黄芪-丹参药对通过下调miR-466b-5p改善高血压肾损害

目的：探讨黄芪-丹参药对通过调节微小核糖核酸-466b-5P（miR-466b-5p）改善高血压肾损害的机制。方法：基于微RNA（miRNA）测序寻找自发性高血压大鼠与Wistar-京都鼠（WKY）的差异基因,构建腺相关病毒转染小鼠,24只C57BL/6小鼠,随机选取12只尾静脉注射小鼠腺相关病毒miR-466b-5P（rAAV-miR-466b-5P）构建miR-466b-5p过表达组,余12只作为空白对照组注射腺相关病毒-9（AAV-9）空载体,转染6周后再各随机分为2组,每组6只,A组（黄芪-丹参+miR-466b-5P过表达组）和C组（黄芪-丹参+空白对照组）以黄芪配方颗粒：2.036 g/kg+丹参配方颗粒：0.255 g/kg灌胃;B组（miR-466b-5P过表达组）和D组（空白对照组）以相同体积生理盐水灌胃;灌胃28 d后观察各组小鼠尿β2-微球蛋白（β2-MG）、N-乙酰-β-D-葡萄糖苷酶（NAG）、微量白蛋白（mALB）,血清胱抑素C（Cys-C）、血管紧张素Ⅱ（AngⅡ）、C反应蛋白（CRP）水平和肾脏病理,进行实时荧光定量(RT-qPCR)与蛋白质印迹法(Western Blotting)寻找其靶基因。结果：1）基于前期miRNA测序选定了miR-466b-5p;2）与D组比较,B组小鼠尿β2-MG、NAG、mALB、血清Cys-C、AngⅡ、CRP均显著升高（P<0.01）;与B组比较,A组小鼠血清Cys-C、CRP明显升高（P<0.05）,尿β2-MG、NAG、mALB、血清AngⅡ无明显改变（P>0.05）;3）与D组比较,B组小鼠出现明显肾小球硬化、小管纤维化,A组较B组有明显改善;4）与D组比较,B组小鼠透明质酸酶2（HAS2）明显下调,而经黄芪-丹参干预后有所回调。结论：miR-466b-5p的过表达会导致高血压肾损害的发生,黄芪-丹参药对通过靶向HAS2下调miR-466b-5p改善高血压肾损害。     

MiR-206 inhibits Head and neck squamous cell carcinoma cell progression by targeting HDAC6 via PTEN/AKT/mTOR pathway

As a kind of endogenous noncoding small RNA, microRNA (miRNA) plays important roles of regulation to various physiological functions, while its affections on senescence of human Head and neck squamous cell carcinoma (HNSCC) are still unknown. The objective of this study was to investigate the effect of miR-206 in HNSCC tissues, adjacent normal tissues and cell lines, and explore its biological functions in HNSCC.In our study, the level of miR-206 in HNSCC tissues and adjacent normal tissues was detected via real-time qPCR. The effect of miR-206 on cell proliferation was tested by MTT assay, colony formation and cell cycle assays. In order to explore the effect of miR-206 on HNSCC cell migration and invasion, we performed wound healing assays and transwell assays. Luciferase reporter assays were designed to identify the interaction between 3′UTR of HDAC6 and miR-206. The level of signaling pathway-related proteins was determined by western blot. The expression of miR-206 was found to be observably decreased in HNSCC tissues and cell lines through real time-PCR. Restoration of miR-206 weaked cell proliferation, invasion and migration in HNSCC cells and the cell cycle was arrest in S phase. Further explores have shown that miR-206 could inhibit HNSCC cells proliferation by targeting the HDAC6 via PTEN/AKT/mTOR pathway. Taken together, our results demonstrated that miR-206 plays a critical role in HNSCC progression by targeting HDAC6 via PTEN/AKT/mTOR pathway, which might be a potential target for diagnostic and therapeutic in HNSCC.     

Silencing of MicroRNA-21 confers the sensitivity to tamoxifen and fulvestrant by enhancing autophagic cell death through inhibition of the PI3K-AKT-mTOR pathway in breast cancer cells

Tamoxifen (TAM) and fulvestrant (FUL) represent the major adjuvant therapy to estrogen receptor-alpha positive (ER⁺) breast cancer patients. However, endocrine resistance to TAM and FUL is a great impediment for successful treatment. We hypothesized that miR-21 might alter the sensitivity of breast cancer cells to TAM or FUL by regulating cell autophagy. Using the ER⁺ breast cancer cells, we knockdown miR-21.by transfection with miR-21 inhibitor, then the cells were exposed to TAM or FUL and the percentages of apoptosis and autophagy were determined. Knockdown of miR-21 significantly increased the TAM or FUL-induced apoptosis in ER⁺ breast cancer cells. Further, silencing of miR-21 in MCF-7 cells enhanced cell autophagy at both basal and TAM or FUL-induced level. The increase of autophagy in miR-21-knockdown MCF-7 cells was also indicated by increase of beclin-1, LC3-II and increased GFP-LC3 dots. Importantly, knockdown of miR-21 contributed to autophagic cell death, which is responsible for part of TAM induced cell death in miR-21 inhibitor-transfected cells. Further analysis suggested that miR-21 inhibitor enhance autophagic cell death through inhibition of PI3K-AKT-mTOR pathway. MiR-21 coordinated the function of autophagy and apoptosis by targeting Phosphatase and tensin homolog (PTEN) through inhibition of PI3K-AKT-mTOR pathway. In conclusion, silencing of miR-21 increased the sensitivity of ER⁺ breast cancer cells to TAM or FUL by increasing autophagic cell death. Targeting autophagy-related miRNAs is a potential strategy for overcoming endocrine resistance to TAM and FUL.     

Up-regulated microRNA-155 expression is associated with poor prognosis in cervical cancer patients

BackgroundMicroRNAs (miRNAs) play important roles in tumor development and progression. The purposes of the study was to investigate the role of miR-155 in cervical cancer.MethodsQuantitative real-time RT-PCR (qRT-PCR) was performed to examine miR-155 expression in cervical cancer tissues and adjacent non-cancerous tissues. The association with overall survival of patients was analyzed by Kaplan-Meier survival analysis. Small interfering RNA (siRNA) was used to suppress miR-155 expression in cervical cancer cells. In vitro assays were performed to further explore the biological functions of miR-155 in cervical cancer.ResultsWe found that miR-155 expression was markedly up-regulated in cervical cancer tissues and correlated with FIGO stage, lymph nodes metastasis, vascular invasion and HPV. Patients with high miR-155 expression level had poorer overall survival than those with low miR-155 expression. Furthermore, multivariate Cox regression analysis suggested that increased miR-155 was an independent prognostic indicator for cervical cancer (P = 0.007; HR = 2.320; 95%CI: 1.259–4.276). Moreover, knockdown of miR-155 was demonstrated to inhibit cell proliferation, migration, and invasion in vitro.ConclusionOur study presents that miR-155 is a novel molecule involved in cervical cancer progression, which provide a potential prognostic biomarker and therapeutic target.     

慢性淋巴细胞性白血病患者miR-9-3甲基化异常及其意义

目的本研究旨在探讨microRNA-9-3(miR-9-3)在慢性淋巴细胞性白血病骨髓细胞中的甲基化异常及其意义。方法采用甲基化特异性聚合酶链反应(MSP)技术检测8例正常骨髓组织、78例新确诊的慢性淋巴细胞性白血病患者和7种白血病细胞株甲基化水平。结果正常对照组miR-9-3呈未甲基化状态;7种白血病细胞株中有5种呈未甲基化状态(71.4%);78例慢性淋巴细胞性白血病患者中65例呈miR-9-3甲基化,MSP阳性率为83%。5-氮杂-2'-脱氧胞苷(5-Aza Dc)处理白血病细胞株后I83-E95和WAC3CD5+细胞株miR-9-3呈未甲基化状态。结论慢性淋巴细胞性白血病患者存在miR-9-3表达受抑,可能与其基因甲基化异常有关。而miR-9-3甲基化在激活慢性淋巴细胞性白血病患者NF-κB1信号通路的作用值得进一步研究。     

脑膜瘤中miR-200b和ZEB1的表达水平变化及其与预后的关系

目的 探讨脑膜瘤组织中miR-200b、ZEB1表达水平变化与脑膜瘤患者预后的关系。方法 选择2013年5月-2014年12月在本院行神经外科手术治疗的脑膜瘤患者42例,采用qRT-PCR法检测各研究对象脑膜瘤组织与瘤旁组织中miR-200b表达水平,采用免疫组织化学染色法检测ZEB1水平,分析miR-200b、ZEB1表达水平与脑膜瘤临床病理特征的相关性,并对出院后的脑膜瘤患者进行随访,分析miR-200b、ZEB1表达水平变化与脑膜瘤患者预后的关系。结果 脑膜瘤组织miR-200b表达水平显著低于瘤旁组织(P<0.05); 脑膜瘤组织ZEB1阳性表达率高于瘤旁组织(P<0.05); miR-200b、ZEB1表达水平与肿瘤分期有关(P<0.05); miR-200b高表达组的5年无瘤生存率显著高于miR-200b低表达组,ZEB1阳性表达组的5年无瘤生存率显著低于ZEB1阴性表达组(P<0.05); miR-200b低表达、ZEB1阳性表达可能是脑膜瘤患者不良预后的独立危险因素。结论 在脑膜瘤组织中miR-200b低表达、ZEB1阳性表达,且与患者无瘤生存期有关,可作为判断脑膜瘤患者预后的潜在标志物。     

miR-26b-3p靶向调控TRA2B表达抑制神经胶质瘤细胞的增殖和转移

目的探讨miR-26b-3p在神经胶质瘤细胞中的表达,以及其对神经胶质瘤细胞增殖和转移的影响。方法采用荧光实时定量PCR(qRT-PCR)检测神经胶质瘤组织与癌旁组织中miR-26b-3p与TRA2B的表达水平;通过向神经胶质瘤细胞细胞系中转染miR-26b-3p mimic和inhibitor干扰内源miR-26b-3p的表达,同时检测其靶基因TRA2B蛋白的表达变化;采用CCK-8法检测各组神经胶质瘤细胞增殖能力的变化,以及划痕实验对癌细胞迁移能力进行评估。结果神经胶质瘤患者组织与癌旁组织比较,miR-26b-3p的表达水平显著上调,TRA2B的表达水平显著下降,差异有统计学意义(P<0.05);Pearson相关性分析显示,在32例神经胶质瘤患者神经胶质瘤组织中,TRA2B的表达水平与miR-26b-3p表达呈显著负相关(r=-0.510;P<0.05);SHG-44细胞体外转染实验结果发现,降低细胞内源miR-26b-3p的表达时,TRA2B的表达水平显著升高,细胞的增殖活性以及转移能力会被抑制;而上调细胞内源miR-26b-3p的表达时,TRA2B的表达水平显著下降,细胞的增殖活性与转移能力会显著提高,相比于对照组,差异有统计学意义(P<0.05)。结论 miR-26b-3p在神经胶质瘤组织中高表达,其可能靶向调控TRA2B的表达抑制神经胶质瘤细胞的增殖活性与转移能力,在神经胶质瘤的发生发展过程中起着重要的生理功能。     

miR-195 inhibits tumor growth and angiogenesis through modulating IRS1 in breast cancer

Angiogenesis has been found as an attractive target for drug therapy as it is necessary for tumor growth. Accumulating evidences show that microRNAs (miRNAs), which are a group of highly conserved, single-stranded, short non-coding RNAs, play important roles through directly targeting angiogenic factors and protein kinases. The purpose of this study is to investigate the role of miR-195 in breast cancer development and angiogenesis through targeting IRS1. We show that miR-195 is inversely related with Insulin receptor substrate 1 (IRS1) in both breast cancer cells and breast cancer tissues. Induction of miR-195 could suppress IRS1 protein expression through binding to its 3′UTR regions either by transfection with miR-195 oligo or by infection with lentivirus encoding miR-195 gene. Moreover, re-expression of IRS1 reverses miR-195-mediated repression of tumor cell growth and miR-195 inhibits tumor angiogenesis through suppressing IRS1-VEGF axis. These data suggest that miR-195 mimics are potential therapeutic agents for breast cancer diagnose.     

miR-7在肾癌中的表达及临床意义研究

目的检测肾细胞癌组织及相应癌旁正常组织中miR-7的表达,分析18达与临床病理特征之间的关系,为进一步研究miR-7在肾癌的发生发展中作用奠定基础。方法收集48例经手术切除的配对肾癌及癌旁组织标本,提取组织中总RNA,经逆转录获得cDNA,采用实时荧光定量PCR（RT-qPCR）方法检测标本中miR-7表达量,并通过统计学软件分析其表达量与患者临床病理特征之间的相关性。结果与相应癌旁组织相比,肾癌组织中miR-7表达量明显升高,其中34例（70.8%）肾癌标本miR-7表达上调,14例（29.2%）表达下调,差异有统计学意义（P〈0.05）。肾细胞癌中miR-7表达上调与患者年龄、性别、肾癌组织类型、TNM分期、AJCC临床分期无相关性（P〉0.05）。结论 miR-7在肾癌组织中明显高表达,提示其可能与肾癌的发生和发展相关,可能成为肾癌早期诊断、治疗乃至预后判断的新的生物标记物。