancer genesis or development or metastasis.

膀胱癌是我国泌尿生殖系统最常见的肿瘤，发病率及死亡率均占泌尿生殖系统肿瘤首位，其较高复发率、进展率及远处转移率严重威胁患者生存率。近年来研究表明，脂肪细胞因子家族如瘦素、脂联素、抵抗素及内脏脂肪素等与肿瘤的发生发展关系密切。抵抗素（Resistin）是具有免疫调节功能的脂肪细胞因子，在肿瘤细胞凋亡调控中发挥着重要作用，可能是通过激活PI3K、MAPK或NF-κB通路而参与肿瘤的发生发展。我们的前期研究证明膀胱癌组织中Resistin表达明显增加，提示Resistin与膀胱癌的发生发展密切相关。课题组拟从DNA遗传变异、蛋白亚细胞定位、mRNA与蛋白表达水平等角度，深入研究Resistin与膀胱癌发生、发展、侵袭与转移的关系及分子机制。采用质粒转染及RNA干扰技术干预Resistin的表达，研究其对膀胱癌细胞凋亡或存活的影响及其信号通路，探索干预Resistin表达调控与膀胱癌发生发展的关系

膀胱癌是我国泌尿生殖系统最常见的肿瘤之一，其发病率及死亡率均占泌尿生殖系统肿瘤首位。抵抗素（resistin）不但参与代谢调节，而且作为一种新型的免疫调节因子参与了机体的大部分炎症反应。本项目研究采用TaqMan SNP分型技术与方法，在膀胱癌患者组及正常对照组样本中对SNP位点rs1862513及rs10401670进行了群体遗传性研究，发现rs10401670位点T等位基因显著降低膀胱癌发病风险（P = 0.03）,CT/TT基因型显著降低膀胱癌发病风险（P = 0.018）；SNP位点rs10401670的CT杂合子携带者早发膀胱癌的风险增加1.97倍。SNP位点rs1862513的CG杂合子比非吸烟患膀胱癌风险增加1.73倍；单因素分析结果表明，SNP位点rs10401670及rs1862513均与膀胱癌患者总体生存、无进展生存不相关。Cox回归分析结果发现，MIBC患者中rs1862513位点CG杂合子携带者肿瘤进展风险显著减低（P = 0.04）；rs10401670位点T等位基因携带者（CT/TT基因型）死亡风险显著增加2.45倍（P = 0.03），特别是TT纯合子，死亡风险增高到3倍（P = 0.02）；ELISA试剂盒检测血浆resistin表达，发现膀胱癌组血浆resistin表达水平明显低于正常对照组，有意义的是，与手术前相比，膀胱癌患者术后血浆resistin蛋白表达上调。免疫组织化学及Western blot检测膀胱癌组织及对照组织中resistin蛋白表达，均发现癌组织中resistin表达显著增高，qPCR检测发现组织中RETN基因mRNA表达显著升高。突变筛查结果在本研究所涉及的332例膀胱癌患者中未筛查到新突变位点。野生型RETN-pcDNA™3.1(+)质粒（WT）及shRNA干扰质粒分别转染T24细胞，干预RETN表达，采用qPCR技术并结合Western Blot，探讨RETN与膀胱癌发生发展的分子机制，结果发现：shRNA干扰质粒组与WT过表达组比较，shRNA成功抑制了resistin表达，且p38、p44及NF-κB蛋白表达显著降低，提示resistin可能通过NF-κB及MAPK信号通路参与膀胱癌的发生发展。本项目研究的顺利完成，为RETN及相关基因的深入研究以及膀胱癌发生发展分子机制研究提供了新的思路与方向