Down-regulation of microRNA-126 and microRNA-133b acts as novel predictor biomarkers in progression and metastasis of non small cell lung cancer

Objective: MiRNAs play crucial roles in progression of cancer. However, the underlying mechanisms of miRNAs in non small cell lung cancer are still poorly understood. The aim of this study was to investigate the expression level of microRNA-126 (miR-126) and microRNA-133b (miR-133b) and also their association with clinicopathological features in patients with non small cell lung cancer (NSCLC). Methods: Total RNA was purified from NSCLC tissues and adjacent non-tumor tissues and then quantitative real-time PCR (qRT-PCR) was used to evaluate the expression rate of microRNAs. Furthermore, the association of miR-126 and miR-133b level with clinicopathological features and prognosis were evaluated. Results: Our findings showed that expression of miR-126 was decreased in NSCLC tissues compared with adjacent non-tumor tissues. On the other hand, a lower expression of miR-133b was seen in NSCLC tissues when compared with adjacent non-tumor tissues. In term of miR-126, our results showed that miR-126 was associated with tumor stage and lymph nodes metastasis (P<0.05). In term of miR-133b, our finding indicated that decreased expression of miR-133b was correlated with advanced tumor stage and lymph nodes metastasis (P<0.05). Kaplan-Meier analysis and log-rank test indicated that patients with low expression of miR-126 and miR-133b had a shorter overall survival (log-rank test; P<0.05). Multivariate Cox proportional hazards model revealed that low expression of miR-126 and miR-133b, advanced tumor stage and lymph nodes metastasis were independent prognostic factors for overall survival of NSCLC patients. Conclusions: These findings suggested that miR-126 and miR-133b might play a key role in the progression and metastasis of NSCLC and would be applied as a novel therapeutic agent.     

Prognostic significance of microRNA-203 in cholangiocarcinoma

MircroRNA functions as a tumor suppressor or a promoter in cholangiocarcinoma (CCA). Researchers have found that miR-203 functioned as tumor suppressor in many types of cancer. However, the role of miR-203 that plays in CCA remains to be clarified. We aimed to detect the expression level and the prognostic significance of miR-203 in CCA tissues. qRT-RCR was performed to examine the miR-203 expression levels in CCA tissue specimens and corresponding normal tissues. Our findings suggest that miR-203 expression was an independent poor prognostic factor for CCA patient overall survival. Therefore, miR-203 may serve as a valuable prognostic marker and promising treatment target for CCA.     

The occurrence of cervical cancer in Uygur women in Xinjiang Uygur Autonomous Region is correlated to microRNA-146a and ethnic factor

Aims: The present study is to investigate the effect of microRNA-146a (miR-146a) and ethnic factor in the occurrence of cervical cancer in Uygur women in Xinjiang Uygur Autonomous Region. Methods: A total of 620 pieces of cervical tissues were obtained between September 2010 and September 2013, including 208 cases of cervicitis, 207 cases of cervical intraepithelial neoplasia, and 205 cases of cervical cancer. The relative expression of miR-146a in tissues was measured using quantitative real-time polymerase chain reaction. Polymerase chain reaction - restriction fragment length polymorphism was used to determine the genotypes of miR-146a (rs2910164). Differences between two groups and multiple groups were compared using t-test and one-factor analysis of variance, respectively. Comparison of genotype compositions and genetic balance examinations were performed using χ² test. Results: Uygur women had earlier age of marriage, more times of pregnancy, and more childbirths than Han women. The miR-146a (rs2910164) genotype composition was significantly different between Uygur and Han, with the ratio of GG genotype in Uygur being higher than that in Han. Logistic regression analysis showed that miR-146a (rs2910164) genotypes were significantly correlated to ethnic factor and tumor sizes. The expression of miR-146a was elevated in cervical intraepithelial neoplasia and cervical cancer, especially for Uygur women, with the GG genotype being the most highly expressed. Conclusions: The miR-146a (rs2910164) polymorphism is significantly correlated to ethnic factor and tumor diameters. miR-146a has differential expression in cervical tissues. Allele G of miR-146a (rs2910164) is related to the high expression of miR-146a, and the progression of cervical cancer.     

Involvement of epigenetics and microRNA-29b in the urethane induced inception and establishment of mouse lung tumors

Epigenetic changes are correlated with tumor development showing aberrations in DNA methylation and histone modifications. To find the early changes, we evaluated the epigenetic events from early to late stage of the urethane induced lung tumor development in mouse model and tried to correlate the molecular events with the progression of tumor. We addressed the hypothesis by examining the tumor development, status of DNMTs, HDACs and MBDs, DNA methylation and expression of microRNA-29b during 1 to 36 weeks after urethane exposure that included the period before and after the tumor appearance. Tumors did not appear after 1 or 4 weeks but well defined tumors appeared after 12 weeks and larger tumors appeared at 36 weeks which was prevented by IP6. DNMT1, DNMT3a and DNMT3b were upregulated after urethane exposure at the time of no tumor till the tumor developed and showed its upregulated functional activity. DNMTs are shown to be the targets of microRNA-29b and we showed that microRNA-29b was downregulated in the line of DNMT upregulation. HDAC, the histone modifier, also showed progressive upregulation. Periodic increase in methyl binding proteins, MBD2, supported the expression of gene silencing pathways in terms of the downregulation of tumor suppressor genes, p16 and MLH1. All these molecular alterations were protected in the presence of IP6. Our results showed that the key steps of epigenetics, DNMTs, mir29b, and HDAC1, are altered both before and after the development of tumors.     

miR-203靶向Survivin抑制卵巢癌细胞增殖、迁移与侵袭

目的 探讨miR-203靶向抑制Survivin对卵巢癌SKOV3及OVCAR3细胞增殖、迁移及侵袭的影响。 方法 构建过表达miR-203、Survivin及空白对照组的慢病毒载体,转染卵巢癌SKOV3和OVCAR3细胞,嘌呤霉素筛选后构建空白对照组、过表达miR-203组、过表达Survivin组及过表达miR-203联合Survivin组卵巢癌细胞系。Western blotting方法测定各组卵巢癌细胞中Survivin及上皮-间质转化(EMT)相关蛋白的表达;MTT和平板克隆实验检测卵巢癌细胞增殖能力的改变;Transwell实验检测对细胞迁移和侵袭能力的影响。 结果 (1) 过表达miR-203抑制Survivin的表达及EMT(P<0.05);(2) MTT、平板克隆实验及transwell实验显示,与空白对照组相比,过表达miR-203组能够抑制卵巢癌细胞的增殖、迁移和侵袭(P<0.05);而过表达Survivin逆转了miR-203对卵巢癌的抑制作用(P<0.05)。 结论 miR-203通过靶向Survivin抑制EMT从而抑制卵巢癌的增殖、迁移及侵袭,miR-203/Survivin/EMT轴有望成为卵巢癌治疗的新靶点。     

miR-200c-3p靶向CCNE2抑制肾母细胞瘤细胞增殖

目的 预测和验证miR-200c-3p的靶基因并探讨其对肾母细胞瘤增殖的抑制作用。方法 应用生物信息学软件对miR-200c-3p在肾母细胞瘤中的靶基因进行预测,建立SK-NEP-1及G401的miR-200c-3p过表达及抑制表达的稳定细胞系。实验设未转染组（Blank）、模拟物阴性对照组（mimic NC）、miR-200c-3p模拟物组（miR-200c-3p mimic）、抑制剂阴性对照组（inhibitor NC）及miR-200c-3p抑制剂组（miR-200c-3p inhibitor）。采用 RT-PCR及Western blot方法检测CCNE2在SK-NEP-1及G401不同组中的表达水平,荧光素酶报告基因检测miR-200c-3p 和CCNE2的靶向关系,细胞计数盒（CCK-8）和软琼脂克隆形成实验检测miR-200c-3p对SK-NEP-1及G401细胞增殖的抑制作用。结果 生物信息学方法预测CCNE2为miR-200c-3p的靶基因之一。RT-PCR实验结果示肾母细胞瘤细胞中miR-200c-3p模拟物组的CCNE2的表达水平低于模拟物阴性对照组（P<0.05）;荧光素酶报告基因检测证实CCNE2是miR-200c-3p的靶基因（P<0.01）。Western blot示在肾母细胞瘤细胞中miR-200c-3p模拟物组的CCNE2蛋白的表达水平低于模拟物阴性对照组（P<0.05）;CCK-8和软琼脂克隆形成实验证实miR-200c-3p对肾母细胞瘤细胞的增殖能力有明显抑制作用（P<0.01）;而miR-200c-3p抑制剂组结果相反。结论 miR-200c-3p通过靶基因 CCNE2抑制肾母细胞瘤细胞的增殖。     

miR-126与心血管疾病关系的研究进展

微小核糖核酸(microRNAs,miRNA)是一类内源性高度保守的非编码小分子RNA,其可通过降解信使RNA并抑制其翻译在转录水平调控靶基因的表达,参与细胞生长、发育、分化和增殖等多个生命过程。MicroRNAs-126(miR-126)是一种内皮特异性miRNA,与高血压、动脉粥样硬化、冠心病、心力衰竭等心血管疾病密切相关。近期研究表明,miR-126能显著影响心血管疾病的发生、发展和疾病的预后。本文就miR-126与心血管疾病的关系进行综述。     

miRNA-205在食管鳞状细胞癌患者组织中的表达及临床意义

目的 探讨miRNA-205在食管鳞状细胞癌(esophageal squamous cell carcinoma,ESCC)患者组织中的表达情况.方法 收集56例经病理确诊的ESCC患者的手术切除组织(癌组织及其癌旁组织),采用实时荧光定量聚合酶链式反应(real-time quantitative polymerase chain reaction,qRT-PCR)检测miRNA-205的表达,并分析miRNA-205在不同临床病理资料之间的差异.结果 ESCC患者组织中miRNA-205相对表达量为1.475(0.444～5.478),显著高于癌旁组织(Z=-3.067,P=0.002);miRNA-205在不同TNM分期ESCC患者组织中存在差异,TNM分期越高,miRNA-205表达量越高,差异具有统计学意义(F=36.335,P＜0.001);miRNA-205在淋巴结转移阴性/阳性组织中存在差异,淋巴结转移阳性组miRNA-205表达量显著高于淋巴结转移阴性组(Z=-3.353,P=0.001).结论 miRNA-205在ESCC患者癌组织中表达升高,并可能参与ESCC的发生与发展.     

基于荧光共振能量转移技术的STAT3二聚化抑制剂筛选模型的建立

目的%20构建STAT3二聚化抑制剂筛选模型,为STAT3抑制剂筛选提供实验方法。方法%20分别构建pECFP-N1-STAT3和pEYFP-N1-STAT3的荧光报告载体,利用脂质体转染技术将二者共转入HEK-293T细胞,利用ECFP和EYFP两种荧光蛋白之间能量共振转移,检测磷酸化STAT3分子的二聚化水平,并检测Stattic对二聚化的影响。结果%20成功构建了pECFP-N1-STAT3和pEYFP-N1-STAT3的荧光报告载体。将两种荧光报告载体共转染HEK-293T细胞后,Western%20Blot%20检测结果显示STAT3以及p-STAT3的表达水平明显增加。以458nm波长激发ECFP,其发射波长可激发EYFP。加入STAT3二聚化抑制剂Stattic后,荧光强度降低,且呈现一定的剂量依赖性。结论%20基于荧光共振能量转移技术的STAT3二聚化抑制剂筛选模型构建成功。