TRPM8离子通道在胰腺癌中高表达的临床意义

发布时间：2018-02-14 23:55

本文关键词： 胰腺癌 TRPM8 Western Blot 膜片钳 CCK8　出处：《吉林大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:1.TRPM8作为临床生物标记物,为准确判断胰腺癌的恶性度及预后提供指导。2.TRPM8作为潜在的药物作用靶点,将来可能通过下调TRPM8的表达,来抑制胰腺癌细胞增殖扩撒,从而最终可以发现有效的胰腺癌个性化治疗的新靶点和新思路。方法:1.购买北京龙迈达斯科技开发有限公司胰腺癌组织77例,正常组织30例,用免疫组织化学技术对标本进行实验,并对结果进行统计分析。2.购买正常胰腺细胞株HPCY-5和胰腺癌细胞株Bx PC-3、PANC-1并进行细胞培养,通过QPCR技术测定TRPM8离子通道m RNA的表达情况。3.用Western bot技术检测细胞株HPCY-5、Bx PC-3、PANC-1通道蛋白的表达情况。4.用细胞膜片钳技术检测转染si RNA前后胰腺癌细胞株PANC1细胞膜电流的改变。5.用CCK8试剂盒检测转染si RNA前后胰腺癌细胞株Bx PC-3、PANC-1,分析细胞增殖能力的改变结果:免疫组化实验、Western Blot实验都提示TRPM通道蛋白在胰腺癌细胞中成高表达状态,且免疫组化结果提示肿瘤细胞中TRPM8通道蛋白的高表达与肿瘤的分化程度具有相关性(中低分化vs高分化P0.05,具有统计学意义)。QPCR检测胰腺癌肿瘤细胞株Bx PC-3、PANC-1中TRPM8 m RNA相对于正常细胞株HPCY-5呈高表达,与免疫组化和Western Blot结果相一致,且P0.05具有统计学意义。膜片钳技术测定薄荷醇激活的TRPM8离子通道电流明显增大。CCK8法提示TRPM8通道基因敲除的癌细胞株增殖能力出现明显减弱,证明TRPM8通道基因的表达对癌细胞的增殖具有一定影响。
[Abstract]:Objective: 1. TRPM8, as a clinical biomarker, provides guidance for accurately judging the malignancy and prognosis of pancreatic cancer. 2. TRPM8 is a potential drug target. It may inhibit proliferation and diffusion of pancreatic cancer cells by down-regulating the expression of TRPM8 in the future. In the end, we can find new targets and new ideas for effective personalized treatment of pancreatic cancer. Methods: 1. Buy 77 cases of pancreatic cancer tissue and 30 cases of normal tissue from Beijing Longmeidas Technology Development Co., Ltd. The results were statistically analyzed by immunohistochemical technique. 2. The normal pancreatic cell line HPCY-5 and the pancreatic cancer cell line Bx PC-3 PANC-1 were purchased and cultured. The expression of m RNA in TRPM8 ion channel was detected by QPCR. The expression of Western bot was detected in HPCY-5 BX PC-3PANC-1 cell line. 4. Cell patch clamp technique was used to detect the cell membrane current of PANC1 cell line before and after transfection of si RNA. CCK8 kit was used to detect the proliferation of pancreatic cancer cell line BxPC-3pPANC-1 before and after transfection of si RNA, and the results showed that the expression of TRPM channel protein in pancreatic cancer cells was high. The results of immunohistochemistry indicated that the high expression of TRPM8 channel protein in tumor cells was correlated with the degree of tumor differentiation (medium and low differentiation vs highly differentiated P0.05, and the detection of TRPM8 m RNA in pancreatic cancer cell line BxPC-3PANC-1 by QPCR was statistically significant. Compared with the normal cell line, the expression of HPCY-5 was higher than that of normal cell line. The results were consistent with the results of immunohistochemistry and Western Blot (P0.05). Patch clamp technique showed that the current of TRPM8 ion channel activated by menthol was significantly increased. CCK8 method indicated that the proliferation ability of TRPM8 channel gene knockout cancer cell line was significantly decreased. The results showed that the expression of TRPM8 channel gene had a certain effect on the proliferation of cancer cells.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.9

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