miR-223在结肠癌组织中的表达及促进结肠癌细胞迁移侵袭的机制研究

目的:探讨微小RNA-223 (mi R-223)在结肠癌组织中的表达及对结肠癌HT-29细胞侵袭、迁移能力的影响及机制。方法:检测mi R-223在结肠癌组织与癌旁组织中的表达。通过脂质体转染法将mi R-223模拟物(mi R-223 mimics,mi R-223 mimics组)及microRNA无关序列(mi R-223 NC,NC组)转染入结肠癌HT-29细胞。采用Real-time PCR检测转染后细胞中mi R-223和TWIST的表达,Western blot检测TWIST的蛋白表达,Tranwell检测细胞的迁移与侵袭能力。双荧光素酶报告基因检测mi R-223对TWIST基因启动子活性的影响。采用Transwell迁移与侵袭实验检测mi R-223 mimic及Twist si RNA共转染后人结肠癌细胞系HT-29迁移与侵袭能力的变化。结果:与癌旁结肠组织比较,mi R-223在结肠癌组织中呈现明显高表达(P0.05);与空白对照组和mi R-223 NC组比较,转染mi R-223 mimics后的HT-29细胞中的mi R-223表达显著增加(P0.05)。与阴性对照组和空载转染组相比较,mi R-223 mimics转染组穿透的细胞数目明显增加(P0.05),且mi R-223 mimics转染组的细胞侵袭能力显著增强(P0.05)。与mi R-223 NC组和空白对照组比较,转染mi R-223 mimics的HT-29细胞的TWIST基因m RNA和蛋白表达均显著增加(P0.05)。双荧光素酶检验结果显示TWIST为mi R-223的下游靶基因。共转染TWIST si RNA和mi R-223 mimics的结肠癌HT-29细胞的迁移与侵袭能力较单独转染mi R-223 mimics的HT-29细胞显著减弱(P0.05)。结论:mi R-223可能通过上调下游靶基因TWIST水平促进结肠癌HT-29细胞的迁移与侵袭。     

miR-448通过抑制上皮间质转化抑制肺癌细胞增殖和运动能力

探讨mi R-448对肺癌细胞增殖和运动的影响及其分子机制。采用实时荧光定量PCR(polymerase chain reaction)检测原发肺癌组织和癌旁正常组织mi R-448表达水平。转染mi R-448 mimic和inhibitor至肺癌A549细胞系,通过MTT(3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazoliumbromide)、平板克隆形成和Transwell实验观察mi R-448表达对A549增殖和运动能力的影响;利用Western blot检测EMT(epithelial-mesenchymal transition)标志物蛋白表达水平,通过实时荧光定量PCR检测EMT相关转录因子m RNA表达水平。实时荧光定量PCR显示较癌旁正常组织相比,mi R-448在原发肺癌组织中表达降低。MTT和平板克隆形成实验显示,过表达mi R-448抑制A549细胞增殖和运动能力;降表达mi R-448增强A549细胞增殖和运动能力。Western blot显示降表达mi R-448能下调上皮标志物E-cadherin,上调间质标志物Vimentin表达水平。实时荧光定量PCR显示降表达mi R-448能上调EMT相关转录因子Twist1和ZEB1 m RNA表达水平。mi R-448可通过抑制EMT抑制肺癌进展。     

非小细胞肺癌组织中miR-19b与miR-20a的表达及临床意义

目的:检测mi R-19b与mi R-20a在非小细胞肺癌(NSCLC)组织中的表达,探讨两者与肺癌临床病理的关系。方法:选择我院NSCLC肺癌患者50例,使用Real-time RT-PCR法对其癌组织及癌旁正常组织中mi R-19b和mi R-20a的含量进行检测,并利用2(-△△CT)法处理结果,分析与临床病理资料的关系。结果:相对于内参U6,mi R-19b基因在NSCLC组织中的表达量明显低于癌旁正常组织,而mi R-20a基因在NSCLC组织中的表达量明显高于癌旁正常组织,差异具有统计学意义(P0.05);在NSCLC组织标本中,mi R-19b基因表达量在临床分期I-II的标本中明显高于临床分期III,而mi R-20a基因表达量在临床分期I-II的标本中明显低于临床分期III,差异均就有统计学意义(P0.05);mi R-19b基因表达量在肿瘤病理低分化组织中明显低于肿瘤病理高分化组织,而mi R-20a基因表达量在肿瘤病理低分化组织中明显高于肿瘤病理高分化组织,差异均具有统计学意义(均P0.05)。结论:mi R-19b低表达和mi R-20a高表达与NSCLC的临床分期、病理分级具有密切关系,早期检测有助于NSCLC诊断和治疗。     

Periostin在非小细胞肺癌中的高表达可增强肺癌细胞的增殖和侵袭能力

为了研究非小细胞肺癌(non-small cell lung cancer,NSCLC)患者的血浆和肺癌组织中Periostin蛋白表达水平以及对癌细胞增殖和侵袭的影响。本研究选取了40例非小细胞肺癌患者作为研究组,同时选取40例同期体检的健康人群作为对照组,采用Real-time PCR的方法比较肺癌患者和健康人群的血浆中,以及肺癌患者的癌组织和癌旁正常组织中Periostin表达水平;采用小分子干扰RNA(small-interfering RNA,si RNA)抑制非小细胞肺癌细胞系A549中Periostin的表达,使用CCK-8法检测A549增值能力的变化,使用Transwell方法观察A549侵袭能力的变化。研究结果显示,Periostin蛋白在非小细胞肺癌患者的血浆中表达水平显著高于正常人群(p0.05),同时在肺癌组织中的含量显著高于癌旁组织(p0.05);导入Periostin的si RNA后,A549细胞的增殖和侵袭能力显著下降(p0.05)。本研究表明,Periostin在非小细胞肺癌患者的血浆和肺癌组织中表达量提高,可以增强肺癌细胞的增殖和侵袭能力。     

外周血miR-17-92基因簇对于早期胃癌的诊断及预测价值

目的:探讨外周血mi R-17-92簇对早期胃癌的诊断价值,为胃癌的早期诊断及治疗提供参考依据。方法:收集胃癌125例(Ⅰ期35例,Ⅱ期28例,Ⅲ期39例,Ⅳ期23例)和癌前病变24例(包括肠化生及上皮内瘤变),同时选择65例慢性胃炎作为对照组。采用实时荧光定量PCR技术(Real-time quantitative PCR,RT-qPCR)检测患者血清中的mi R-17-92基因簇的表达水平。通过受试者工作曲线(Receiver Operating Curve, ROC)及曲线下的面积(Area Under the Curve,AUC)评估mi R-17-92基因簇表达水平诊断早期胃癌的敏感性和特异性。结果:(1)慢性胃炎与癌前病变mi R-17-92基因簇表达比较无显著差异(P0.05);(2)早期胃癌及进展期胃癌mi R-17-5p表达明显高于慢性胃炎(P0.05),mi R-19a-3p、mi R-19b-3p、mi R-20a-5p和mi R-92a-3p表达则显著低于慢性胃炎及进展期胃癌(P0.05);(3)miR-17-5p诊断早期胃癌的曲线下面积较mi R-19a-3p、mi R-19b-3p、mi R-20a-5p、mi R-92a-3p及CEA更高;(4)miR-19a-3p、mi R-19b-3p、mi R-20a-5p、mi R-92a-3p高低表达组与在胃癌的浸润深度间有显著性差异(P0.05),mi R-19b-3p高低表达组在胃癌的临床分期间有显著性差异(P0.05);(5)miR-17-5p、mi R-19a-3p、mi R-19b-p、mi R-20a-5p、mi R-92a-3p诊断早期胃癌的阳性率较CEA、CA199高。结论:外周血mi R-17-92基因簇对于早期胃癌的诊断价值明显优于CEA和CA199,这可能为胃癌的早诊早治提供新的策略。     

miR-106b-93-25簇对子宫内膜癌细胞的调控作用研究

目的:检测mi R-106b-93-25基因簇对子宫内膜癌细胞增殖及凋亡的影响,并探讨其机制。方法:q RT-PCR检测临床子宫内膜癌标本及癌旁正常组织中mi R-106b、mi R-93和mi R-25及其宿主基因MCM7的表达情况。将micro RNA及其拮抗剂转染ECC-1细胞后,MTT实验检测ECC-1细胞增殖情况,流式细胞术检测ECC-1细胞周期及细胞凋亡情况。荧光素酶报告系统验证mi R-106b和mi R-25分别直接调控p21和Bim。结果:临床标本子宫内膜癌组织与癌旁正常组织相比mi R-106b-93-25簇及其宿主基因MCM7的表达明显增高。mi R-106b-93-25簇能够促进ECC-1细胞增殖,减少凋亡。转染mi R-106b和mi R-93的细胞出现明显的S期阻滞,过表达mi R-25的细胞凋亡明显减少。mi R-106b-93-25簇通过抑制靶基因p21和Bim的表达,引起促增殖、抗凋亡作用。结论:mi R-106b-93-25簇能够促进子宫内膜癌细胞增殖,抑制凋亡,并使细胞发生S期阻滞。mi R-106b-93-25簇在子宫内膜癌的发生与发展中具有重要的作用。     

Hsa-miR-5195-3p对人宫颈癌细胞SiHa增殖、迁移与侵袭的影响

目的:探讨mi R-5195-3p对人宫颈癌细胞系Si Ha增殖、迁移与侵袭的影响。方法:采用qRT-PCR检测人宫颈癌细胞SiHa和正常上皮细胞HaCaT中mi R-5195-3p的表达水平。将mi R-5195-3p mimic转染至Si Ha细胞中构建外源性过表达细胞株,阴性对照组中则转染NC mimic,并用q RT-PCR验证转染效率;通过MTT和集落形成实验检测细胞增殖能力;划痕愈合实验检测细胞横向迁移能力; Transwell小室实验检测细胞纵向迁移能力和侵袭能力;采用qRT-PCR和Western blot检测E-cadherin、Vimentin与snail m RNA转录水平及蛋白表达水平。结果:宫颈癌细胞Si Ha中的mi R-5195-3p表达水平较HaCaT偏低(P 0. 05)。与阴性对照组相比,转染mi R-5195-3p mimic的SiHa细胞中mi R-5195-3p水平显著增高(P 0. 01);并且其体外增殖(P 0. 001),迁移(P 0. 001)与侵袭能力(P 0. 001)明显减弱;同时E-cadherin表达水平上调而Vimentin、snail表达水平下调。结论:过表达mi R-5195-3p可能通过阻碍EMT通路抑制宫颈癌细胞Si Ha的增殖,迁移与侵袭。     

食道癌组织miR-21、miR-182表达与临床病理特征及预后的关系

目的:探讨食道癌组织微小RNA-21(miR-21)、微小RNA-182(miR-182)表达与临床病理特征及预后的关系。方法:选取2011年4月到2013年7月期间在我院接受手术治疗的食道癌患者84例,取患者的癌组织和癌旁正常组织作为检验标本,比较癌组织和癌旁正常组织中miR-21、mi R-182的表达水平,并分析食道癌组织中mi R-182、mi R-21的表达与临床病理特征及预后的关系。结果:癌组织中mi R-21、mi R-182的相对表达量明显高于癌旁正常组织,差异有统计学意义(P<0.05)。食道癌组织中mi R-21的表达与淋巴结转移、临床分期有关(P<0.05),与性别、年龄、分化程度、肿瘤大小无关(P>0.05);食道癌患者癌组织中miR-182的表达与年龄、性别、肿瘤大小无关(P>0.05),与分化程度、临床分期、淋巴结转移有关(P<0.05)。食道癌癌组织中miR-21、miR-182高表达患者的中位生存时间均低于低表达患者,差异有统计学意义(P<0.05)。结论:食道癌组织mi R-21、mi R-182表达与患者的部分临床病理特征及预后有关,两者有望成为食道癌新的治疗靶点。     

孕激素诱导的microRNA-1a抑制子宫内膜上皮细胞增殖

本文旨在研究孕激素诱导micro RNA-1a(mi R-1a)在子宫内膜上皮细胞(endometrial epithelial cells,EECs)的表达及其对EECs增殖的作用。用雌激素(E2组)或雌、孕激素联合(E2P4组)处理去卵巢小鼠,q PCR检测mi R-1a的成熟体mi R-1a-3p的表达;将mi R-1a-3p拟似剂或抑制剂注入各组小鼠的一侧子宫角,另一侧注入拟似剂或抑制剂对照,流式细胞术检测mi R-1a-3p对EECs细胞周期的影响;免疫组化检测mi R-1a-3p对细胞周期蛋白cyclin D2、cyclin E1和cyclin E2表达的影响。在体外实验,将分离培养的原代小鼠EECs分为两组,分别给予雌激素处理(E2组)和雌、孕激素联合处理(E2P4组),q PCR检测mi R-1a-3p的表达;再用mi R-1a-3p拟似剂作用于雌激素预处理细胞,对照组给予非特异性拟似剂,Ed U掺入实验检测细胞增殖活性。体内q PCR结果显示,E2P4组小鼠EECs mi R-1a-3p的表达量相比E2组明显增加(P0.05)。流式细胞术结果显示mi R-1a-3p拟似剂能够将E2处理细胞的细胞周期进程阻滞在G1/S期(P0.05);mi R-1a-3p抑制剂能够部分逆转P4对细胞周期进程的G1/S期阻滞(P0.05)。小鼠子宫组织免疫组化结果显示mi R-1a-3p拟似剂能较明显地抑制cyclin E1和cyclin E2蛋白的表达(P0.05),而对cyclin D2蛋白表达无影响(P0.05)。体外q PCR结果显示E2处理组和E2P4联合处理组细胞中都未检测到mi R-1a-3p表达,体外Ed U掺入实验表明外源性mi R-1a-3p拟似剂能够一定程度抑制雌激素促细胞增殖作用(P0.05)。以上结果表明,孕激素可通过诱导mi R-1a-3p表达引起细胞周期G1/S期阻滞,从而抑制EECs细胞增殖,这种抑制增殖作用与其抑制cyclin E1和cyclin E2蛋白表达有关。     

MiR-216a-3p regulates the proliferation,apoptosis, migration,and invasion of lung cancer cells via targeting COPB2

ABSTRACT

Effect of miR-216a-3p on lung cancer hasn’t been investigated. Here, we explored its effects on lung cancer. MiR-216a-3p expression in lung cancer tissues and cells was detected by RT-qPCR. The target gene of miR-216a-3p was predicted by bioinformatics and confirmed by luciferase-reporter assay. After transfection, cell viability, migration, invasion, proliferation, and apoptosis were detected by MTT, scratch, transwell, colony formation, and flow cytometry. The expressions of COPB2 and apoptosis-related factors were detected by RT-qPCR or western blot. MiR-216a-3p was low-expressed and COPB2 was high-expressed in lung cancer tissues and cells. MiR-216a-3p targeted COPB2 and regulated its expression. MiR-216a-3p inhibited lung cancer cell viability, migration, invasion, and proliferation, while promoted apoptosis. Effect of miR-216a-3p on lung cancer was reversed by COPB2. MiR-216a-3p regulated proliferation, apoptosis, migration, and invasion of lung cancer cells via targeting COPB2.     

miR-125a-3p调控人结肠癌细胞浸润转移的作用及机制研究

目的:探讨miR-125a-3p在结肠癌细胞浸润与转移中的作用及其可能机制。方法:通过qRT-PCR方法检测miR-125a-3p在结肠癌细胞及组织样本中的表达;在结肠癌细胞过表达或沉默miR-125a-3p后,通过平板克隆实验、MTT实验、划痕实验、Transwell实验检测结肠癌细胞增殖、迁移及侵袭能力的变化;采用Western blot方法检测miR-125a-3p过表达后相关标志分子的表达水平变化情况。结果:miR-125a-3p在结肠癌细胞及组织呈现异常低表达;过表达miR-125a-3p抑制结肠癌细胞HCT116及SW480的增殖能力;过表达或沉默miR-125a-3p分别抑制或增强结肠癌细胞的迁移与侵袭能力;过表达miR-125a-3p在mRNA及蛋白水平均能够显著抑制Snail、N-cadherin及Vimentin的表达,而增加E-cadherin的表达。结论:miR-125a-3p参与调节结肠癌细胞浸润与转移,其机制可能是通过调控上皮间质转化途径介导的。     

HuR facilitates cancer stemness of lung cancer cells via regulating miR-873/CDK3 and miR-125a-3p/CDK3 axis

Objectives

To study the roles and mechanisms of HuR in cancer stem cell maintenance of lung cancer.

Results

HuR expression was increased in tumor spheres of lung cancer cells. Knockdown of HuR suppressed spheroid formation and size, inhibited the expression of stemness-related marker, Oct4, Nanog and ALDH in lung cancer cells. Importantly, HuR and CDK3 expressions were increased in lung cancer tissues compared with normal adjacent tissues, and positively correlated. Mechanistically, HuR directly bound to CDK3, and increased CDK3 mRNA stability and expression. Additionally, miR-873 or miR-125a-3p attenuated the promotion of HuR on CDK3 expression and lung cancer stemness. Furthermore, HuR facilitated lung cancer stemness dependent on CDK3 expression. miR-873 or miR-125a-3p level was negatively correlated with HuR and CDK3 expression levels in lung cancer tissues.

Conclusions

HuR facilitates lung cancer stemness via regulating miR-873/CDK3 and miR-125a-3p/CDK3 axis.

     

microRNA-199a-3p,DNMT3A,and aberrant DNA methylation in testicular cancer

It was previously demonstrated that miR-199a was downregulated in testicular germ cell tumor (TGCT), probably due to hypermethylation of its promoter. Further study found that re-expression of miR-199a in testicular cancer cells (NT2) led to suppression of cell growth, cancer migration, invasion and metastasis. More detailed analyses showed that these properties of miR-199a could be assigned to miR-199a-5p, one of its two derivatives. The biological role of the other derivative, miR-199a-3p in TGCT, remains largely uncharacterized. In this report, we identified DNA (cytosine-5)-methyltransferase 3A (DNMT3A), the de novo methyltransferase, as a direct target of miR-199a-3p using a 3′-UTR reporter assay. Transient expression of miR-199a-3p in NT2 cells led to decrease, while knocking down of miR-199a-3p in a normal human testicular cell line (HT) led to elevation, of DNMT3A2 (DNMT3A gene isoform 2) mRNA and protein levels. In clinical samples, DNMT3A2 was significantly overexpressed in malignant testicular tumor, and the expression of DNMT3A2 was inversely correlated with the expression of miR-199a-3p. However, DNMT3A did not affect miR-199a expression in NT2 cells. Further characterization of miR-199a-3p revealed that it negatively regulated DNA methylation, partly through targeting DNMT3A. Overexpression of miR-199a-3p restored the expression of APC and MGMT tumor-suppressor genes in NT2 cells by affecting DNA methylation of their promoter regions. Our studies demonstrated the deregulation of miR-199a-3p expression in TGCT may provide novel mechanistic insights into TGCT carcinogenesis and suggested a potentially therapeutic use of synthetic miR-199a-3p oligonucleotides as effective hypomethylating compounds in the treatment of TGCT.     

间充质干细胞外泌体对肺脏上皮钠离子转运的调控作用研究

该文讨论了小鼠骨髓间充质干细胞来源的外泌体(bone mesenchymal stem cell-exo-some,BMSC-exo)对肺损伤引起的肺泡上皮钠离子转运障碍的调控.从BMSCs的条件培养基中分离外泌体,利用透射电镜技术对其形态结构以及大小进行了鉴定;对培养的经典肺上皮细胞系H441细胞分别给予脂多糖或外泌...     

miR-182在前列腺癌组织中的表达及其对前列腺癌细胞增殖和凋亡的影响

目的:观察前列腺癌组织及不同前列腺癌细胞系中miR-182的表达,并探讨下调其表达对前列腺癌细胞增殖和凋亡的影响及机制。方法:采用实时荧光定量PCR(q RT-PCR)检测30例前列腺癌组织和30例相应的癌旁组织以及前列腺正常上皮RWPE-1细胞、前列腺癌PC-3、LNCa P和DU145细胞中miR-182的表达,进一步采用Lipfectamine 2000脂质体转染miRNA-182 inhibitor和阴性对照miRNA于PC-3细胞后,通过噻唑蓝(MTT)比色法检测细胞增殖情况,流式细胞术检测细胞凋亡率,免疫印迹(Western blot)法检测转录因子FOXO1、血管内皮生长因子(VEGF)和抑癌基因p53蛋白的表达。结果:miR-182在前列腺癌组织中的表达明显高于癌旁组织(P0.05);miR-182在前列腺癌细胞系PC-3、LNCa P和DU145中的表达均高于前列腺正常上皮细胞RWPE-1(P0.05),其中PC-3细胞中miR-182表达水平最高。转染miRNA-182 inhibitor至PC-3细胞成功下调miR-182表达后,细胞的增殖能力明显受到抑制,细胞凋亡能力明显增强,FOXO1表达水平显著升高,VEGF和p53的表达明显降低,差异均具有统计学意义(P0.05)。结论:miR-182在前列腺癌组织及细胞中呈高表达,下调miR-182的表达可能通过增加FOXO1的表达并减少VEGF和p53的表达,抑制前列腺癌细胞增殖并诱导细胞凋亡。     

Licochalcone A reverses NNK-induced ectopic miRNA expression to elicit in vitro and in vivo chemopreventive effects

BackgroundChemoprevention is the best cost-effective way regarding cancers. MicroRNAs (miRNAs) have been reported to be differentially expressed during the development of lung cancer. However, if lung cancer prevention can be achieved through modulating miRNAs expression so far remains unknown.PurposeTo discover ectopically expressed miRNAs in NNK-induced lung cancer and clarify whether Licochalcone A (lico A) can prevent NNK-induced lung cancer by modulating miRNA expression.Study design and methodsA/J mice were used to construct a lung cancer model by intraperitoneal injection with physiological saline NNK (100 mg/kg). Chemopreventive effects of lico A against lung cancer at 2 mg/kg and 20 mg/kg doses were evaluated in vivo. MicroRNA array and RT-qPCR were used to assess the expression levels of miRNAs. MLE-12 cells were treated with 0.1 mg/ml NNK, stimulating the ectopic expression pattern of miR-144-3p, miR-20a-5p, miR-29c-3p, let-7d-3p, and miR-328-3p. miR-144-3p mimics and inhibitors were used to manipulate miR-144-3p levels. The effects of lico A (10 μM) on cell cycle distribution, apoptosis, and the expression of CK19, RASA1, miR-144-3p, miR-20a-5p, miR-29c-3p, let-7d-3p, and miR-328-3p in NNK-treated MLE-12 cells were studied.ResultsThe expression levels of miR-144-3p, miR-20a-5p, and miR-29c-3p increased, while those of let-7d-3p and miR-328-3p decreased in both NNK-induced A/J mice and MLE-12 cells. Lico A could reverse the NNK-induced ectopic miRNA (miR-144-3p, miR-20a-5p, miR-29c-3p, let-7d-3p, and miR-328-3p) expression both in vivo and in vitro and elicit in vivo lung cancer chemopreventive effect against NNK. In MLE-12 cells, the overexpression of miR-144-3p elicited the same effect as NNK regarding the expression of lung cancer biomarker CK19; the silencing of miR-144-3p reversed the effect of NNK on cell cycle distribution and apoptosis. Lico A could reverse the effect of NNK on the expression of miR-144-3p, CK19, and RASA1 (predicted target of miR-144-3p).ConclusionThe present study suggests that miR-144-3p, miR-20a-5p, miR-29c-3p, let-7d-3p, and miR-328-3p were involved in the in vivo pathogenesis of NNK-induced lung cancer, and lico A could reverse the effect of NNK both in vivo and in vitro to elicit lung cancer chemopreventive effects through, at least partially, these five ectopically expressed miRNAs, especially miR-144-3p.     

circ_0001461靶向抑制miR-30a-5p对骨肉瘤细胞增殖和凋亡的影响

摘要 目的：探讨circ_0001461对骨肉瘤细胞增殖和凋亡的影响及调控机制。方法：采用实时荧光定量聚合酶反应（qRT-PCR）检测检测circ_0001461在骨肉瘤组织和细胞中的表达水平。在U2OS和HOS细胞中转染sh-NC和sh-circ_0001461后,采用CCK8检测细胞增殖情况,流式细胞术检测细胞凋亡情况,qRT-PCR检测增殖相关分子Ki-67 mRNA的表达水平,Western Blot检测凋亡相关分子Cleaved-caspase-3蛋白的表达水平。采用双荧光素酶报告基因检测circ_0001461和miR-30a-5p的结合情况。结果：circ_0001461在骨肉瘤组织中的表达水平明显高于癌旁正常组织（P<0.05）,circ_0001461在骨肉瘤细胞U2OS和HOS中的表达水平均明显高于成骨细胞NHOst（P<0.05）。低表达circ_0001461能够抑制骨肉瘤细胞U2OS和HOS的增殖和增殖相关分子Ki-67的表达（P<0.05）;促进骨肉瘤细胞U2OS和HOS的凋亡和凋亡相关分子Cleaved-caspase-3蛋白的表达（P<0.05）。双荧光素酶结果显示circ_0001461能够靶向结合miR-30a-5p。低表达circ_0001461能够促进miR-30a-5p的表达（P<0.05）,circ_0001461和miR-30a-5p在骨肉瘤组织中的表达呈负相关（P<0.05）。在U2OS细胞中共转染sh-circ_0001461和miR-30a-5p mimics后能够进一步加强单独转染sh-circ_0001461对U2OS细胞增殖和凋亡的影响（P<0.05）;在HOS细胞中共转染sh-circ_0001461和miR-30a-5p inhibitors后能够逆转单独转染sh-circ_0001461对U2OS细胞增殖和凋亡的影响（P>0.05）。结论：circ_0001461在骨肉瘤组织和细胞中明显高表达,低表达circ_0001461能够靶向促进miR-30a-5p的表达进而抑制骨肉瘤细胞增殖和促进细胞凋亡。     

环状RNA circMRPS35通过miR-130a-3p/ZNRF3轴调节胃癌细胞的增殖、凋亡、迁移和侵袭

摘要 目的：探讨环状RNA MRPS35（circMRPS35）对胃癌（GC）细胞增殖、凋亡、迁移和侵袭的调控机制。方法：体外培养人GC细胞系（HGC-27、MGC-803、MKN45和AGS）和正常胃上皮GES-1细胞,实时荧光定量PCR（RT-qPCR）检测circMRPS35、miR-130a-3p和锌环指蛋白3（ZNRF3）mRNA表达。另取MGC-803细胞,分为对照组、pc-NC组、pc-circMRPS35组、pc-circMRPS35+miR-NC组、pc-circMRPS35+miR-130a-3p组,采用Lipofectamine 3000进行质粒转染。RT-qPCR检测circMRPS35、miR-130a-3p和ZNRF3 mRNA表达,Western blot检测ZNRF3蛋白表达,CCK-8法、流式细胞术检测细胞增殖与凋亡,划痕实验和Transwell小室实验检测细胞迁移与侵袭能力,裸鼠移植瘤实验探究circMRPS35对GC细胞体内生长的影响。双荧光素酶报告基因检测miR-130a-3p与circMRPS35或ZNRF3的靶标关系。结果：GC细胞系中circMRPS35和ZNRF3 mRNA呈低表达,miR-130a-3p呈高表达（均P＜0.05）。过表达circMRPS35可降低miR-130a-3p,上调ZNRF3 mRNA和蛋白水平,抑制细胞增殖、迁移和侵袭,并促进细胞凋亡（均P＜0.05）;circMRPS35过表达对GC细胞恶性行为和裸鼠移植瘤生长的抑制作用可被miR-130a-3p mimic逆转（P＜0.05）。双荧光素酶实验结果显示,过表达miR-130a-3p可降低circMRPS35-WT和ZNRF3-WT的荧光素酶活性（P＜0.05）。结论：circMRPS35可能通过miR-130a-3p/ZNRF3轴抑制GC细胞的增殖、迁移和侵袭,并促进细胞凋亡。