【作者】 周颖；

【导师】 李晓波；

【作者基本信息】 上海交通大学 ， 内科学（消化系病）， 2017， 硕士

【摘要】 目的:研究LRG1在结直肠癌患者血浆和组织中的表达情况以及LRG1对结直肠癌细胞增殖、细胞周期和细胞凋亡的影响,探讨LRG1与结直肠癌发生发展的关系。方法:利用ELISA实验检测结直肠癌患者血浆LRG1分泌情况,并用Real-time PCR和Western blot检测60对新鲜结直肠癌和配对正常组织标本中LRG1的m RNA和蛋白表达水平。对结肠癌细胞SW480及HCT116行si LRG1、si RUNX1、si NC转染处理或用人重组LRG1蛋白刺激,采用CCK-8、细胞周期及细胞凋亡实验评估结肠癌细胞的增殖和凋亡情况。通过Real-time PCR和Western blot检测细胞周期关键基因cyclin D1,cyclin B和cyclin E以及细胞凋亡相关基因Bax、Bcl-2和cleaved caspase-3的表达水平。结果:结直肠癌患者血浆LRG1分泌水平升高,且肿瘤切除术后患者血浆LRG1分泌水平显著下降(P<0.001)。LRG1在结直肠癌组织中的m RNA和蛋白表达水平均显著高于配对正常组织(P<0.001)。沉默LRG1后显著抑制SW480和HCT116细胞的增殖能力,诱导G0/G1期捕获,并促进细胞凋亡。另外,下调LRG1表达后SW480及HCT116细胞中细胞周期蛋白cyclin D1,cyclin B,cyclin E和抑凋亡基因Bcl-2的表达均显著下调,促凋亡基因Bax及cleaved caspase-3的表达显著上调。RUNX1的表达随人重组LRG1刺激浓度的增加而不断升高,沉默RUNX1能逆转LRG1对结肠癌细胞的促增殖和凋亡抑制作用。结论:LRG1在结直肠癌患者血浆及组织中均表达上调,并通过调控RUNX1促进结直肠癌细胞增殖,抑制其凋亡,提示LRG1可能成为潜在的结直肠癌早期诊断和术后随访标记物以及结肠癌治疗的靶向分子。更多还原

【Abstract】 Aims: To investigate the plasma and tissue level of LRG1 in colorectal cancer(CRC)patients,and to detect the influence of LRG1 on the proliferation and apoptosis of colorectal cancer cells,therefore to explore the relationship between LRG1 and the tumorigenesis of CRCMethods: Enzyme-linked immunosorbent assay was used to measure the plasma level of LRG1 in the CRC patients.Real-time PCR and western blot were performed to evaluate LRG1 m RNA and protein expression in 60 cases of fresh CRC and matched normal tissues.SW480 and HCT116 cells were treated with si LRG1,si RUN1,si NC,or human recombinant LRG1.CCK-8 assays,cell cycle assays and apoptosis assays were performed to evaluate the proliferation and apoptosis of CRC cells.Cell cycle factors cyclin D1,cyclin B,cyclin E as well as apoptotic key genes Bcl-2,Bax and cleaved caspase-3 were detected by Real-time PCR and western blot.Results: The plasma level of LRG1 was significantly increased in CRC patients,while it was remarkably decreased in patients with resected colorectal cancers.In addition,both m RNA and protein levels of LRG1 were remarkable overexpressed in CRC tissues than normal tissues(P<0.001).The knockdown of LRG1 significantly inhibited cell proliferation,induced cell cycle arrest at the G0/G1 phase,and promoted apoptosis in SW480 and HCT116 cells in vitro.In addition,LRG1 silencing led to the downregulation of the levels of cyclin D1,B,and E and anti-apoptotic Bcl-2 while it up-regulated the expression of pro-apoptotic Bax and cleaved caspase-3.Furthermore,RUNX1 could be induced by LRG1 in a concentration-dependent manner,while the knockdown of RUNX1 blocked the promotion of the proliferation and inhibition of apoptosis induced by LRG1.Conclusions: The plasma and tissue level of LRG1 was significantly increased in CRC patients,and LRG1 plays a crucial role in the proliferation and apoptosis of CRC by regulating RUNX1 expression.Thus,LRG1 may be a potential detection biomarker as well as a marker for monitoring recurrence and therapeutic target for CRC.更多还原