转染miRNAs降低脂多糖诱导的大鼠肠微血管内皮细胞水通道蛋白1表达

目的探讨大鼠肠微血管内皮细胞miRNAs对脂多糖(LPS)诱导的水通道蛋白1(AQP1)表达的影响。方法用机械吹打法及胰蛋白酶消化法从大鼠空肠分离微血管内皮细胞,用免疫荧光染色鉴定细胞;用荧光实时定量PCR(RT-qPCR)检测AQP1和miRNAs的表达; miRNAs芯片分析结合在线软件筛选调控AQP1表达的miRNAs,瞬时转染miRNAs模拟体及对照序列进行进一步验证。结果肠微血管内皮细胞v WF免疫荧光染色为阳性。用LPS(0、0. 1、1和10 mg/L)处理细胞后,AQP1 m RNA表达水平随LPS浓度增加呈下降趋势,其中1和10 mg/L LPS处理细胞后AQP1 m RNA表达水平与对照组相比有明显差异(P0. 05和P0. 001);与对照组相比,LPS处理组有22个表达上调2倍以上的miRNAs及9个下调2倍以上的miRNAs;软件预测显示芯片结果中的miR-423-5p、miR-874-5p、miR-361-3p、miR-219a-5p、miR-27b-3p、miR-133b和miR-666可与AQP1启动子区互补配对; miR-874-5p、miR-361-3p、miR-133b和miR-666在LPS处理后的肠微血管内皮细胞中表达上调;转染miR-133b和miR-666模拟体后可使AQP1 m RNA表达水平下调。结论 LPS可通过上调miR-133b和miR-666表达水平,间接抑制AQP1的表达,继而影响肠黏膜通透性。     

急性心肌梗死大鼠缺血心肌中差异microRNA的表达谱分析

 目的：筛选急性心肌梗死（AMI）大鼠缺血心肌中差异表达的microRNAs（miRNAs）,预测其靶基因并分析其可能的生物学功能。方法：结扎冠状动脉左前降支建立雄性Wistar大鼠AMI模型,同时检测其心电图和颈总动脉血压变化,并用TTC法测定心肌梗死面积;假手术（sham）组除不结扎冠状动脉左前降支外,其它实验程序与AMI组相同。心肌缺血4 h后取梗死区心肌组织,提取总RNA进行microRNA芯片杂交检测,并用real-time PCR进行验证;生物信息学方法预测差异miRNAs的靶点并分析其生物学功能。结果：心电图、血压检测及病理切片证实AMI模型制备成功。Microarray检测结果表明,与sham组相比,获得11个与急性心肌梗死相关的miRNAs,其中6个miRNAs上调表达,5个miRNAs下调表达;已知3个miRNAs（rno-miR-181c、rno-miR-146b和rno-miR-208）参与了心血管功能的调节,8个miRNAs（rno-miR-672*、rno-miR-743b、rno-miR-128、rno-miR-138-1*、rno-miR-336、rno-miR-138-2*、rno-miR-325-3p和rno-miR-3572）是否与心血管功能有关尚不清楚,可能是心肌梗死相关的新的生物标志物。预测的miRNA靶基因中的一部分亦与心血管功能相关。结论：本研究获得的与AMI相关的差异miRNAs,可能是急性心肌梗死新的生物标志物和潜在的治疗靶点。     

结肠癌化疗耐药相关miRNA表达谱的初步研究

目的:应用基因芯片技术筛选结肠癌耐药相关微小RNAs (miRNAs),探究miRNAs对化疗耐药的调控机制。方法:采用基因芯片技术分析结肠癌细胞系HCT8及其耐长春新碱细胞系HCT8/v中miRNAs的表达差异,对部分差异表达的miRNAs应用RT-qPCR进行验证,对表达差异显著的miRNAs进行靶基因预测,利用Gene Ontology (GO)和京都基因与基因组百科全书(KEGG)数据库对预测到的靶基因进行生物信息学分析。结果:筛选出342个差异表达miRNAs,其中190个表达上调,152个表达下调。RT-qPCR验证结果示miR-125-5p、miR-181c-5p和miR-153-3的表达情况和芯片检测结果一致;miR-130a-3p和miR-149-3p的表达与芯片检测结果不一致。GO分析结果显示,耐药相关基因主要富集的旁路是RNA聚合酶II调控区序列特异性DNA结合旁路,主要通过正向调节发挥作用,位置主要是在细胞内有界细胞器上。KEGG分析结果显示,耐药相关基因最为富集的是轴突导向通路、胰岛素信号通路及磷脂酶D信号通路。结论:miRNAs与结肠癌化疗耐药密切相关。对这些miRNAs的研究能使我们对结肠癌的化疗耐药机制有更深入的理解,并为逆转化疗耐药提供新的思路。     

人脐带间充质干细胞移植肝硬化大鼠肝脏miRNA的差异表达

背景：研究表明脐带间充质干细胞可以显著改善肝硬化的程度,进而修复肝损伤,但其治疗肝硬化的分子调控机制,尤其是非编码RNA调控的肝内基因变化,目前并没有得到详细的阐释。 目的：分析人脐带间充质干细胞移植肝硬化大鼠肝细胞中微小RNA基因表达的变化。 方法：采用四氯化碳皮下注射联合乙醇喂服方法建立肝硬化大鼠模型,造模8周后经尾静脉输注人脐带间充质干细胞,每周1次,连续注射4周。最后一次注射治疗1周后收集大鼠肝脏组织进行石蜡切片和提取肝脏RNA进行表达谱基因芯片分析,同时收集血清利用自动生化分析仪测定肝功能指标变化。 结果与结论：人脐带间充质干细胞治疗可以显著降低谷丙转氨酶、谷草转氨酶和转肽酶水平,脂肪病变和肝细胞坏死显著减少。微小RNA表达谱芯片杂交分析和PCR验证结果显示rno-miR-369-5p、rno-miR-3584-5p和rno-miR-153*这3种微小RNA基因在造模过程中先下调表达,并在人脐带间充质干细胞治疗后显著上调;而rno-miR-93、rno-miR-199a-3p、rno-miR-195、rno-let-7a和rno-miR-19a这5种微小RNA基因在造模过程中先上调表达,并在人脐带间充质干细胞治疗后显著下调。以上结果表明人脐带间充质干细胞逆转肝硬化和肝细胞损伤过程中,可能通过上调rno-miR-369-5p、rno-miR-3584-5p和rno-miR-153*等miRNA基因表达,下调rno-miR-93、rno-miR-199a-3p、rno-miR-195、rno-let-7a和rno-miR-19a等相关miRNA基因表达发挥治疗作用。 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

rno-miR-161靶向EGLN2抑制血管性痴呆大鼠额叶铁死亡

目的 探讨rno-miR-161调控egl-9家族缺氧诱导因子2(EGLN2)对血管性痴呆（vascular dementia,VD）大鼠额叶铁死亡的影响。方法 24只雄性SD大鼠按随机原则分为模型组和假手术组,模型组结扎双侧颈总动脉,假手术组不结扎双侧颈总动脉。术后4周水迷宫检测行为学。RT-qPCR检测VD模型大鼠额叶rno-miR-161的变化,RT-qPCR和Western blotting检测EGLN2和谷胱甘肽过氧化物酶4(GPX4)的表达。生物信息学预测及双荧光素酶报告基因验证rno-miR-161与EGLN2是否存在靶向调控关系。RT-qPCR及Western blotting检测rno-miR-161-mimic和rno-miR-161-inhibitor对EGLN2表达的影响。siEGLN2转染PC12细胞,检测GPX4的表达。结果 与假手术组相比,模型组大鼠额叶rno-miR-161表达下降、EGLN2表达上升、GPX4表达下降。rno-miR-161直接靶向EGLN2。rno-miR-161-mimic抑制EGLN2的表达,rno-miR-161inhibito...     

严重烫伤大鼠早期胫骨前肌组织中微小RNA表达谱数据的靶基因分析

目的初步确定严重烫伤大鼠早期胫骨前肌组织中差异表达微小RNA(miRNA)的靶基因,从而为进一步机制研究指明方向。 方法采用高通量芯片技术检测严重烫伤大鼠早期胫骨前肌miRNA表达谱数据,筛选部分差异明显且相关性较好的miNRA,包括miR-1-3p、miR-206-3p、miR-133a-3p、miR-22-3p、miR-30a-5p、miR-30d-3p、miR-190b-5p、miR-628和miR-678,利用互联网络检索数据库Mirbase、Miranda和Mirdb,对这些差异miRNA进行靶基因预测。 结果3大数据库预测结果显示差异miRNA主要调控79个靶基因,网络关系图显示了miR-1-3p、miR-206-3p、miR-133a-3p、miR-22-3p、miR-30a-5p、miR-30d-3p、miR-190b-5p、miR-628和miR-678等miRNA及其预测靶基因之间复杂的调控网络关系。 结论根据初步的预测结果,miR-1-3p和miR-206-3p共同调控的靶基因较多,可作为进一步机制研究的理论依据。     

慢性神经病理性疼痛对大鼠脊髓背角miRNA表达的影响

目的 筛选慢性神经病理性疼痛大鼠脊髓背角差异表达的miRNA,并预测其调控的靶基因.方法 建立大鼠坐骨神经慢性压迫损伤CCI模型,在术后疼痛高峰期取腰膨大脊髓背角,用miRNA芯片筛选CCI大鼠差异表达的miRNAs,再用荧光实时定量RT-PCR验证差异表达的miRNAs,并利用MIRANDA、TARGETSCAN、PICTAR 3个数据库找出这些miRNA可能调控的靶基因.结果 CCI大鼠表达上调的有miR-99b,表达下调的有miR-674-3p、miR-879与miR-325-5p.RT-qPCR验证结果与芯片基本相符.预测这些miRNA可能的靶基因约26个,这些基因功能广泛.结论 慢性神经病理性疼痛可导致miRNA的表达发生变化,这些miRNA及其调控的靶基因为进一步研究奠定了基础.     

微小RNAs对肺癌细胞系侵袭、迁移及凋亡的影响

目的 探讨微小RNAs（miRNAs）对肺癌细胞系侵袭、迁移及凋亡的影响。方法 通过hsa-mir-933、hsa-mir-4700-3p、hsa-mir-3144-3p、hsa-mir-3972和hsa-mir-548a-5p感染目的细胞，流式细胞术检测5种miRNAs对肺癌细胞系的细胞凋亡的影响；Transwell分析5种miRNAs对肺癌细胞系的细胞侵袭效果；划线分析5种miRNAs对肺癌细胞系的细胞迁移的影响；Real-time PCR检测miRNA表达情况。结果 与感染空载体组和无感染的对照组相比，感染了hsa-mir-933、hsa-mir-4700-3p、hsa-mir-3144-3p、hsa-mir-3972和hsa-mir-548a-5p的细胞凋亡明显增加，其中NCI-H460和A549均以转染hsa-mir-4700-3p凋亡率最高；5种miRNAs对肺癌细胞的侵袭数目明显减小；5种miRNAs对肺癌细胞的细胞的迁移效果减弱； Real-time PCR检测hsa-mir-933、hsa-mir-4700-3p、hsa-mir-3144-3p、hsa-mir-3972和hsa-mir-548a-5p处理后相应miRNA的水平表达均相应增加。 结论 5种miRNAs能够促进肺癌细胞系的凋亡，降低肺癌细胞系的高侵袭性，抑制肺癌细胞系的迁移，增加对应miRNA的表达。     

miR-133a-3p靶向调控CD2AP基因减轻RSV感染的人支气管上皮细胞系16-HBE损伤

目的探讨miR-133a-3p对呼吸道合胞病毒(RSV)感染的支气管上皮细胞凋亡、炎性反应的影响及其对CD2相关蛋白(CD2AP)的调控作用。方法收集59例支气管哮喘儿童的支气管黏膜组织标本为AsP组,同时选取59例支气管扩张非哮喘儿童的支气管黏膜组织标本为NC组,用RT-qPCR法检测组织中miR-133a-3p、CD2AP的表达量;RSV感染支气管上皮细胞(16-HBE)建立细胞损伤模型(RSV组),分别将miR-NC、miR-133a-3p mimics、si-NC、si-CD2AP、pcDNA-NC、pcDNA-CD2AP、miR-133a-3p mimics与pcDNA-NC、miR-133a-3p mimics与pcDNA-CD2AP转染至感染RSV的16-HBE细胞;用RT-qPCR法与Western blot法分别检测细胞中miR-133a-3p、CD2AP的表达量;用ELISA法检测TNF-α、IL-6、IL-1α的水平;用流式细胞术检测细胞凋亡率;双荧光素酶报告实验检测miR-133a-3p与CD2AP的靶向关系;Western blot法检测cleaved-caspase-3蛋白表达量。结果与NC组比较,AsP组支气管黏膜组织中miR-133a-3p的表达水平降低(P0.05),CD2AP mRNA的表达水平升高(P0.05);RSV感染的16-HBE细胞中miR-133a-3p的表达水平降低(P0.05),CD2AP的表达水平及TNF-α、IL-6、IL-1α的水平升高(P0.05),凋亡率及cleaved-caspase-3蛋白水平升高(P0.05);RSV感染的16-HBE细胞中转染miR-133a-3p mimics或转染si-CD2AP可明显降低TNF-α、IL-6、IL-1α的水平、凋亡率及cleaved-caspase-3蛋白水平(P0.05);双荧光素酶报告实验证实CD2AP是miR-133a-3p的靶基因;共转染miR-133a-3p mimics与pcDNA-CD2AP可明显逆转miR-133a-3p mimics对16-HBE细胞凋亡及炎性反应。结论 miR-133a-3p过表达可通过负向调控CD2AP的表达而抑制RSV感染的支气管上皮细胞炎性反应及凋亡,从而减轻细胞损伤。     

头颈部鳞状细胞癌预后相关的miRNAs的生物信息学分析

目的 利用癌症基因组图谱（TCGA）数据库微小核糖核酸（miRNAs）表达谱数据分析头颈部鳞状细胞癌（HNSCC）与癌旁正常组织间差异表达的miRNAs,结合临床信息寻找与HNSCC预后相关的miRNAs。方法 从TCGA中下载miRNAs表达数据,包括39例HNSCC患者和39个肿瘤邻近正常组织样本筛选差异表达的miRNAs,应用481例HNSCC患者的miRNAs表达谱和临床信息来评估找到的差异表达miRNAs的预后作用。结果 共筛选出114个差异表达的miRNAs,包括60个上调和54个下调的miRNAs。Kaplan-Meier生存分析显示miR-4652-5p和miR-99a-3p与HNSCC患者预后相关,单因素和多因素Cox回归分析显示,miR-4652-5p和miR-99a-3p是HNSCC的重要预后因素。结论 miR-4652-5p和miR-99a-3p与HNSCC患者预后相关,但miR-4652-5p和miR-99a-3p在头颈鳞状细胞癌发生发展中的分子机制仍需更全面的基础和临床研究进行探讨。     

减体积肝移植模型大鼠肝脏组织miRNAs表达谱变化

BACKGROUND: Liver regeneration is the key factor influencing the prognosis of living donor liver transplantation. There has not been the research on special miRNA of liver regeneration after living donor liver transplantation. OBJECTIVE: To analyze the variation of miRNAs expression profile after rat reduced-size liver transplantation at certain time point, select and verify target miRNA which can provide targeting intervention strategies in liver regeneration after rat reduced-size liver transplantation and provide theoretical evidence for liver regeneration after living donor liver transplantation. METHODS: The reduced-size liver transplantation models were established. miRNAs microarray was used to detect miRNA expression. In differentially expressed microRNAs, real-time quantitative PCR was utilized to detect target miRNAs. The credibility of miRNAs microarray results was verified. RESULTS AND CONCLUSION: Compared with rat liver tissue in the sham operation group, 11 miRNAs up-regulated in reduced-size liver transplantation, including let-7b-5p, let-7c-5p, miR-101a-3p, miR-103-3p, miR-130a-3p, miR-142-5p, miR-186-5p, miR-199a-3p, miR-21-5p, 221-3p and miR-34a-5p. Four miRNAs were down-regulated, including miR-26b-5p, miR-150-5p, miR-19a-3p and rno-miR-146-5p. PCR test further verified that miR-221-3p and miR-199a-3p expression changes approximated the chip results at 24, 48 hours and 1 week, indicating that results of miRNA microarray were believable. These results verified that it exists variation of miRNAs expression profile after rat reduced-size liver transplantation, which picked out and verified the target miRNAs.        

Global microRNA profiles and signaling pathways in the development of cardiac hypertrophy

Hypertrophy is a major predictor of progressive heart disease and has an adverse prognosis. MicroRNAs (miRNAs) that accumulate during the course of cardiac hypertrophy may participate in the process. However, the nature of any interaction between a hypertrophy-specific signaling pathway and aberrant expression of miRNAs remains unclear. In this study, Spague Dawley male rats were treated with transverse aortic constriction (TAC) surgery to mimic pathological hypertrophy. Hearts were isolated from TAC and sham operated rats (n=5 for each group at 5, 10, 15, and 20 days after surgery) for miRNA microarray assay. The miRNAs dysexpressed during hypertrophy were further analyzed using a combination of bioinformatics algorithms in order to predict possible targets. Increased expression of the target genes identified in diverse signaling pathways was also analyzed. Two sets of miRNAs were identified, showing different expression patterns during hypertrophy. Bioinformatics analysis suggested the miRNAs may regulate multiple hypertrophy-specific signaling pathways by targeting the member genes and the interaction of miRNA and mRNA might form a network that leads to cardiac hypertrophy. In addition, the multifold changes in several miRNAs suggested that upregulation of rno-miR-331*, rno-miR-3596b, rno-miR-3557-5p and downregulation of rno-miR-10a, miR-221, miR-190, miR-451 could be seen as biomarkers of prognosis in clinical therapy of heart failure. This study described, for the first time, a potential mechanism of cardiac hypertrophy involving multiple signaling pathways that control up- and downregulation of miRNAs. It represents a first step in the systematic discovery of miRNA function in cardiovascular hypertrophy.     

MiR-34a-5p and miR-452-5p: The Novel Regulators of Pancreatic Endocrine Dysfunction in Diabetic Zucker Rats?

Objective: The pancreatic endocrinal system dominates the regulation of blood glucose levels in vivo, and the dysfunction of pancreatic endocrine β-cells is a major cause of the occurrence and development of Type 2 diabetes (T2D). Although microRNA (miRNA) have been found to be key regulators of pancreatic β-cells proliferation, differentiation and apoptosis, the underlying mechanism remains enigmatic. The aim of this study was to identify several novel miRNAs which might be involved in the etiopathogenesis of diabetic β-cells dysfunction.Methods: The miRNA expression profiles in the pancreas of high-fat diet (HFD) fed Zucker diabetic fatty (ZDF) rats and Zucker lean (ZL) rats feed with normal-fat diet (NFD) were detected by using miRNA microarray chip, and individually verified the most significant factors by quantitative real-time polymerase chain reaction (qRT-PCR) assay. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were used to predict the target genes related to each of the identified miRNAs and the functions of these target genes in different metabolic signaling pathways.Results: Compared with the ZL rats, a total of 24 differentially expressed miRNAs were detected in ZDF rats. Among which miR-34a-5p and miR-452-5p were the most significantly up-regulated and down-regulated respectively. These miRNAs have not been reported in rats'' pancreas before. By GO and KEGG enrichment analyses, we found that miR-34a-5p could negatively regulate pancreatic β-cell proliferation through the involvement of Wnt signaling pathway. In addition, it was also found to regulate insulin secretion through the insulin signaling pathway to modulate blood glucose levels. At the same time, miR-452-5p was found to positively regulate the activity of the key rate-limiting enzyme branched-chain α-keto acid dehydrogenase-β (BCKDHB) in the catabolism of branched chain amino acids (BCAA), leading to mitochondrial dysfunction in pancreatic β-cells.Conclusions: miR-34a-5p and miR-452-5p were identified as the novel regulators of pancreatic endocrine dysfunction. These miRNAs might have the potential to be utilized as the new predictive biomarkers for the diagnosis of the occurrence and development of T2D, as well as the therapeutic targets for T2D treatment.     

Identification of microRNAs in Throat Swab as the Biomarkers for Diagnosis of Influenza

Background: Influenza is a serious worldwide disease that captures global attention in the past few years after outbreaks. The recent discoveries of microRNA (miRNA) and its unique expression profile in influenza patients have offered a new method for early influenza diagnosis. The aim of this study was to examine the utility of miRNAs for the diagnosis of influenza.Methods: Thirteen selected miRNAs were investigated with the hosts'' throat swabs (25 H1N1, 20 H3N2, 20 influenza B and 21 healthy controls) by real-time quantitative polymerase chain reaction (RT-qPCR) using U6 snRNA as endogenous control for normalization, and receiver operating characteristic (ROC) curve/Area under curve (AUC) for analysis.Results: miR-29a-3p, miR-30c-5p, miR-34c-3p and miR-181a-5p are useful biomarkers for influenza A detection; and miR-30c-5p, miR-34b-5p, miR-205-5p and miR-449b-5p for influenza B detection. Also, use of both miR-30c-5p and miR-34c-3p (AUC=0.879); and miR-30c-5p and miR-449b-5p (AUC=0.901) are better than using one miRNA to confirm influenza A and influenza B infection, respectively.Conclusions: Given its simplicity, non-invasiveness and specificity, we found that the throat swab-derived miRNAs miR-29a-3p, miR-30c-5p, miR-34b-5p, miR-34c-3p, miR-181a-5p, miR-205-5p and miR-449b-5p are a useful tool for influenza diagnosis on influenza A and B.     

Combined characterization of microRNA and mRNA profiles delineates early differentiation pathways of CD133+ and CD34+ hematopoietic stem and progenitor cells

MicroRNAs (miRNAs) have been shown to play an important role in hematopoiesis. To elucidate the role of miRNAs in the early steps of hematopoiesis, we directly compared donor-matched CD133(+) cells with the more differentiated CD34(+) CD133(-) and CD34(-) CD133(-) cells from bone marrow on the miRNA and mRNA level. Using quantitative whole genome miRNA microarray and sequencing-based profiling, we found that between 109 (CD133(+) ) and 216 (CD34(-) CD133(-) ) miRNAs were expressed. Quantification revealed that the 25 highest expressed miRNAs accounted for 73% of the total miRNA pool. miR-142-3p was the highest expressed miRNA with up to 2,000 copies per cell in CD34(+) CD133(-) cells. Eighteen miRNAs were significantly differentially expressed between CD133(+) and CD34(+) CD133(-) cells. We analyzed their biological role by examining the coexpression of miRNAs and its bioinformatically predicted mRNA targets and luciferase-based reporter assays. We provide the first evidence for a direct regulation of CD133 by miR-142-3p as well as tropomyosin 1 and frizzled homolog 5 by miR-29a. Overexpression of miRNAs in CD133(+) cells demonstrated that miR-142-3p has a negative influence on the overall colony-forming ability. In conclusion, the miRNAs expressed differentially between the CD133(+) and CD34(+) CD133(-) cells are involved in inhibition of differentiation, prevention of apoptosis, and cytoskeletal remodeling. These results are highly relevant for stem cell-based therapies with CD133(+) cells and delineate for the first time how the stem cell character of CD133(+) cells is defined by the expression of specific miRNAs.     

喉癌组织中的microRNA差异表达谱及miR-125a-5p抑制喉癌细胞增殖的初步研究

 目的：筛选并分析喉癌组织与周围正常喉黏膜的微小RNA(microRNAs,miRNAs) 之间的表达谱差异,为进一步研究miRNA与喉癌发生、发展的关系提供线索。方法：收集喉癌组织和癌旁正常喉黏膜标本共42对,随机选取10对标本进行miRNA微阵列基因芯片分析, 另选取32对标本进行实时荧光定量PCR （qRT-PCR）验证,获得喉癌组织中的miRNA差异表达谱。应用MTT法和克隆形成实验检测miR-125a-5p对喉癌Hep2细胞增殖的影响。结果：喉癌组织中的let-7f-5p、miR-10a-5p、miR-125a-5p、miR-144-3p、miR-195-5p、miR-203等6个miRNA在基因芯片以及qRT-PCR中表达均显著下调。与对照组相比,转染miR-125a-mimics组的喉癌Hep2细胞增殖能力受到抑制,而转染miR-125a-inhibitor组Hep2细胞增殖能力增强。结论：基因芯片与qRT-PCR结果一致;喉癌与正常喉黏膜之间存在明显的miRNA差异表达,这些miRNA的差异性表达可能与喉癌的发病、侵袭等相关。miR-125a可以抑制喉癌Hep2细胞的增殖,可能作为喉癌生物治疗的新靶点。     

蜂胶黄酮PB3A作用下结肠癌细胞株中miRNA-198和miRNA-296-5p的表达及功能预测

目的:探讨蜂胶黄酮短叶松素-3-乙酸酯(pinobanksin-3-acetate,PB3A)对结肠癌SW480细胞微小RNA(microRNA,miRNA)表达谱的影响,为结肠癌的治疗及靶向药物研发提供理论依据。方法:使用miRNA芯片技术分析检测蜂胶黄酮PB3A处理人类结肠癌SW480细胞后miRNA表达谱的变化。通过实时荧光定量PCR方法检测miRNA-198和miRNA-296-5p的表达水平,以此来验证miRNA芯片结果的准确性和可靠性。利用miRWalk、Micro T、miRanda等12个网上数据库预测这2条miRNAs的靶基因并进行靶基因功能富集分析。结果:miRNA芯片分析结果显示,蜂胶黄酮PB3A干预24 h后结肠癌SW480细胞中差异表达倍数在2倍及以上的miRNA有267条,其中差异表达倍数达10倍及以上的miRNA有30条,28条为上调表达,2条下调表达;RT-qPCR实验结果显示miRNA-198和miRNA-296-5p的表达量趋势跟miRNA芯片结果一致,表达差异有统计学意义(P0.05)。miRNA靶基因预测发现miRNA-198有859个靶基因,miRNA-296-5p有906个靶基因;对这些可能被调控的靶基因进行Gene Class分析,结果显示miRNA-198和miRNA-296-5p的靶基因功能主要为转录因子、拷贝数变异、细胞分化、癌基因、蛋白激酶、组蛋白、转移癌基因、肿瘤抑制基因等(P0.05)。信号通路富集分析结果显示,miRNA-198靶基因显著富集于肿瘤通路、Wnt信号通路、胞吞通路、Erb B信号通路、黏着斑通路、黑素生成通路等信号通路,而miRNA-296-5p靶基因在MAPK信号通路、胞吞通路、轴突导向通路、Wnt信号通路、胰岛素信号通路、钙离子信号通路等信号通路中出现聚集(P0.05)。结论:蜂胶黄酮PB3A影响结肠癌SW480细胞的miRNA表达谱。PB3A作用下miRNA-198和miRNA-296-5p的异常表达可能参与PB3A抗结肠癌的过程。