冠心病患者白细胞miR-223-3p水平与氯吡格雷治疗后的血小板反应无关

目的：探讨冠心病患者白细胞miR-223-3p水平与氯吡格雷治疗后血小板反应之间的潜在相关性。方法：收 集188名择期经皮冠状动脉介入治疗(percutaneous coronary intervention,PCI)术后接受双重抗血小板治疗的门诊患者的 一般资料和血标本,检测二磷酸腺苷(adenosine diphosphate,ADP)诱导的全血血小板聚集率。选取血小板反应有显著 差异的患者(超反应组和无反应组),检测其白细胞miR-223-3p水平。结果：除ADP诱导的全血血小板聚集率外,两组 患者的一般资料和白细胞miR-223-3p水平差异无统计学意义(P>0.05)。结论：冠心病择期PCI术后门诊患者外周血白细 胞miR-223-3p水平与氯吡格雷治疗后的血小板反应无关。     

7- 二氟亚甲基-5, 4'- 二甲氧基金雀异黄素对大鼠压力性尿失禁模型的疗效及其机制

目的：探讨7-二氟亚甲基-5, 4'-二甲氧基金雀异黄素(7-difluoromethy-5, 4'-dimethoxygenistein,DFMG)对SD大 鼠压力性尿失禁(stress urinary incontinence,SUI)模型的疗效及其机制。方法：采用模拟妊娠、难产产伤及卵巢去势 建立SD大鼠SUI模型,分为正常对照组、SUI组、DFMG(10,20 mg/kg)组,模型鼠行DFMG隔日灌胃治疗。采用膀胱 最大容积(maximal bladder capacity,MBC)、漏尿点压力(leak point pressure,LPP)、腹部漏尿点压力(abdominal leak point pressure,ALPP)以及HE染色和Masson染色检测建模效果;采用RT-PCR检测尿道括约肌细胞(urethral sphincter muscles cells,USMCs)miR-26b及其下游靶基因磷酸酶和肌腱同源染色体(phosphatase and tensin homolog deleted on chromosome 10,PENT)mRNA表达;用Western印迹检测USMCs细胞PENT,磷脂酰肌醇3激酶(phosphatidylinositol 3-kinase,PI3K), 蛋白激酶B(protein kinase B,AKT),B细胞淋巴瘤/白血病-2(B-cell lymphoma 2,Bcl-2),Bcl-2相关X蛋白(Bcl-2 associated X protein,Bax),细胞色素C(cytochrome C,Cyt-C)和含半胱氨酸的天冬氨酸蛋白水解酶(cysteinyl aspartate specific proteinase,caspase-9)的蛋白表达;采用流式细胞仪(flow cytometry,FCM)检测USMCs细胞凋亡率,采用噻唑蓝[3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide,MTT]检测尿USMCs细胞增殖率。结果：成功建立SD大鼠 SUI模型。HE染色和Masson染色显示：与SUI组比较,DFMG组尿道括约肌病理症状显著改善,MBC,LPP和ALPP 均有显著提高(均P<0.05)。RT-PCR结果显示：与SUI组比较,DFMG组USMCs细胞miR-26b mRNA表达升高(P<0.05), PENT mRNA表达下调(P<0.05)。Western印迹显示：与SUI组比较,DFMG组PENT,Bax,Cyt-C和caspase-3蛋白均下调 (均P<0.05);而PI3K,AKT和Bcl-2蛋白表达均上调(均P<0.05);并伴随USMCs细胞凋亡率降低(P<0.05),增殖率增高 (P<0.05)。结论：DFMG可以显著改善SUI模型鼠尿动力学症状,其机制可能与上调miR-26b表达、调控PI3/AKT-Bcl-2/ Bax信号通路而抑制细胞凋亡有关。     

Expression and clinicopathological significance of miR-146a in hepatocellular carcinoma tissues

Background. Aberrant expression of microRNA-146a (miR-146a) has been found in several classes of cancers. However, its expression and clinicopathological contribution in hepatocellular carcinoma (HCC) has not been fully elucidated.

Objective. To explore the clinicopathological significance of the miR-146a level in HCC formalin-fixed paraffin-embedded (FFPE) tissue.

Methods. Eighty-five HCC samples and their para-cancerous normal liver tissues were collected. Total mRNA including miRNA was extracted, and miR-146a expression was determined using real-time RT-PCR. Furthermore, the correlation between the miR-146a expression and clinicopathological parameters was investigated.

Results. MicroRNA-146a expression in HCC tissues was lower compared with that in adjacent non-cancerous hepatic tissues. MicroRNA-146a expression was also related to clinical TNM stage, metastasis, portal vein tumor embolus, and number of tumor nodes.

Conclusions. Down-regulation of miR-146a is related to HCC carcinogenesis and deterioration of HCC. MicroRNA-146a may act as a suppressor miRNA of HCC, and it is therefore a potential prognostic biomarker for HCC patients.     

miRNA-145通过调控IRS-1影响葡萄膜黑色素瘤细胞增殖

背景：微小RNA（miRNA）在肿瘤发生过程中可以作为癌基因或者抑癌基因发挥作用。本文中我们研究了miR-145在葡萄膜黑色素瘤发生过程中所起的作用。 方法：采用安捷伦miRNA芯片在葡萄膜黑色素瘤中检测miRNA的表达谱,并用实时定量聚合酶链式反应（RT-PCR）验证miR-145在正常葡萄膜组织,葡萄膜黑色素瘤组织和葡萄膜黑色素瘤细胞系中的表达水平。使用慢病毒表达系统构建了稳定过表达miR-145的葡萄膜黑色素瘤细胞系MUM-2B和OCM-1。使用MTT方法检测细胞增殖。使用流式细胞术分析细胞周期和细胞凋亡。通过生物信息学预测miR-145的靶基因,并通过双荧光素酶报告基因检测来验证。使用Western blot检测潜在的miR-145靶蛋白IGF-1R和IRS-1的表达。在OCM-1细胞系中敲除IRS-1基因通过TUNEL, BrdU以及流式细胞术验证其在细胞中的功能。 结果：与正常葡萄膜组织样本比较,葡萄膜黑色素瘤组织中发现了47种miRNA上调,61种miRNA下调。其中miR-145在葡萄膜黑色素瘤组织和细胞系中表达比正常葡萄膜组织显著下调。在葡萄膜黑色素瘤细胞系中过表达miR-145可以抑制细胞增殖,并将阻断细胞从G1期进入S期,同时还可以促进肿瘤细胞的凋亡。鉴定了IRS-1可能为miR-145在葡萄膜黑色素瘤中的靶基因。敲除IRS-1基因与过表达miR-145具有类似的效果。 结论：我们的数据首次表明了miR-145可能通过下调IRS-1而抑制葡萄膜黑色素瘤细胞增殖。     

Intermittent hypoxia with or without hypercapnia is associated with tumorigenesis by decreasing the expression of brain derived neurotrophic factor and miR-34a in rats

Background Very recent studies revealed that obstructive sleep apnoea （OSA） is a contributor of the increased incidence and mortality of cancer in humans,but mechanisms of how OSA promotes tumorigenesis remains largely unknown.We investigated whether intermittent hypoxia with and without hypercapnia plays a role in tumorigenesis.Methods First,Sprague-Dawley （SD） male rats （12 weeks old） were subjected to different hypoxia exposures：intermittent hypoxia and intermittent hypoxia with hypercapnia; continuous hypoxia and normal air.The systemic application of chronic fast rate hypoxia with or without hypercapnia mimicked severe OSA patients with apnoea/hypopnea index equivalent to 60 events per hour.Then routine blood tests were performed and the levels of brain derived neurotrophic factor （BDNF） and miR-34a were examined.Results In contrast to intermittent hypoxia with hypercapnia,both intermittent hypoxia and continuous hypoxia treatments caused significantly higher levels of haematology parameters than normoxia treatments.Compared to normoxia,intermittent hypoxia with hypercapnia exposure resulted in substantial decrease of serum BDNF and,miR-34a in the lower brainstem,while less pronounced results were found in intermittent hypoxia and continuous hypoxia exposure.Conclusions The exposure of intermittent hypoxia with or without hypercapnia,mimicking the situations in severe OSA patients,was associated with,or even promoted tumorigenesis.     

miR-9对神经纤毛蛋白-1的靶向调控及其在辐射效应中的作用

目的：构建has-miR-9表达质粒和NRP1-3’UTR荧光素酶报告质粒,探讨miR-9对 NRP1的靶向调控作用及其在A549 细胞中的辐射效应。方法：利用生物信息学方法预测has-miR-9与NRP1-3’UTR的结合位点;将miR-9序列插入载体pcDNA-DEST47中构建真核表达质粒,同时构建NRP1-3’UTR的野生型和突变型荧光素酶报告质粒,并与pcDNA-DEST-miR-9质粒共转染至A549细胞,分析miR-9对其调控作用;miR-29b作为阴性对照,观察miR-9对NRP1的靶向作用。采用Western blot方法,验证miR-9对NRP1蛋白表达的抑制作用及照射后A549细胞NRP1蛋白表达变化;采用实时定量PCR方法检测10 Gy电离辐射照射后A549细胞miR-9表达量。结果：荧光素酶活性实验结果显示,miR-9可以显著下调野生型NRP1-3’UTR质粒的荧光素酶活性（t=3.906,P<0.05）,而不影响突变型质粒的荧光素酶活性,同时证实miR-9以外的miRNA（miR-29b）不能抑制野生型NRP1-3’UTR质粒的荧光素酶活性。转染miR-9 mimic后,在A549细胞中,靶基因NRP1蛋白的表达受抑制。10 Gy照射后,A549细胞中miR-9表达量下调（t=37.319,P<0.05）,而NRP1蛋白表达升高。结论：在A549辐射效应中miR-9通过靶向结合NRP1基因3’UTR,特异性调控NRP1蛋白表达。     

Serum MicroRNA-210 as a Predictive Biomarker for Treatment Response and Prognosis in Patients with Hepatocellular Carcinoma undergoing Transarterial Chemoembolization

PurposeTo investigate whether serum microRNA-210 (miR-210) level can serve as an indicator of prognosis and a predictor of efficacy of transarterial chemoembolization in patients with hepatocellular carcinoma (HCC).Materials and MethodsSerum miR-210 level was measured in 113 patients with HCC before transarterial chemoembolization (t1), 3 days after transarterial chemoembolization (t2), and 4 weeks after transarterial chemoembolization (t3) and compared with 39 healthy control subjects. The correlations between miR-210 levels and clinicopathologic factors, tumor responsiveness, and prognosis were analyzed. The modified Response Evaluation Criteria in Solid Tumors assessment was conducted at t3.ResultsA higher mean baseline miR-210 level was observed in patients with HCC compared with control subjects (3.69 ± 2.04 vs 1.08 ± 0.45, P < .001). A positive correlation between baseline miR-210 level and tumor size (P < .001), vascular invasion (P = .005), tumor differentiation (P = .037), and Barcelona Clinic Liver Cancer stage (P < .001) was observed. Elevated baseline miR-210 level also served as an independent prognostic factor predicting poor overall survival (risk ratio, 2.082; P = .003). Patients who did not respond to transarterial chemoembolization had higher baseline miR-210 levels than patients who did respond to treatment (4.34 ± 1.67 vs 3.28 ± 2.15, P < .001). In addition, miR-210 levels increased significantly 4 weeks after transarterial chemoembolization in nonresponders (5.79 ± 2.06 at t3 vs 4.34 ± 1.67 at t1, P < .001), whereas no significant change was observed in responders (3.53 ± 2.20 at t3 vs 3.28 ± 2.15 at t1, P = .116). Lastly, an inverse correlation was identified between miR-210 change t1–t3 with the time to radiologic progression (P < .001).ConclusionsSerum miR-210 may represent a novel biomarker for predicting efficacy of transarterial chemoembolization and overall survival for patients with HCC.     

CircRNA pappalysin 1 facilitates prostate cancer development through miR-515-5p/FKBP1A axis

The role of circular RNA (circRNA) pappalysin 1 (circ-PAPPA; hsa_circ_0088233) in prostate cancer (PCa) cells was explored in the current study. Circ-PAPPA abundance was markedly enhanced in PCa. Circ-PAPPA interference restrained cell viability, proliferation, motility and glycolysis while elevated the apoptosis rate of PCa cells. Circ-PAPPA negatively regulated microRNA-515-5p (miR-515-5p) abundance. MiR-515-5p silencing largely diminished circ-PAPPA knockdown-mediated effects in PCa cells. MiR-515-5p directly bound to FKBP prolyl isomerase 1A (FKBP1A). MiR-515-5p overexpression-mediated impacts were partly counteracted by FKBP1A overexpression. Circ-PAPPA silencing reduced FKBP1A protein level partly by elevating miR-515-5p expression. Circ-PAPPA knockdown significantly restrained the tumour growth in vivo. Circ-PAPPA elevated the malignant phenotypes of PCa cells by sequestering miR-515-5p to induce the expression of FKBP1A.     

miR-29a抑制胃癌细胞内靶基因VEGF-A的表达及意义

目的 验证胃癌细胞株SGC-7901内miR-29a对靶基因VEGF-A的直接调控作用.方法 采用负载miR-29a的腺病毒载体（Ad-miR29a）感染人胃癌细胞株SGC-7901,上调SGC-7901细胞内miR-29a的表达丰度后,采用RT-qPCR及Western blot法分别检测SGC-7901细胞内VEGF-A在mRNA及蛋白水平的表达;进一步采用双荧光素酶实验及突变实验,验证miR-29a是否通过结合在VEGF-A 3＇UTR而直接抑制靶基因表达.结果 与感染阴性对照Ad-LacZ的SGC-7901细胞相比,感染Ad-miR29a的SGC-7901细胞内VEGF-A蛋白表达下降（0.42±0.02 vs.0.91±0.03,P＜0.01）,且VEGF-A mRNA水平亦下调（0.75±0.21 vs.1.15±0.25,P＜0.05）;荧光素酶实验显示,与阴性对照相比,转染miR-29a mimic后,HEK293细胞萤火虫荧光素酶活性明显下降（0.56±0.13 vs.0.93±0.06,P＜0.05）;而在VEGF-A 3＇UTR与miR-29a的结合位点被突变之后,HEK293细胞萤火虫荧光素酶活性得以恢复.结论 miR-29a通过结合于VEGF-A 3＇UTR相应位点,直接抑制VEGF-A在mRNA及蛋白水平的表达.miR-29a可能成为胃癌基因治疗的有效靶标.     

miR-212在老年前列腺癌患者中的表达及其对癌细胞增殖、侵袭、转移的影响机制研究

目的探讨miR-212在老年前列腺癌患者中的表达及其对癌细胞增殖、侵袭、转移的影响机制。方法选择2017年10月至2019年10月于厦门市第五医院行手术确诊的前列腺癌患者60例,经病理档案室收集其癌组织标本及配对癌旁正常组织标本,采用实时荧光定量PCR检测前列腺组织miR-212的相对表达水平。将购自上海北诺生物科技有限公司的前列腺癌PC-3细胞24株随机分为空白组(不作任何处理)、对照组(转染空白miR-212对照)、miR-212组(转染miR-212 mimics),每组各8株;检测并比较3组转染后前列腺癌细胞增殖率、侵袭能力和转移率;采用荧光素酶报告实验验证miR-212下游靶基因。结果miR-212在前列腺癌组织中呈低表达,在癌旁正常组织中呈高表达,前列腺癌组织中的miR-212相对表达水平低于癌旁正常组织(P<0.05)。转染miR-212后,PC-3细胞增殖、侵袭、转移均受到抑制。miR-212组PC-3细胞增殖率、侵袭能力、转移率低于对照组、空白组(P<0.05);空白组和对照组比较,PC-3细胞增殖率、侵袭能力、转移率差异无统计学意义(P>0.05)。miR-212组与上皮-间质转化(EMT)-WT(野生型)共转染后细胞荧光活性显著低于对照组与EMT-WT共转染后细胞荧光活性(P<0.05);空白组与对照组比较,与EMT-WT共转染后细胞荧光活性差异无统计学意义(P>0.05)。空白组、对照组、miR-212组与EMT-MUT(突变型)共转染后细胞荧光活性比较,差异无统计学意义(P>0.05)。结论miR-212在前列腺癌组织中呈低表达,上调miR-212的相对表达水平可抑制前列腺癌细胞增殖、侵袭、转移能力,其作用与EMT密切相关。