【作者】 王慧；

【导师】 沈啸洪；

【作者基本信息】 南开大学 ， 生理学， 2018， 硕士

【摘要】 背景:胰腺癌是一种死亡率非常高的胃肠道消化肿瘤,在全球和癌症相关的死亡中,胰腺癌排第四位,而且预后效果极差,5年生存率小于6%。而因为癌细胞无限的生长增殖,转化和转移,所以癌细胞很难消除。但是目前为止,胰腺癌细胞迅速生长增殖以及凋亡过程的发展机制尚不清楚。目的:探究趋化因子Fractalkine(FKN,CX3CL1)及其受体CX3CR1对胰腺癌细胞生长增殖以及凋亡过程的影响,并探讨FKN调控胰腺癌细胞增殖凋亡过程的分子机制,确定一种与胰腺癌病情进展及预后相关的特异性蛋白质,进而为早期胰腺癌的靶点治疗提供依据。方法:首先我们收集了切除并经病理确诊的胰腺癌及癌旁组织标本25例,并对收集的胰腺癌组织及癌旁组织标本中的FKN及CX3CR1进行免疫组化染色(Immunohistochemistry)分析,确定它们的表达量。然后,选取了 3种胰腺癌细胞系 Capan-2,MIA PaCa-2 和 Aspc-1,通过免疫印迹(Western Blotting)检测胰腺癌细胞内FKN及CX3CR1的表达情况。同时又通过免疫印迹、CCK-8、流式细胞术以及免疫荧光等技术检测过表达FKN质粒、外源蛋白FKN及FKN的小干扰RNAs对胰腺癌细胞增殖凋亡以及细胞周期的影响。最后,通过PI3K以及NF-κB/p65的抑制剂阻断该信号通路,进而研究FKN影响胰腺癌增殖凋亡过程的生理机制。结果:免疫组化实验和免疫印迹实验结果显示,FKN和CX3CR1在胰腺癌组织和癌旁组织中都表达,但是在癌组织中的表达显著高于癌旁组织。在胰腺癌细胞Capan-2,MIA PaCa-2 和 Aspc-1 中,通过 Western Blotting 实验发现 FKN 和CX3CR1依然是高表达。流式细胞技术、免疫荧光共聚焦实验以及免疫印迹实验结果显示,FKN可以促进胰腺癌细胞的增殖并且抑制胰腺癌细胞的凋亡,同时还会促进细胞周期中G1期向S期的转化,但是加入PI3K或NF-κB/p65抑制剂后会影响FKN对癌细胞的作用。结论:该研究证实,FKN与它的受体CX3CR1结合以后通过AKT/NF-κB/p65信号通路,促进胰腺癌细胞的增殖并抑制细胞的凋亡,进而影响胰腺癌的发展,以上结果表明,FKN/CX3CR1或许可作为临床上早期胰腺癌治疗的有效靶点。更多还原

【Abstract】 Background:Pancreatic cancer is one of the most lethal gastrointestinal tract malignancies and the fourth most common cause of cancer-related death worldwide.The overall prognosis for patients diagnosed with pancreatic cancer remains dismal,with 5-year survival rates averaging<6%.Cancer cells are difficult to eliminate because of the unlimited growth and proliferation,transformation and metastasis of cancer cells.But till now,the exact mechanism of occurrence and development is still not clear.Objective:The aim of this study was to explore the role of FKN on the proliferation and apoptosis in pancreatic cancer cell;and explore the physiological mechanism of FKN affecting the proliferation and apoptosis in pancreatic cancer cells.So we find FKN/CX3CRlassociated with tumor progression and prognosis,which will provide the effective targeted for cancer therapy.Methods:In this study,we detected the expression levels of FKN and CX3CR1 by immunohistochemistry in fresh pancreatic cancer and paracarcinoma tissues which obtained from 25 patients who were verified by surgery and pathology diagnosis.We chose three pancreatic cancer cell lines Capan-2,MIA PaCa-2,Aspc-1.We detected the expression of FKN and CX3CR1 in pancreatic cancer cell lines.By western blotting,CCK-8,flow cytometry,immunofluorescence and other techniques we tested the effects of FKN plasmid,exogenous FKN and FKN small interfering RNAs on pancreatic cancer cell proliferation,apoptosis and cell cycle.Finally,we studied the physiological mechanism of FKN in the process of proliferation and apoptosis of pancreatic cancer by the inhibitors of PI3K and NF-kappa B/p65.Results:FKN and CX3CR1 have expressed in pancreatic cancer and paracarcinoma tissues,and the expression levels of them were all higher in carcinoma tissues than that in paracarcinoma tissues.In the pancreatic cancer cell Capan-2,MIA PaCa-2 and Aspc-1,FKN and CX3CR1 were also highly expressed by Western Blotting.the results of flow cytometry,immune fluorescent confocal experiment and western blotting showed that FKN can promote pancreatic cancer cell proliferation and inhibit apoptosis of pancreatic cancer cells and promote cell cycle G1 phase to S phase.However,this effect was significantly reduced after the addition of PI3K or NF-kappa B/p65 inhibitors.Conclusion:The study confirmed that FKN combined with its receptor CX3CR1 can promote pancreatic cancer cell proliferation and inhibit apoptosis through the AKT/NF-kappaB/p65 signal pathway,which influenced the development of pancreatic cancer.The above results indicate that FKN/CX3CR1 may be an effective target for early treatment of pancreatic cancer.更多还原