MiR-21/RASA1 axis affects malignancy of colon cancer cells via RAS pathways

AIM:To determine how the oncogene mi R-21 regulates the RAS signaling pathways and affects colon cancer cell behaviors.METHODS:RAS p21 GTPase activating protein 1(RASA1) protein expression in six colon cancer cell lines was assessed by Western blot.Colon cancer RKO cells were chosen for transfection because they are KRAS wild type colon cancer cells whose RASA1 expression is significantly decreased.RKO cells were transfected with vectors overexpressing or downregulating either mi R-21 or RASA1.Furthermore,a luciferase reporter assay was used to determine whether RASA1 is a gene target of mi R-21.Then,changes in m RNA and protein levels of RASA1,RASGTP,and other components of the RAS signaling pathways were assessed in transfected RKO cells by real-time quantitative reverse transcription-polymerase chain reaction,Western blot and immunoprecipitation.Finally,cell proliferation,apoptosis,invasion,and tumorformation ability w ere assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide dye assay,flow cytometry,transwell assay,and animal experiment,respectively.RESULTS:RASA1 protein levels were significantly decreased in RKO cells compared with the other 5 colon cancer cell lines,and RASA1 was confirmed as a target gene of mi R-21.Interestingly,RASA1 m RNA and protein levels in pre-mi R-21-LV(up-regulation of mi R-21) cells were lower than those in anti-mi R-21-LV(down-regulation of mi R-21) cells(P < 0.05).In addition,pre-mi R-21-LV or si RASA1(down-regulation of RASA1) cells showed higher cell proliferation,reduced apoptosis,increased expression of RAS-GTP,p-AKT,Raf-1,KRAS,and p-ERK1/2,and higher invasion and tumor formation ability,compared with control,antimi R-21-LV or pc DNA3.1-RASA1(up-regulation of RASA1) cells(P < 0.05).CONCLUSION:RASA1 is a target gene of mi R-21,which promotes malignant behaviors of RKO cells through regulation of RASA1 expression.

Lnc-TCL6是肝硬化患者早期诊断和病变分级的潜在生物学标志物

背景:长链非编码RNAs(lncRNAs)作为生物学标志物已在许多疾病中得到应用。然而,目前并没有发现某个lncRNA差异表达可用于肝硬化临床分期。本研究旨在鉴定出一些lncRNA用作肝硬化早期诊断和分级的非侵入性敏感标志物。方法:在包括305例研究对象(包括健康对照85例,乙肝病毒携带者44例、慢性乙肝患者40例、肝硬化患者136例)的3个队列中评估血液lncRNA表达情况。首先,采用CapitalBiotech芯片筛选出诊断肝硬化的候选lncRNAs。然后,采用定量逆转录PCR法分析这些候选lncRNAs在所有肝硬化及不同Child-Pugh分级者中的表达情况。最后,采用Mann-Whitney检验和ROC分析,对其中最有应用前景的lncRNA的诊断准确性进行验证。结果:筛选出lnc-TCL6作为肝硬化早期诊断(Child-Pugh A)的分子标志物,其与健康对照相鉴别的ROC曲线下面积(AUC)为0.636,与乙肝病毒携带者相鉴别的AUC为0.671,与慢性乙肝患者相鉴别的AUC为0.672。而且,lnc-TCL6还可以鉴别不同Child-Pugh分级的肝硬化患者(AUC:0.711-0.837)。与健康对照、乙肝病毒携带者及慢性乙肝患者相比,Child-Pugh A级肝硬化患者lnc-TCL6表达显著升高,但Child-Pugh C级肝硬化患者lnc-TCL6表达却显著降低。结论:lnc-TCL6是一种可用于肝硬化早期诊断且可能提示疾病进展的新的潜在生物学标志物。