miR-135a在调节小细胞肺癌多药耐药中的作用

目的:探讨miR-135a在小细胞肺癌(small cell lung cancer,SCLC)多药耐药中的作用。方法:收集2008年1月至2013年12月本院肿瘤科及呼吸内科就诊的48例SCLC患者外周血液标本,将其分为化疗敏感组(28例)和耐药组(20例),采用QRT-PCR法检测患者血液标本、SCLC敏感细胞株H69及耐药细胞株H69AR中miR-135a的表达,分析其与临床预后的关系,并应用miR-135a模拟物和抑制剂分别上、下调细胞株中miR-135a的表达,采用CCK-8法检测细胞对各种化疗药物(ADM、DDP和VP-16)敏感性的影响。结果:与化疗敏感组相比,miR-135a在化疗耐药患者血液标本中的表达明显增高[(2.174±3.981)vs(-1.963±3.750),P<0.001];在临床患者血液标本中,miR-135a的表达与患者的性别、年龄无关(P>0.05),与疾病的分期、化疗敏感性及患者的生存时间相关(均P<0.01)。miR-135a在H69AR细胞中的表达明显高于H69细胞[(7.841±0.392)vs(1.047±0.081),P<0.01];通过miR-135a抑制剂下调H69AR中miR-135a的表达能够显著增加细胞对化疗药物的敏感性,H69AR细胞对DDP、ADM及VP-16的IC50值明显下降[(247.09±11.55)vs(76.64±10.00)、(150.83±8.03)vs(40.72±5.06)、(436.63±61.19)vs(163.35±20.00);H69细胞转染miR-135a mimics后对化疗药物DDP、ADM和VP-16的IC50值明显增加[(18.58±1.37)vs(159.27±3.29)、(24.37±2.63)vs(129.19±2.64),(48.55±2.59)vs(359.87±2.92)μg/ml,P<0.01]。结论:miR-135a参与调节SCLC的多药耐药,下调miR-135a基因的表达可能增加细胞对化疗药物的敏感性。     

miR-34a调控肺癌EGFR耐药细胞株中MET信号通路探讨

[目地]研究肺癌耐药细胞株HCC827/GR中MET信号通路的调控,明确肺癌获得性耐药的分子机制。[方法]使用HCC827细胞[epidermal growth factor receptor（EGFR）基因19外显子缺少的肺腺癌细胞株],在此细胞的基础上培养吉非替尼耐药细胞株。检测耐药细胞株中MET的表达,并使用RT-PCR的方法检测miR-34a的表达;使用TargetScan等生物信息学软件预测miR-34a的下游靶点,并在细胞内验证miR-34a是否可以调控MET的表达。[结果]与HCC827细胞相比,HCC827/GR细胞株对吉非替尼明显耐药。在耐药HCC827/GR细胞株中,miR-34a低表达,MET高表达,而在HCC827细胞株中,miR-34a高表达,MET低表达。Target Scan生物信息学软件提示,MET是miR-34a的下游靶点之一。将携带荧光报告基团和MET 3′UTR的载体与miR-34a共转染入细胞后荧光度下降。[结论]miR-34a可能通过调控靶基因MET而参与EGFR-TKI的获得性耐药。     

大肠癌多药耐药基因的定量分析及其临床意义

目的：探讨多药耐药（ｍｕｌｔｉｄｒｕｇ ｒｅｓｉｓｔａｎｃｅ，ＭＤＲ）基因及其编码的糖蛋白（Ｐ－ｇｌｙｃｏｐｒｏｔｅｉｎ，Ｐ－ｇｐ）在结直肠癌中的表达及其临床意义。方法：采用逆转录－聚合酶链式反应（ＲＴ－ＰＣＲ）方法定量分析ｍｄｒ１基因在不同分化程度、不同Ｄｕｋｅ’ｓ分期的癌组织、癌旁组织和正常组织中的表达量；应用免疫组化方法检测相应的Ｐ－ｇｐ表达率。结果：大肠癌组织、癌旁组织正常组织的ｍｄｒ１基     

ERK信号传导抑制逆转大肠癌多药耐药的研究

促分裂原激活的蛋白激酶(mitogen-activated protein kinase,MAPK)的信号传导途径,是细胞生长和分化的重要环节.Ras/Raf-1/Mek1/2/Erk1/2,即ERK1/2信号传导通路,是MAKP众多通路中的一个.肿瘤细胞在化疗药物作用下,出现P-糖蛋白(P-glycoprotein,P-gp)表达增加的同时,也发现了MAPK活性的增加.     

骨肉瘤患者血清中miR-34a、miR-449a表达及与新辅助化疗耐药的关系

目的 分析骨肉瘤患者血清中miR-34a、miR-449a表达水平及与新辅助化疗耐药的关系.方法 随机选择骨肉瘤83例,将其纳入研究组.另选取同期体检健康受试者83例,将其纳入对照组.检测血清miR-34a、miR-449a表达水平.问卷调查骨肉瘤患者临床病理特征,给予新辅助化疗,统计耐药情况.结果 研究组患者血清mi...     

miR-135a 通过下调SOX2 抑制喉癌Hep-2 细胞的恶性生物学行为和增强对奥沙利铂的敏感性

[摘要] 目的：探讨敲降miR-135a 对人喉癌上皮Hep-2 细胞的恶性生物学行为和奥沙利铂敏感性的影响。方法：收集2018年1 月至2018 年6 月郑州大学附属南阳医院南阳市中心医院行喉癌切除术10 例患者的喉癌组织和癌旁组织标本。采用qPCR检测喉癌组织和Hep-2 细胞中miR-135a 的表达水平。miR-135 inhibitor 转染喉癌Hep-2 细胞后,采用CCK-8 检测Hep-2 细胞活性,集落形成实验检测Hep-2 细胞集落形成能力,Transwell 实验检测Hep-2 细胞的侵袭及迁移能力,WB实验检测Hep-2 细胞中SOX2蛋白的表达水平。0.5、1.0、1.5、2.0 μmol/L的奥沙利铂处理已转染miR-135 inhibitor 的Hep-2 细胞,CCK-8 实验检测Hep-2 细胞的增殖活性,Annexin-V-FITC/PI 染色流式细胞术检测Hep-2 细胞的凋亡率。采用miR-135a inhibitor 质粒、对照pcDNA 空载体（SOX2-Con）质粒、pcDNA-SOX2（SOX2-OE）质粒共转染Hep-2 细胞,构建miR-135a inhibitor+SOX2-Con 组和miR-135a inhibitor+SOX2-OE组,检测此2 组细胞的增殖活性、集落形成能力、侵袭及迁移能力。结果：与癌旁组织比较,喉癌组织中miR-135a表达水平显著升高（P<0.01）;与正常NHP细胞比较,miR-135a 在Hep-2 细胞中表达水平明显上调（P<0.01）;转染miR-135a inhibitor导致Hep-2 细胞中miR-135a 表达水平明显降低（P<0.01）。敲降miR-135a 明显降低Hep-2 细胞的增殖活性、细胞集落数、迁移、侵袭和SOX2 表达（均P<0.01）;明显增强细胞对奥沙利铂敏感性（P<0.01）;与miR-135a inhibitor+SOX2-Con 组比较,miR-135a inhibitor+SOX2-OE 组的Hep-2 细胞的增殖活性、细胞集落数、迁移与侵袭能力均明显增加（均P<0.01）;同时,用不同浓度的奥沙利铂处理此2 组细胞,相对于miR-135a inhibitor+SOX2-Con组,miR-135a inhibitor+SOX2-OE组的Hep-2 细胞存活率显著升高（P<0.01）。结论：敲降miR-135a 可能通过下调转录因子SOX2 的表达抑制Hep-2 细胞的恶性生物学行为,并增强其对奥沙利铂的敏感性。     

miR-21通过靶向SOX2增强胃癌细胞的迁移能力

目的:探讨miR-21在胃癌细胞系中对SOX2的调控作用及其对胃癌细胞迁移的影响。方法:在胃癌细胞系AZ-521、AGS中通过转染miR-21 mimic或者miR-21 antagomir构建miR-21过表达和低表达模型,RT-qPCR检测转染效果,并通过蛋白免疫印迹法检测SOX2在转染细胞中表达的变化。构建miR-21双荧光素酶报告基因,在工具细胞HEK293T中检验miR-21是否通过直接结合SOX2的3′-UTR区域来对其实现调控。应用Transwell迁移实验检测miR-21及SOX2对胃癌细胞迁移能力的影响。结果:miR-21直接结合于SOX2的3′-UTR区域来抑制SOX2在胃癌细胞系中的表达。迁移实验发现上调miR-21或者下调SOX2都能促进胃癌细胞的迁移。结论:miR-21可促进胃癌细胞的迁移能力,其机制可能通过抑制SOX2的表达而实现。     

miR-103与胃癌多药耐药作用机制探讨

目的 既往研究肿瘤耐药均集中在DNA水平,而DNA又影响mRNA及蛋白质表达.但是mRNA与编码蛋白质的表达并不成比例,而非编码RNA恰好能解释两者之间的不平衡.非编码RNA即微小RNA(microRNA,miRNA),是一类内源性短链非编码小分子核糖核苷酸,长度约18~25个核糖核苷酸,是转录后基因表达调节的关键分子,参与胃癌的发生发展.本研究旨在探讨miR-103在胃癌多药耐药中作用及其分子机制.方法 miRNA芯片筛选SGC7901/ADR及SGC7901细胞差异表达miRNA,应用qRT-PCR验证miR-103表达;将miR-103的拟似物及抑制物分别转染SGC7901/ADR及SGC7901细胞,MTT法检测转染前后对多柔比星敏感性变化;蛋白质印迹法检测miR-103对caveolin-1表达的影响.结果 miR-103在SGC7901/ADR细胞中表达(0.32±0.04)显著低于SGC7901细胞.miR-103拟似物转染SGC7901/ADR细胞后对多柔比星的敏感性较对照组显著提高,IC50由(18.83±0.32) μg/mL下降到(4.54±0.29) μg/mL,t=1.04,P<0.05;而miR-103抑制物转染SGC7901细胞后对多柔比星的敏感性显著降低,IC50由(1.65±0.03) μg/mL上升到(15.27±0.26) μg/mL,t=1.25,P<0.05.miR-103靶向作用于caveolin-1的3'-UTR,并在转录后水平负向调控caveolin-1表达.结论 miR-103通过靶向负调控caveolin-1表达,增加SGC7901/ADR细胞对多柔比星敏感性,从而逆转胃癌多药耐药.     

miR-187通过抑制胃癌细胞凋亡影响胃癌多药耐药

目的:探讨miR-187对胃癌多药耐药的影响。方法:应用qPCR检测miR-187在胃癌SGC7901细胞和SGC7901ADR细胞中的表达。miR-187 mimics转染上述细胞,MTT检测转染后细胞对多种化疗药物的敏感性,流式细胞仪检测细胞凋亡。结果:SGC7901ADR细胞中miR-187的表达明显高于SGC7901细胞。MTT实验显示转染miR-187 mimics后胃癌细胞对化疗药物的敏感性均明显降低(P＜0.05)。流式细胞检测表明miR-187 mimics转染可以减少SGC7901细胞的凋亡(P＜0.05)。结论:miR-187通过抑制胃癌细胞凋亡参与胃癌多药耐药,可能成为逆转胃癌多药耐药的新靶点。     

miR-34b和miR-320a的表达与前列腺癌组织临床病理特征及预后的相关性

目的 探讨前列腺癌组织中miR-34b、miR-320a的表达与其临床病理特征及预后的相关性.方法 选择前列腺癌患者80例,另选40例良性前列腺增生患者作为对照组.采用定量聚合酶链式反应(RT-PCR)检测两组患者组织中miR-34b、miR-320a的表达量,分析前列腺癌组织中miR-320a、miR-34b与各临床...     

Long noncoding RNA SOX2OT promotes the proliferation of pancreatic cancer by binding to FUS

Pancreatic ductal adenocarcinoma (PDAC) is one of the most malignant tumors has one of the worst prognoses, and the role of long noncoding RNAs (lncRNAs) in the biological and pathological processes of pancreatic cancer, including tumor cell proliferation, is a popular topic in tumor research. Our previous study revealed the correlation between high levels of the lncRNA-SOX2OT (SOX2OT) with poor survival outcomes. Cell Counting Kit-8, EdU, Flow cytometry and Colony formation assays as well as Xenograft growth of PDAC cells in mice were used for the detection of PDAC cells proliferation progression. Fluorescence in situ hybridization, RNA-binding protein pulldown and RNA immunoprecipitation assays were also used to identify the putative mechanisms of SOX2OT participating in the tumor progression. SOX2OT and its potential downstream targets were verified by Western blot and quantitative real-time polymerase chain reaction (qRT-PCR). SOX2OT was confirmed to promote the proliferation of PDAC cells. It was found to directly physically bind to FUS and we also demonstrated that FUS protein stability was affected by binding with SOX2OT and FUS could suppressed PDAC tumor by regulating cell cycle-associated factors CCND1 and p27. Our findings suggest that SOX2OT may act as a tumor promoter in PDAC through physically binding FUS and regulating its downstream cell cycle-associated factors CCND1 and p27. It may serve as an effective target for antitumor treatment for pancreatic cancer.     

SOX2蛋白在结直肠癌中的表达及临床意义

目的 研究SOX2蛋白在结直肠癌中的表达及临床意义.方法 收集80例结直肠癌手术标本及20例正常的肠黏膜标本,采用免疫组化法检测SOX2蛋白在标本中的表达情况.结果 SOX2蛋白在正常肠黏膜中不表达,在结直肠癌中高表达,表达率为41.3％.SOX2表达水平与TNM分期、肿瘤浸润深度、淋巴结转移、组织学分化程度、Ki-6...     

The SOX17/miR-371-5p/SOX2 axis inhibits EMT,stem cell properties and metastasis in colorectal cancer

Cancer stem cells (CSCs) and EMT-type cells, which share molecular characteristics with CSCs, have been believed to play critical roles in tumor metastasis. Although much progress has been garnered in elucidating the molecular pathways that trigger EMT, stemness and metastasis, a number of key mechanistic gaps remain elusive. In the study, miR-371-5p was obviously down-regulated in primary CRC tissues compared with matched adjacent normal mucosa and correlated significantly with differentiation, tumor size, lymphatic and liver metastases. MiR-371-5p could attenuate proliferation, invasion in vitro and metastasis in vivo in CRC cells. It also suppressed EMT by regulating Wnt/β-catenin signaling and strongly decreased the CRC stemness phenotypes. Moreover, demethylation of SOX17 induced miR-371-5p expression and consequently suppressed its direct target SOX2 in CRC cells. MiR-371-5p was necessary for SOX17 mediated cancer-related traits and SOX2 was a functional target of miR-371-5p. A positive relationship between SOX17 and miR-371-5p expression and a negative one between miR-371-5p and SOX2 expression were observed in CRC cell lines and tissues. In conclusion, we identified miR-371-5p as an important “oncosuppressor” in CRC progression and elucidated a novel mechanism of the SOX17/miR-371-5p/SOX2 axis in the regulation of EMT, stemness and metastasis, which may be a potential therapeutic target.     

结直肠癌中医辨证分型与miR-29、miR-34a表达的相关性分析

目的:探究结直肠癌患者血清miR-29、miR-34a表达水平及其与中医辨证分型的相关性。方法:选取2014年06月至2015年06月本院收治的127例结直肠癌患者作为研究对象（结直肠癌组）,并根据中医辨证分型标准分为湿热内蕴型21例、气滞血瘀型40例、脾肾阳虚型29例、气血两虚型18例、肝肾阴虚型19例;另选取同期在本院体检的127例健康者作为健康对照组。采用实时荧光定量PCR（qRT-PCR）技术检测血清miR-29、miR-34a表达水平,并进行比较;采用Pearson法分析血清miR-29与miR-34a表达水平的相关性;采用Logistic回归模型分析影响结直肠癌患者不良预后发生的危险因素。结果:结直肠癌组患者血清miR-29、miR-34a表达水平均明显低于健康对照组（P＜0.05）。低分化、浸润程度T3-T4、TNM分期Ⅲ-Ⅳ期、淋巴结转移、远处转移患者血清miR-29、miR-34a表达水平明显低于高中分化、浸润程度T1-T2、TNM分期Ⅰ-Ⅱ期、无淋巴结转移、无远处转移患者（P＜0.05）。结直肠癌患者血清miR-29与miR-34a表达水平呈正相关（r=0.529,P=0.000）。低分化、浸润程度T3-T4、TNM分期Ⅲ-Ⅳ期、淋巴结转移、低miR-29水平、低miR-34a水平是影响结直肠癌患者不良预后发生的危险因素（P＜0.05）。脾肾阳虚型、气滞血瘀型、湿热内蕴型预后不良患者比例依次明显增加（P＜0.05）。肝肾阴虚型、气血两虚型、脾肾阳虚型、气滞血瘀型、湿热内蕴型结直肠癌患者血清miR-34a表达水平依次显著降低（P＜0.05）,miR-29表达水平差异无统计学意义（P＞0.05）。结论:结直肠癌患者血清中miR-29、miR-34a低表达,与肿瘤分期升高、分化程度降低、浸润程度增加、淋巴结转移、远处转移、不良预后等密切相关,且miR-34a对结直肠癌不同中医辨证分型有一定的分型参考意义。     

miR-34a对结肠癌细胞SW480增殖、迁移和侵袭能力的影响及机制探讨

目的 探讨miR-34a对结肠癌细胞株SW480增殖、侵袭和迁移能力的影响及可能的作用机制。方法将miR-34a过表达慢病毒、空病毒载体转染SW480细胞,未做处理细胞作为空白对照组。Real-time PCR检测各组细胞内miR-34a的表达;CCK8法检测细胞增殖能力;划痕实验、Transwell小室实验检测细胞迁移和侵袭能力;Western blot实验检测细胞内E-cadherin和Vimentin蛋白表达。结果 与空病毒载体组及空白对照组相比,转染组细胞中miR-34a的表达增高,且细胞增殖效率、侵袭和迁移能力降低(P＜0.05),miR-34a使E-cadherin蛋白表达增加、Vimentin蛋白表达降低。结论 miR-34a可抑制结肠癌细胞SW480增殖、侵袭和迁移能力,并能影响E-cadherin和Vimentin的表达,miR-34a有望成为干预结肠癌转移和复发的分子靶点。     

SOX2在结直肠癌表达的临床观察

目的：观察SOX2在结直肠癌组织的表达情况,探讨SOX2在结直肠癌中表达的临床意义。方法收集103例有60～125个月（中位随访时间84个月）随访资料的结直肠癌及其相应的正常结直肠黏膜标本,应用免疫组化方法检测SOX2的表达,分析SOX2表达与临床特征的关系。结果结直肠癌组织中SOX2蛋白的表达率显著高于癌旁正常肠黏膜（P＜0.001）;SOX2表达与结直肠癌分化程度显著负相关（P＜0.01）,与结直肠癌TNM分期、淋巴结转移个数及远处转移程度正相关（P＜0.05）。SOX2表达与结直肠癌患者术后5年生存率和3年无瘤生存率也具有相关性（P＜0.05）。结论 SOX2蛋白异常表达可作为提示结直肠癌预后判断的重要参考指标。     

miR-497 and miR-34a retard lung cancer growth by co-inhibiting cyclin E1 (CCNE1)

Cyclin E1, encoded by the CCNE1 gene, promotes G1/S transition, chromosome instability, and oncogenesis. Here, we show that miR-497 and miR-34a target the 3′-UTR of CCNE1. miR-497 and miR-34a are downregulated in cancer cells and their ectopic expression inhibited cell proliferation and colony formation in vitro, and inhibited tumor growth in a xenograft model. The effect of simultaneous overexpression of miR-497 and miR-34a on the inhibition of cell proliferation, colony formation, and tumor growth, and the downregulation of cyclin E1 was stronger than the effect of each miRNA alone. The synergistic actions of miR-497 and miR-34a partly correlated with cyclin E1 levels. When cells stably expressing CCNE1 were transfected with the Hi-miR-497/34a plasmid, there was no effect on colony formation, compared with that of cells transfected with either Hi-miR497 or Hi-miR34a. These results indicate cyclin E1 is downregulated by both miR-497 and miR-34a, which synergistically retard the growth of human lung cancer cells.     

miR－320a表达与结直肠癌患者预后的关系

目的：探讨miR－320a与结直肠癌患者的临床病理特征及预后的相关性。方法选取2010年1月至2013年3月在本科室收治的65例已经配对的癌旁正常组织和结直肠癌组织,采用实时定量PCR的方法检测癌旁正常组织和癌组织中miR－320a的表达情况,分析miR－320a表达情况与结直肠癌患者的临床病理特点及生存预后的关系。结果结直肠癌组织中miR－320a表达量明显低于癌旁正常组织（P＜0．001）。结直肠癌患者组织中miR－320a的表达与肿瘤分化程度、TNM分期和CEA水平有关（均P＜0．05）,而与年龄、性别、肿瘤部位无关（均P＞0．05）。生存分析结果显示miR－320a高表达的结直肠癌患者术后总体预后明显优于miR－320a低表达者,两组患者的5年生存率分别为54．7％和31．4％（P＝0．022）。结论 miR－320a 可能作为结直肠癌患者预后的预测指标。miR－320a过表达可能提示结直肠癌患者肿瘤生物学行为较佳。     

Dysregulation of miR-106a and miR-591 confers paclitaxel resistance to ovarian cancer

Background:

MicroRNAs are noncoding regulatory RNAs strongly implicated in carcinogenesis, cell survival, and chemosensitivity. Here, microRNAs associated with chemoresistance in ovarian carcinoma, the most lethal of gynaecological malignancies, were identified and their functional effects in chemoresistant ovarian cancer cells were assessed.

Methods:

MicroRNA expression in paclitaxel (PTX)-resistant SKpac sublines was compared with that of the PTX-sensitive, parental SKOV3 ovarian cancer cell line using microarray and qRT–PCR. The function of differentially expressed microRNAs in chemoresistant ovarian cancer was further evaluated by apoptosis, cell proliferation, and migration assays.

Results:

Upregulation of miR-106a and downregulation of miR-591 were associated with PTX resistance in ovarian cancer cells and human tumour samples. Transfection with anti-miR-106a or pre-miR-591 resensitized PTX-resistant SKpac cells to PTX by enhancing apoptosis (23 and 42% increase), and inhibited their cell migration (43 and 56% decrease) and proliferation (64 and 65% decrease). Furthermore, ZEB1 was identified as a novel target gene of miR-591, and BCL10 and caspase-7 were target genes of miR-106a, as identified by immunoblotting and luciferase assay.

Conclusion:

MiR-106a and miR-591 have important roles in conferring PTX resistance to ovarian cancer cells. Modulation of these microRNAs resensitizes PTX-resistant cancer cells by targeting BCL10, caspase-7, and ZEB1.