Pleiotrophin在两种胰腺癌细胞系中的表达及意义

发布时间：2018-12-17 13:00

【摘要】：目的研究PTN(Pleiotrophin,多效生长因子)在胰腺癌细胞株Miapaca-2和Capan-1中的表达情况,利用免疫荧光技术、Q-PCR、Western blot法检测PTN mRNA和蛋白的表达,从而筛选较稳定的高表达细胞系,为后期建立小鼠原位胰腺癌模型并通过干扰原位胰腺癌中PTN蛋白的表达,进一步验证PTN与其受体结合是否激活相关信号传导通路在胰腺癌神经浸润发生中的作用提供科学依据。PTN可作为潜在的药物作用靶点,进一步对相关信号通路进行干预,抑或体外合成相应的PTN抑制剂,从而达到下调PTN或完全抑制的目的,阻止肿瘤的进一步生长及胰腺癌神经浸润的发生,为胰腺癌的早期诊断、分期及治疗寻找新途径提供科学依据。方法用DMEM及1640培养液对Miapaca-2和Capan-1细胞进行传代培养,取指数生长期细胞,利用Q-PCR法和Western blot方法检测PTN m RNA和蛋白在胰腺癌细胞株中的表达,并采用免疫荧光化学法检测PTN在胰腺癌细胞中的表达位置。结果Q-PCR结果显示,PTN mRNA在MIA Pa Ca-2细胞中表达明显较高,且与Capan-1细胞相比,MIA Pa Ca-2中PTN m RNA表达量为Capan-1的1.193倍。Western blot结果显示:目的蛋白PTN的分子量大小约为19KDa,内参GAPDH蛋白大小为36KDa。然后应用Image J软件进行半定量分析,结果发现,PTN蛋白在MIA PaCa-2和Capan-1细胞均有表达,且与Capan-1相比,MIA Pa Ca-2细胞系中PTN蛋白的表达更加明显。免疫荧光结果显示PTN的阳性表达呈红色颗粒,主要定位于胞浆,部分胞核亦可见阳性着色。随后我们将Q-PCR、Western blot、免疫荧光结果进行对比,结果表明胰腺癌细胞中PTN在mRNA及蛋白水平表达具有一致性。结论Mia Pa Ca-2细胞中PTN mRNA和蛋白均呈更高表达,主要定位于胞浆中,因此,该细胞可作为良好的研究工具,为进一步建立小鼠原位胰腺癌模型,以研究PTN与受体结合是否促进胰腺癌神经浸润的发生提供依据。
[Abstract]:Objective to study the expression of PTN (Pleiotrophin, multipotent growth factor (PTN (Pleiotrophin,) in pancreatic cancer cell line Miapaca-2 and Capan-1, and to detect the expression of PTN mRNA and protein by Q-PCR Western blot. Thus, a stable high expression cell line was screened to establish a mouse model of pancreatic cancer in situ and to interfere with the expression of PTN protein in pancreatic carcinoma in situ. To further verify whether the binding of PTN to its receptor can activate the role of related signal transduction pathway in the neurogenesis of pancreatic cancer, PTN can be used as a potential drug target to further interfere with the related signal pathway. Or the synthesis of corresponding PTN inhibitors in vitro, so as to down-regulate PTN or complete inhibition, prevent the further growth of tumors and the occurrence of pancreatic cancer neural infiltration, which is the early diagnosis of pancreatic cancer. Staging and treatment to find new ways to provide scientific basis. Methods Miapaca-2 and Capan-1 cells were subcultured with DMEM and 1640 medium. The expression of PTN m RNA and protein in pancreatic cancer cell lines was detected by Q-PCR and Western blot methods. The expression of PTN in pancreatic cancer cells was detected by immunofluorescence method. Results Q-PCR showed that the expression of, PTN mRNA in MIA Pa Ca-2 cells was significantly higher than that in Capan-1 cells. The expression of PTN m RNA in MIA Pa Ca-2 was 1.193 times of that of Capan-1. The results showed that the molecular weight of the target protein PTN was about 19K Daa and the internal reference GAPDH protein size was 36KDa. Then the semi-quantitative analysis was carried out by Image J software. The results showed that PTN protein was expressed in both MIA PaCa-2 and Capan-1 cells, and the expression of PTN protein in, MIA Pa Ca-2 cell line was more obvious than that in Capan-1 cell line. The results of immunofluorescence showed that the positive expression of PTN was red granules, mainly located in the cytoplasm, and the positive staining was also seen in some nuclei. The results showed that the expression of PTN in pancreatic cancer cells was consistent at the level of mRNA and protein. Conclusion the expression of PTN mRNA and protein in Mia Pa Ca-2 cells is higher, which is mainly located in the cytoplasm. Therefore, this cell can be used as a good tool for the establishment of in situ pancreatic cancer model in mice. To study whether the binding of PTN to receptor can promote the neurogenesis of pancreatic cancer.
【学位授予单位】：河南科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.9

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