FERM和merlin蛋白对不同分化程度胃癌细胞生物学功能的调节

目的研究FERM和merlin蛋白对不同分化程度胃癌细胞生物学功能的调节作用。方法利用Lipofectamine 2000将重组质粒FERM、merlin-1和-2,以及空白对照质粒分别转染到不同分化程度胃癌细胞BGC-823、MGC-803、SGC-7901和正常胃粘膜细胞GES-1中,培养36 h后用MTT检测法检测细胞增殖,用transwell方法检测细胞黏附侵移能力。结果与对照组相比,转染FERM、merlin-2蛋白到不同分化程度胃癌细胞(BGC-823、MGC-803、SGC-7901)、正常胃粘膜细胞(GES-1)后,发现对细胞增殖、细胞黏附侵移能力没有影响(P0.05);而转染merlin-1蛋白后,无论对不同分化程度胃癌细胞、还是正常胃粘膜细胞的增殖、细胞黏附能力均有显著抑制作用(P0.05)。结论 merlin-1对胃癌细胞和正常胃粘膜细胞增殖、黏附侵移能力有抑制作用。     

lncRNA HOTAIR在胃癌中的表达及对胃癌细胞增殖及自噬的影响

目的研究lncRNA HOTAIR在胃癌中的表达及其对胃癌细胞增殖和自噬的影响,探讨lncRNA HOTAIR调控胃癌细胞自噬的机制。方法收集2015年1月—2017年12月于中国医科大学附属肿瘤医院确诊为胃癌的60例癌组织及其配对癌旁组织;实时荧光定量PCR (RT-PCR)检测HOTAIR表达水平,CCK-8实验检测siHOTAIR及si-ATG3对细胞增殖影响,蛋白免疫印迹实验检测HOTAIR对细胞自噬水平及ATG3表达水平影响,免疫荧光实验检测HOTAIR对胃癌细胞自噬的影响。结果 HOTAIR在胃癌组织及癌细胞株MGC-803、BGC-823、SGC-7901中表达水平显著高于癌旁正常组织及人胃黏膜细胞株GES-1。通过转染si-HOTAIR,构建胃癌HOTAIR干扰细胞株;与空载对照组相比,HOTAIR干扰组胃癌细胞增殖水平降低,胃癌细胞自噬标志蛋白LC3表达水平降低,LC3-Ⅱ/Ⅰ比例降低,P62表达增高,自噬相关蛋白ATG3表达亦降低。结论胃癌组织中HOTAIR表达上调,抑制HOTAIR促进了胃癌细胞增殖能力;且HOTAIR能够通过ATG3促进自噬水平。     

沉默TROP2基因抑制胃癌BGC-823细胞体外迁移侵袭能力

目的探讨TROP2基因在胃癌细胞迁移侵袭过程中的作用。方法用基因克隆技术构建针对TROP2的小干扰RNA(siRNA),瞬时转染胃癌BGC-823细胞系,荧光实时定量PCR和Western blot法检测细胞TROP2mRNA和蛋白表达水平,MTT法测定细胞增殖能力,Transwell小室实验观察细胞迁移侵袭能力。结果酶切鉴定和DNA测序分析显示,TROP2靶向RNA干扰重组质粒构建成功。筛选得到最佳干扰效果质粒,该质粒转染细胞后,细胞增殖和迁移侵袭能力显著下降(P<0.05)。结论干扰TROP2基因具有抑制胃癌BGC-823细胞迁移侵袭的作用,TROP2基因可能成为癌基因靶向治疗的分子靶点。     

姜黄素调控Wnt信号通路抑制胃癌细胞上皮间质转化的研究

目的探讨姜黄素对人胃癌细胞株BGC-823上皮间质转化(EMT)的影响机制。方法将细胞分为姜黄素干预组、TGF-β组、BGC-823细胞对照组、GES-1细胞对照组。采用姜黄素(10μmol/L)干预人胃癌细胞株BGC-823,通过MTT试验检测胃癌细胞增殖能力;通过Transwell小室试验检测胃癌细胞侵袭功能改变;通过Western blot检测各组细胞E-cadherin、Vimentin蛋白及Wnt信号通路相关蛋白表达。结果 TGF-β诱导能够促进人胃癌细胞株BGC-823增殖(P 0.05),胃癌细胞表现出更高的侵袭能力(P 0.01),而姜黄素干预后胃癌细胞的增殖及侵袭能力均明显受抑制。Western blot检测表明,TGF-β诱导干预后,BGC-823细胞E-cadherin、Vimentin蛋白水平升高,与此同时GSK3β、β-catenin及c-Myc的表达明显上调(P 0.01);姜黄素干预能有效抑制胃癌细胞EMT作用,Wnt信号通路蛋白水平显著下调(P 0.05)。结论姜黄素可以通过调控Wnt/β-catenin信号通路抑制胃癌细胞EMT作用,从而抑制肿瘤细胞的侵袭、转移,实现抗肿瘤作用。     

胃癌中REIC蛋白分泌水平及其增殖抑制作用研究

目的:研究胃癌患者血清和胃癌细胞培养液上清中分泌型REIC含量及重组REIC对胃癌细胞增殖的影响.方法:ELISA检测血清和细胞上清中的REIC表达;WST-8法检测胃癌细胞的增殖.结果:REIC在胃癌患者中的分泌水平比正常健康人明显降低(P<0.05),并且与肿块大小呈负相关(P<0.05);胃癌细胞系培养液上清中的REIC表达低于正常的胃勃膜细胞(P<0.05);胃癌AGS细胞系转染REIC后,细胞培养液上清中的REIC浓度升高(P<0.05);加人外源性融合的REIC可以抑制胃癌细胞AGS和BGC-823的增殖(P<0.05).结论:分泌型的REIC在胃癌细胞中表达降低,重组REIC蛋白可抑制胃癌细胞的增殖,REIC蛋白可以作为胃癌早诊、生物学和基因治疗的靶点.     

HOXB7基因沉默对人胃癌细胞活力、迁移和侵袭的影响

目的:探讨靶向沉默HOXB7基因对人胃癌SGC-7901细胞活力、迁移和侵袭作用的影响及潜在机制。方法:设计靶向HOXB7的siRNA,然后瞬时转染胃癌SGC-7901细胞,实时荧光定量PCR和Western blot法检测siRNA靶向沉默的效果。MTT法检测细胞的活力,Western blot法检测细胞周期相关蛋白CDK4和cyclin D1蛋白的表达水平,Transwell小室侵袭实验检测SGC-7901细胞的侵袭能力,实时荧光定量PCR和Western blot法检测胃癌SGC-7901细胞PTEN和VEGF的表达水平以及p-AKT的蛋白水平。结果:胃癌组织中HOXB7基因在mRNA和蛋白水平的表达均显著高于对照组织。siRNA可有效靶向沉默SGC-7901细胞中HOXB7的表达,且沉默HOXB7表达后SGC-7901细胞活力明显下降,同时细胞周期相关蛋白CDK4和cyclin D1的表达亦下调。靶向沉默HOXB7可明显抑制SGC-7901细胞中AKT的磷酸化水平,抑制胃癌细胞的侵袭转移,同时下调VEGF的表达水平,抑制胃癌组织中的肿瘤血管生成。结论:HOXB7作为一个癌基因,可上调细胞周期相关蛋白CDK4、cyclin D1和侵袭相关分子VEGF的表达,上调AKT的磷酸化水平,进而抑制PTEN的功能,促进胃癌细胞SGC-7901的生长、迁移和侵袭能力。     

EZH2在胃癌组织中的表达及对胃癌细胞增殖的影响

目的探讨EZH2表达与胃癌临床病理特征、预后的关系,及其对胃癌细胞增殖的影响。方法采用免疫组化SP法检测EZH2在65例胃癌组织及20例正常胃黏膜组织中的表达,分析其表达与胃癌临床病理特征的相关性,应用Kaplan-Meier法及Log-rank检验分析EZH2表达与胃癌患者生存率的关系。采用Western blot法检测胃癌细胞株SGC7901、BGC823中EZH2的表达。EZH2 Sh RNA转染胃癌细胞株后,采用MTT法检测细胞的增殖率。结果胃癌组织中EZH2蛋白阳性率为63.07%,正常胃黏膜组织中EZH2蛋白阳性率为10%;胃癌组织中EZH2蛋白表达明显高于正常胃黏膜组织(P0.000 1)。胃癌组织中EZH2蛋白表达与肿瘤直径(P0.000 1)、肿瘤浸润深度(P0.000 1)、淋巴结转移(P=0.001)及分期相关(P=0.001)。EZH2阳性者5年总生存率明显低于阴性者(P=0.001)。胃癌细胞株SGC7901和BGC823中EZH2蛋白高表达,Sh RNA介导的EZH2表达沉默可明显抑制胃癌细胞的增殖。结论 EZH2高表达是胃癌患者预后不良的重要因子,EZH2可明显促进胃癌细胞的增殖。     

抑瘤素M对人胃癌细胞迁移及COX-2表达的影响

研究抑瘤素M(oncostatin M,OSM)对人胃癌细胞株MKN-28体外侵袭能力和环氧合酶-2(COX-2)表达的影响,探讨其在胃癌浸润转移中的作用及可能机制。运用Transwell小室法检测OSM对胃癌细胞迁移的影响;将靶向COX-2基因的小分子干扰RNA(siRNA)瞬时转染胃癌细胞,采用蛋白印迹法(western blot)和MTT法分析转染后胃癌细胞COX-2蛋白表达情况和细胞生长情况;荧光定量聚合酶链反应(RT-PCR)和western blot分别检测25 ng/ml的OSM对胃癌细胞COX-2 mRNA和蛋白表达的影响。研究发现,25 ng/ml OSM能促进胃癌细胞的迁移(P0.05)。用针对COX-2的siRNA转染胃癌细胞后,COX-2蛋白表达降低,细胞生长变慢。经25 ng/ml OSM作用的胃癌细胞COX-2 mRNA及蛋白的表达升高(P0.05)。结果提示,一定剂量的OSM可能通过诱导COX-2的表达,促进胃癌细胞侵袭和转移。     

小分子化合物E-039抑制胃癌细胞迁移侵袭及其分子机制

目的研究小分子化合物E-039对人胃癌细胞增殖、迁移侵袭的影响及其分子机制。方法用MTT检测不同浓度小分子化合物E-039对胃癌细胞BGC823和MKN-45增殖能力的影响。分别用0、20、40、60μmol/L浓度的E-039处理BGC823和MKN-45两种胃癌细胞,应用Transwell小室检测其对上述细胞迁移侵袭能力的抑制作用,Western Blot检测化合物对金属基质蛋白酶2(MMP2)表达及丝切蛋白(Cofilin)磷酸化水平的影响。结果化合物E-039对于BGC823和MKN-45两种胃癌细胞的增殖能力无明显影响,却能够以剂量依赖的方式抑制BGC823及MKN-45细胞的迁移和侵袭。Western Blot结果显示E-039下调胃癌细胞BGC823中MMP2的表达及Cofilin的磷酸化水平。结论化合物E-039能够通过下调胃癌细胞中MMP2的表达及Cofilin的磷酸化水平,抑制胃癌细胞BGC823、MKN-45的迁移和侵袭。     

lncRNA XIST通过靶向miR-3666调控胃癌细胞增殖、侵袭转移机制北大核心CSCD

目的:探讨lncRNA XIST对胃癌细胞增殖、侵袭转移的影响及分子机制。方法:收集武汉市第四医院30例胃癌患者癌组织及癌旁组织,培养正常胃黏膜上皮细胞MRG-1、胃癌细胞SGC-7901和AGS,采用实时荧光定量PCR(RT-qPCR)检测lncRNA XIST和miR-3666表达水平;通过敲减胃腺癌细胞AGS中lncRNA XIST表达以及下调miR-3666表达水平,分析lncRNA XIST及miR-3666对胃癌细胞的调控作用,将细胞AGS分为si-NC组、si-XIST组、miR-NC组、miR-3666组、si-XIST+antimiR-NC组、si-XIST+anti-miR-3666组;MTT检测细胞增殖情况;Transwell检测细胞迁移和侵袭;流式细胞术检测细胞凋亡;双荧光素酶报告实验检测lncRNA XIST和miR-3666的靶向关系。结果:与癌旁组织和正常胃黏膜上皮细胞MRG-1比较,胃癌组织和AGS、SGC-7901细胞中XIST表达水平升高,miR-3666表达水平降低(P<0.05)。敲除lncRNA XIST或过表达miR-3666,AGS细胞增殖能力减弱,迁移和侵袭能力降低,细胞凋亡率升高(P<0.05)。lncRNA XIST靶向调控miR-3666,抑制miR-3666逆转了XIST基因敲除对胃癌AGS细胞增殖、运动性及凋亡的影响。结论:敲除lncRNA XIST可通过上调miR-3666抑制胃癌细胞增殖、迁移和侵袭,并促进细胞凋亡。     

MicroRNA与胃癌的关系

微小RNA(microRNA)是一类内源性、长度约为16～29 nt非编码小RNA,在细胞发育、增殖、分化、凋亡等方面都起了重要作用.通过与靶mRNA完全或不完全互补配对,引起mRNA的降解或翻译抑制,从而对基因转录后水平进行调控.microRNA既可作为促癌基因参与恶性肿瘤的发生和发育过程,又可作为抑瘤基因控制恶性肿...     

信号通路对胃癌干细胞及胃癌发生的影响

背景：Hedgehog与Wnt/β-catenin信号通路对胃癌干细胞的影响及在胃癌发生发展中的作用机制少见报道。 目的：探讨Hedgehog与Wnt/β-catenin信号通路在胃癌发生发展中的作用机制。 方法：采用肿瘤球悬浮分选法从胃癌组织标本中分选胃癌干细胞。采用免疫组化SP法检测Hedgehog 及Wnt/β-catenin信号通路主要分子SHH、GLI1、Wnt2 及β-catenin在胃癌干细胞中的表达。Spearman相关分析各细胞因子间的相关性分析。 结果与结论：SHH、GLI1、Wnt2及β-catenin 在胃癌干细胞中的阳性表达率分别为74.7%,78.3%,85.5%和83.3%,均显著高于癌旁组织的阳性表达率(P < 0.05)。各细胞因子间在胃癌干细胞中的表达均呈正相关(P < 0.05)。说明在胃癌干细胞中Hedgehog及Wnt/β-catenin信号通路均被激活,二者互相作用可能参与了胃癌的发生发展,为胃癌的干细胞治疗提供了新的研究方向。     

胃癌干细胞CSC-G在胃癌侵袭和转移中的作用

BACKGROUND: Studies have shown that cancer stem cells play an important role in tumor invasion and metastasis, but studies on the role of gastric cancer stem cells in invasion and metastasis of gastric cancer cells were rarely reported. OBJECTIVE: To study the effect of gastric cancer stem cells CSC-G on invasion and metastasis of gastric cancer cells. METHODS: Gastric cancer stem cells CSC-G and gastric cancer cells SGC7901 were cultured in vitro for 10 days followed by spherical colony formation assay, western blot assay for detecting OCT4, SOX2, E-cadherin and CD44 protein expression levels in gastric cancer stem cells CSC-G and gastric cancer cells SGC7901, and Transwell assay for detecting the invasion and migration of gastric cancer stem cells and gastric cancer cells SGC7901. RESULTS AND CONCLUSION: Gastric cancer cells SGC7901 in RPMI1640 medium presented with adherent growth and were quadrilateral or polygonal after passage; gastric cancer stem cells CSC-G in serum-free medium presented with suspension growth, and adherent gastric cancer stem cells were spindle-shaped or round. Compared with gastric cancer cells SGC7901, the protein expressions of OCT4, SOX2 and CD44, the number of cancer cell spheres and the number of trans-membrane cells were significantly increased in the gastric cancer stem cells CSC-G (P < 0.05), and the expression of E-cadherin protein was significantly decreased (P < 0.05). These findings indicate that the gastric cancer stem cells CSC-G can be successfully cultured in vitro, and have enhanced invasion and migration compared with the gastric cancer cells SGC7901, which play an important role in gastric cancer invasion and metastasis.      

Advanced gastric cancer and prognosis

Summary The morphological features of 158 gastric carcinomas were analyzed in an attempt to identify patterns best correlated with prognosis. To this end, the depth of infiltration, vascular invasion, intra- and perineoplastic lymphocytic infiltrate, lymph node metastases and number of metastatic lymph nodes were evaluated according to the several classifications for advanced gastric cancer. A good correlation between prognosis and histological features of malignancy were observed, as well as different five-year survival rates for Mulligan, Lauren and Ming histotypes However, when the influence of each single morphological criteria of malignancy was examined, these differences disappeared for Mulligan and Lauren histotypes. On the other hand, the better prognosis for Ming expanding type carcinomas appeared unrelated to any individual feature of malignancy.     

胃癌干细胞在胃癌侵袭、转移中及血管形成中的作用和机制

BACKGROUND: Although tumor stem cells and their differentiated vascular cells, which are not sensitive to chemotherapy and molecular targeted therapy, reduce overall blood supply of the tumor, these cells facilitate the invasion and metastasis of the tumor, eventually resulting in treatment failure.     

Features of gastritis predisposing to gastric adenoma and early gastric cancer

BACKGROUND/AIMS: Helicobacter pylori gastritis is a risk factor for the development of gastric cancer. The results of several studies indicate that gastric adenomas, which are considered premalignant lesions, may also be associated with H pylori gastritis. However, it is not clear whether there are different patterns of gastritis in these patients compared with patients with gastric cancer or patients with H pylori gastritis alone. Therefore, this study was designed to investigate the patterns of gastritis in these three groups of patients. METHODS: The histological features of gastric mucosa at a distance from the tumour were analysed prospectively in 118 patients with gastric adenoma (mean age, 71.8; female to male ratio, 6 : 4). In addition, for every patient with H pylori associated gastric adenoma an age and sex matched control patient with either H pylori associated early gastric cancer of the intestinal type or H pylori gastritis only was investigated. RESULTS: Only 60 patients (50.9%) with gastric adenoma were infected with H pylori. In the remaining patients, complete atrophic gastritis predominated. In those patients with adenoma and H pylori infection, the gastritis was similar to that seen in patients with early gastric cancer (median score, 2 for activity and degree of gastritis in the antrum and corpus); intestinal metaplasia was common to both groups. These two groups differed significantly from patients with H pylori gastritis only (median grade and activity of gastritis, 1 in antrum and corpus), in whom intestinal metaplasia was rare. CONCLUSIONS: It appears that gastric adenomas and gastric intestinal cancer arise by analogous mechanisms. However, owing to severe atrophic gastritis and a lower incidence of H pylori, adenomas do not appear to be definite precursor lesions for gastric cancer.