miRNA-221 promotes proliferation,migration and invasion by targeting TIMP2 in renal cell carcinoma

Introduction: MicroRNAs (miRNAs) play important roles in tumorigenesis. In this study, we investigated the role of miR-221 in the development and progression of clear cell renal cell carcinoma (ccRCC). Methods: Quantitative real-time PCR (qRT-PCR) was used to measure the expression level of miR-221 in ccRCC tissues and cell lines. Then, we investigated the role of miR-221 to determine its potential roles on renal cancer cell proliferation, migration and invasion in vitro. A luciferase reporter assay was conducted to confirm the target gene of miR-221 and the results were validated in renal cancer cells. Results: In the present study, we found that miR-221 was significantly increased in ccRCC tissues and cell lines. Knocked-down expression of miR-221 remarkably inhibited cell proliferation, migration and invasion of renal cancer cells. Moreover, at the molecular level, our results suggested that TIMP2 as a direct target of miR-221 through which miR-221 promoted tumor cell proliferation, migration and invasion. Conclusions: These findings suggested that miR-221 play an oncogenic role in the renal cancer cell proliferation, migration and invasion by directly inhibiting the tumor suppressor TIMP2, indicating miR-221 act as a potential new therapeutic target for the treatment of ccRCC.     

miR-338-3p inhibits the proliferation and migration of gastric cancer cells by targeting ADAM17

Background: MicroRNAs (miRNA) have been documented playing a critical role in cancer progression. Although miR-338-3p has been implicated in several cancers, its role in gastric cancer is still unknown. The aim of our study was to investigate the role of miR-338-3p in gastric cancer progression. Methods: Expression levels of miR-338-3p in gastric cancer cell lines and tissues were determined by quantitative real-time PCR (qRT-PCR). The effect of miR-338-3p on proliferation was evaluated by MTT assay, cell migration and invasion were evaluated by transwell migration and invasion assays. Furthermore, luciferase reporter assay was conducted to confirm the target gene of miR-338-3p, and the results were validated in gastric cancer cells. Results: In the present study, we found that miR-338-3p was down-regulated in both gastric cancer cell lines and tissues. Enforced expression of miR-338-3p inhibited proliferation, migration and invasion of gastric cancer cells in vitro. Moreover, we identified A disintegrin and metalloproteinase 17 (ADAM17) gene as potential target of miR-338-3p. Importantly, ADAM17 rescued the miR-338-3p mediated inhibition of cell proliferation, migration and invasion. Conclusions: Our study suggested that miR-338-3p is significantly decreased in gastric cancer, and inhibits cell proliferation, migration and invasion partially via the downregulation of ADAM17. Thus, miR-338-3p may represent a potential therapeutic target for gastric cancer intervention.     

miR-200b调控PDCD4的表达对乳腺癌细胞增殖和侵袭的影响

目的探讨miR-200b对乳腺癌细胞增殖及侵袭的影响及其可能的分子机制。方法利用荧光实时定量PCR检测人乳腺癌组织及乳腺癌细胞株中miR-200b的表达差异;利用生物信息学方法预测miR-200b的靶基因,并使用免疫蛋白印迹实验对靶基因的表达进行验证;分别将miR-200b siRNA、PDCD4 mimics以及相应对照miRNA转染MCF-7细胞,通过CCK-8实验检测细胞的增殖能力;采用Transwell侵袭实验检测细胞的侵袭能力。结果miR-200b在乳腺癌组织中呈低表达水平,进一步抑制miR-200b的表达,乳腺癌细胞的增殖及侵袭能力显著增强(P<0.05);将miR-200b siRNA、PDCD43′-UTR共转染到乳腺癌MCF-7细胞中,双荧光素酶报告基因结果显示抑制miR-200b的表达后,PDCD4的表达及活性相应上调(P<0.05);同时通过蛋白免疫印迹实验可以发现,抑制miR-200b表达后,PDCD4蛋白表达含量降低,乳腺癌细胞的增殖及侵袭能力明显增强(P<0.05);而过表达PDCD4后PDCD4蛋白表达含量增加(P<0.05),乳腺癌细胞的增殖及侵袭能力明显降低(P<0.05)。结论miR-200b可靶向上调PDCD4基因的表达,从而抑制乳腺癌细胞的增殖及侵袭能力。     

Down-regulation of miR-497 is associated with poor prognosis in renal cancer

Introduction: Clear cell renal cell carcinoma (ccRCC) is the most common type of cancer in the adult kidney, and the prognosis of metastatic ccRCC remains poor with high mortality. Recent study indicated that microRNAs (miRNAs) played critical roles in tumor progression. The aim of this study was to investigate the expression, biological role and clinical significance of miR-497 in ccRCC. Methods: Quantitative real-time PCR (qRT-PCR) was performed to detect the expression of miR-497 in renal cancer cell lines and ccRCC tissues. The association between miR-497 expression and overall survival was estimated by the Kaplan-Meier method. Gain of function assays were performed in the 786-O renal cancer cell line. Results: Expression of the miR-497 was significantly decreased in renal cancer cell lines and ccRCC tissues when compared with normal human proximal tubule epithelial cells and adjacent non-tumor tissues. Decreased miR-497 expression was significantly associated with tumor stage, histological grade and lymph node metastases. Significantly shorter overall survival was observed in patients with lower expression of the miR-497. Overexpression of miR-497 significantly inhibited renal cancer cell proliferation, migration and invasion. Conclusions: Our results demonstrated that miR-497 was decreased in ccRCC tissues and may provide a potential prognostic biomarker and a potential target for therapeutic intervention.     

MicroRNA-145 suppresses cell migration and invasion by targeting paxillin in human colorectal cancer cells

A number of cancers show increased expression of paxillin which plays a central role in tumor progression, including colorectal cancer. However, the mechanisms causing paxillin upregulation remains unclear. In our study, bioinformatics analyses suggested that paxillin is predicted to be a direct target of miR-145. We firstly identified paxillin as a new target of miR-145 and demonstrated that miR-145 inhibits paxillin expression by binding to the paxillin mRNA 3’UTR. Therefore, we assume overexpression of paxillin induced by suppression of miR-145 may promote cell migration and invasion. We detected the expression of paxillin and miR-145 in human colorectal cancer tissues by real-time quantitative PCR. Higher expression of paxillin and lower expression of miR-145 was observed in colorectal cancer tissues than corresponding paracancerous tissue. Moreover, the expression of paxillin was negatively correlated with miR-145 expression. A dual-luciferase reporter assay was used to confirm that paxillin was a direct target of miR-145. In CRC cell lines, overexpression of miR-145 could downregulate paxillin protein expression levels, and ectopic overexpression of miR-145 mimics or inhibitor could inhibit or promote cell migration, invasion, proliferation and clone formation in vitro. Taken together, these data suggested that miR-145 plays a pivotal role in colon cancer through inhibiting cell proliferation migration and invasion, and miR-145 may serve as a tumor suppressor by targeting paxillin gene.     

microRNA-182 inhibits the proliferation and invasion of human lung adenocarcinoma cells through its effect on human cortical actin-associated protein

Lung cancer is one of the main causes of cancer death worldwide. The cortactin gene, CTTN, may play a pivotal role in the proliferation and invasion of tumors. A microRNA (miR-182) was cloned and used to study the expression of CTTN and its regulatory effects on the proliferation and invasion of the lung cancer cell line, A549. Cortactin protein and CTTN mRNA expression decreased in A549 cells that were transfected with the miR-182 expression plasmid. A cell proliferation assay indicated that miR-182 expression affected cell cycle regulation and suppressed proliferation of lung cancer cells in vitro. In addition, xenograft experiments confirmed the suppression of tumor growth in vivo, which was due to the promotion of apoptosis. In conclusion, endogenous mature miR-182 expression may have an important role in the pathogenesis of lung cancer through its interference with the target gene CTTN by epigenetic modification.     

MicroRNA-940在乳腺癌中的表达变化及作用机制

目的 探讨microRNA-940(miR-940)在乳腺癌组织和细胞中的表达以及对乳腺癌细胞增殖、侵袭、迁移能力的影响及其相关分子机制。 方法 实时定量聚合酶链反应（Real-time PCR）检测2016年1月~2017年1月我院手术切除的78例患者的乳腺癌组织、癌旁组织和人乳腺癌细胞系MCF-7、SK-BR-3、MDA-MB-231、BT-549及人正常乳腺细胞系MCF-10 A中miR-940的表达情况。在乳腺癌细胞系MDA-MB-231中利用脂质体LipofectaminsTM 2000转染miR-940模拟物上调miR-940的表达,CCK-8实验检测细胞增殖活性的改变,小室侵袭及迁移实验（transwell）检测细胞侵袭、迁移能力的改变。生物学信息法预测miR-940的可能作用靶基因,双荧光素酶报告实验检测miR-940与CXC趋化因子受体2 (CXCR2)的3’UTR区结合情况,Western blotting检测miR-940对CXCR2 蛋白表达的影响。 结果 miR-940在乳腺癌组织和细胞中表达明显降低（P<0.01）,并且miR-940的表达与TNM分期和淋巴结转移密切相关（P<0.01）。上调MDA-MB-231细胞miR-940表达后,细胞的增殖活性明显下降（P<0.01）,侵袭及迁移能力明显下降（P<0.01）。双荧光素酶报告显示,miR-940可与CXCR2 的3’UTR区特定序列结合显著抑制荧光素酶活性(P<0.01),上调miR-940后细胞中CXCR2 蛋白的表达均明显下降(P<0.01)。 结论 miR-940在乳腺癌中的表达降低,miR-940可以通过靶向CXCR2抑制乳腺癌细胞的增殖、侵袭及迁移能力。     

miR-126 对结肠癌细胞生物学行为的影响及其在调控结肠癌EMT 转化中的作用

目的:观察miR-126 在不同转移潜能的人结肠癌细胞系中的表达情况及其对结肠癌细胞增殖、侵袭转移能力的影响并探讨可能的作用机制。方法:采用实时荧光定量PCR 检测结肠癌细胞系(SW480、SW620 及HCT116)中miR-126 的表达量。通过脂质体瞬时转染法将miR-126 过表达(miR-126 mimics),并设置阴性对照组,然后采用CCK8 法检测细胞的增殖能力,细胞划痕实验检测细胞的迁移能力,Transwell 侵袭小室实验检测细胞的侵袭能力,Western blot 实验检测E-cadherin 和Vimentin 蛋白表达量的变化。结果:相对于低转移潜能的结肠癌细胞株SW480,miR-126 在高转移潜能的SW620 和HCT116细胞中的表达降低。过表达miR-126 可使SW620 细胞增殖、迁移和侵袭能力降低,E-cadherin 蛋白表达增加,Vimentin 蛋白表达降低,差异具有统计学意义(P<0.05)。结论:低表达的miR-126 与结肠癌的转移密切相关,miR-126 影响结肠癌细胞生物学行为的作用可能是通过调控EMT 进程实现的。     

MicroRNA-17 promotes osteosarcoma cells proliferation and migration and inhibits apoptosis by regulating SASH1 expression

MicroRNAs (miRNAs) are abnormally expressed in numerous diseases, which are intimately associated with cell proliferation, migration and invasion. Recent study indicated that miR-17 may be involved in regulating osteosarcoma (OS) occurrence and development, but its function and mechanism have not been reported. In this study, quantitative real-time PCR (qRT-PCR) was used to measure the expression of miR-17, and Western blotting assay was performed to measure the expressions of SAM and SH3 domain containing 1 (SASH1), phosphoinoinositide-3 kinase (PI3K), protein kinase B (AKT), Caspase3, Bcl-2 gene family (Bcl-2, Bax) and matrix metalloprotein (MMP-2, MMP-9) in MG-63 cells. Luciferase reporter assay was conducted to confirm the target of SASH1 by miR-17. Cell proliferation, migration, invasion and apoptosis assay was performed to investigate the role of miR-17 in OS cells. We found that the expression of miR-17 was significantly up-regulated in OS cell lines. MiR-17 inhibitor inhibited the proliferation ability, and induced apoptosis of OS cells. Besides, miR-17 inhibitor prevented the migration and invasion of OS cells. Further, we identified that SASH1 was a target gene of miR-17. In addition, knockdown of miR-17 increased the protein expression of SASH1, and regulate related genes of cell proliferation, invasion and anti-apoptosis in the downstream of OS cells. These findings indicated that miR-17 was over-expressed and promoted cell proliferation, migration and inhibited cell apoptosis by targeting SASH1 in OS cells.     

微小RNA-138-5p抑制肺癌细胞增殖、迁移和侵袭能力的机制研究

目的:探讨微小RNA-138-5p(miR-138-5p)抑制肺癌细胞增殖、迁移和侵袭能力的相关机制。方法:以肺癌细胞A549和H460作为研究对象,分别转染miR-NC(对照组)或miR-138-5p(实验组);生物信息学技术预测miR-138-5p的靶基因;RT-qPCR检测转染后细胞miR-138-5p、叉头框蛋白C1(FOXC1)mRNA和波形蛋白(vimentin)mRNA的相对表达量;Western blot法检测FOXC1、vimentin、E-cadherin、N-cadherin和β-catenin蛋白表达变化;MTS法和集落形成实验分别检测细胞的增殖能力;划痕愈合实验和Transwell法检测细胞迁移和侵袭能力。结果:miR-138-5p过表达显著降低FOXC1和vimentin的mRNA及蛋白的表达(P0.05),E-cadherin和β-catenin蛋白表达上调,N-cadherin蛋白表达下调,显著抑制肺癌细胞的增殖、迁移和侵袭能力(P0.05)。结论:miR-138-5p可以通过靶向干扰FOXC1和vimentin的表达抑制肺癌细胞的增殖、迁移和侵袭,可能是肺癌基因治疗的潜在靶点。     

miR-454-3p靶向BPTF表达调控肺癌细胞A549增殖、迁移和侵袭

目的探讨miR-454-3p对肺癌细胞增殖、迁移和侵袭的影响及分子机制。方法采用RT-PCR技术检测miR-454-3p在肺癌组织以及肺癌细胞株中的表达,以表达量最低的肺癌细胞A549为后续分析对象,将miR-454-3p mimic转入A549细胞,RT-PCR验证miR-454-3p的过表达效率;采用CCK-8、迁移、侵袭等实验,观察转染组与对照组肺癌细胞的增殖、迁移和侵袭情况;生物信息学预测BPTF是miR-454-3p的靶标,构建BPTF 3′UTR荧光素酶载体,通过双荧光素酶报告基因验证miR-454-3p和BPTF的靶向关系;应用Western blot法检测BPTF的表达及迁移、侵袭相关蛋白的变化。结果RT-PCR实验表明miR-454-3p在肺癌组织和细胞中表达下调,且在A549细胞中表达最低(P<0.05)。与对照组相比,过表达miR-454-3p的肺癌细胞A549增殖能力明显降低,迁移和侵袭能力均受到抑制(P<0.05)。生物信息学软件分析BPTF为miR-454-3p的潜在靶基因,过表达miR-454-3p后BPTF的表达水平明显受到抑制。同时,迁移相关蛋白MMP-2和MMP-9表达明显下调,E-cadhern表达明显上调,而E-cadherin的负性调控N-cadherin表达下降。结论miR-454-3p可能通过靶向下调BPTF的表达,抑制肺癌细胞的增殖、迁移和侵袭能力,为肺癌的靶向治疗提供潜在靶标。     

MicroRNA-133b inhibits proliferation and invasion of ovarian cancer cells through Akt and Erk1/2 inactivation by targeting epidermal growth factor receptor

Aberrant expression of microRNA-133b (miR-133b) has been frequently reported in some cancers excluding ovarian cancer (OC). The role and its molecular mechanism of miR-133b in OC have not been reported. In this study, we explored the effects of miR-133b overexpression on proliferation and invasion in OC cells. The mRNA level of miR-133b in OC cell lines was determined by real-time PCR. The miR-133b mimic was transiently transfected into OC cells using Lipofectamine™ 2000 reagent. Subsequently, cell proliferation and invasion were assessed by MTT, Brdu-ELISA and Transwell assays. Moreover, the effects of miR-133b overexpression on the MAPK and PI3K/Akt signaling pathways were determined by Western blot. Protein level of EGFR was also measured by Western blotting. Meanwhile, luciferase assays were performed to validate EGFR as miR-133b target in OC cells. Our results showed that the mRNA level of miR-133b was remarkably decreased in OC cell lines compared with normal colon epithelium cells, whereas the protein expression of EGFR was significantly increased. Up-regulation of miR-133b inhibited the proliferation and invasion of OC cells. We also found that miR-133b overexpression evidently decreased the phosphorylation of Erk1/2 and Akt. Bioinformatics analysis predicted that the EGFR was a potential target gene of miR-133b. Luciferase reporter assay demonstrated that miR-133b could directly target EGFR. Altogether, our results indicated that miR-133b overexpression was shown to inhibit proliferation and invasion of OC cells through suppression of the MAPK and PI3K/Akt signaling pathways by targeting EGFR.     

MiR-124-3p inhibits the migration and invasion of Gastric cancer by targeting ITGB3

BACKGROUNDGastric cancer is one of the major malignant tumors in the world. Integrins expressed in cancer cells can promote tumor progression and migration. MiRNAs can inhibit the expression of target genes by directly binding to their mRNAs and can affect various important biological processes. The aim of this study was to investigate the role of miR-124- 3p and ITGB3 in gastric cancer.METHODSRT-PCR and western blot are used to detect the expression of miR-124-3p, ITGB3 and integrin β3 in gastric cancer tissues and cells. The wound healing, CCK-8 assay, transwell migration and invasion assay were performed to determine the cell proliferation, migration and invasion. What’s more, bioinformatics prediction and luciferase assay was conducted to demonstrated the binding efficiency between miR-124-3p and ITGB3.RESULTSWe verified that ITGB3 and miR-124-3p changes the migration and invasion of gastric cancer cells in vitro. The overexpression or silencing of miR-124-3p inhibited or promoted the proliferation, migration and invasion of both selected gastric cancer cells, and ITGB3 is just the reverse. Meanwhile, we validated that ITGB3 is the target of miR-124-3p by bioinformatics prediction and luciferase assay. Lastly, the expression of ITGB3 in 40 pairs of gastric cancer tissues were significantly higher than that in the adjacent normal tissues, while the expression level of miR-124-3p was significantly decreased in cancer tissues.CONCLUSIONSmiR-124-3p inhibits the migration and invasion of Gastric cancer by targeting ITGB3 in gastric cancer cells. Our results suggested that miR-124-3p and ITGB3 may reasonably serve as a promising therapeutic target.     

MiR-873-5p inhibits cell migration,invasion and epithelial-mesenchymal transition in colorectal cancer via targeting ZEB1

Recent studies have demonstrated that dysregulation of mircoRNAs (miRNAs) greatly affected biological processes of human cancers, including colorectal cancer. As a member of miRNAs family, miR-873-5p has been proved to be a tumor suppressor in some human cancers. Here, we aim to investigate the effects of miR-873-5p on the migration, invasion and epithelial-mesenchymal transition (EMT) of colorectal cancer cells. The low expression of miR-873-5p in colorectal cancer cells was identified by conducting qRT-PCR analysis. Gain of function assays were designed and conducted to demonstrate the specific function of miR-873-5p overexpression in colorectal cancer progression. Transwell assay and western blot assay were conducted and revealed that miR-873-5p inhibited cell migration, invasion and EMT formation. To find the downstream molecular mechanism of miR-873-5p, mechanism assays were designed and performed to find the downstream target of miR-873-5p. ZEB1 (Zinc finger E-box-binding homeobox 1) was certified to be the target of miR-873-5p through bioinformatics analysis, luciferase activity assay and pull-down assay. Finally, rescue assays were carried out to demonstrate the effects of miR-873-5p-ZEB1 axis on the migration, invasion and EMT process of colorectal cancer cells. In conclusion, we confirmed that miR-873-5p suppressed cell migration, invasion and EMT in colorectal cancer via targeting ZEB1.     

miR-130b-3p有望作为人膀胱癌的标志物

目的进一步了解miRNA在膀胱癌中的潜在机制。方法芯片分析4对人膀胱癌组织和相邻正常组织中的miRNA的表达。并用RT-q PCR来验证两个最上调的miRNA及其靶基因的表达是否符合miRNA/mRNA芯片结果。通过相关性分析和双荧光素酶报告实验推断并验证miR-130b-3p可以靶向PTEN。应用CCK8、EDU、流式细胞术、划痕、Transwell和细胞骨架等实验证明miR-130b可以影响膀胱癌细胞的增殖、凋亡、迁移和侵袭。用Western blot检测PI3K/AKT和整合素β1/FAK信号通路的关键靶蛋白。结果人膀胱癌中miR-130b-3p表达高于癌旁且与PTEN表达呈负相关。miR-130b-3p可下调PTEN表达,导致PI3K/AKT和整合素β1/FAK信号通路的激活,且与膀胱癌EJ细胞的增殖、迁移和侵袭相关。细胞转染miR-130b-3p抑制剂时,可以重排细胞骨架。结论本结果揭示miR-130b/PTEN有望用于人膀胱癌诊断和治疗的标志物。     

透明细胞肾细胞癌的表观遗传学研究进展

正肾脏肿瘤的发病率在人类泌尿系统肿瘤中排名第3,约占恶性肿瘤的3%,发病年龄主要在50~70岁,每年导致90 000多例患者死亡,且呈递增趋势~([1])。肾癌的具体发病机制不详,除遗传因素外,吸烟、肥胖、污染和辐射等也是重要因素。大多数透明细胞肾细胞癌(clear cell renal cell carcinoma,ccRCC)患者在早期无任何症状,20%~30%患者在     

miR-197-3p调控DMBT1抑制甲状腺癌细胞系增殖、迁移和侵袭

目的探讨miR-197-3p是否通过靶向调控恶性脑瘤缺失1基因(DMBT1)影响甲状腺癌细胞增殖、迁移和侵袭。方法RT-qPCR检测健康人甲状腺细胞Nthy-ori 3-1和甲状腺癌细胞SW579、CGTHW-1中miR-197-3p表达;MTT法检测SW579细胞增殖;Transwell小室法检测SW579细胞迁移和侵袭;双荧光素酶报告基因实验验证miR-197-3p是否靶向DMBT1;Western blot检测细胞DMBT1、cyclinD1、p21、MMP-2和E-cadherin蛋白表达。结果与Nthy-ori 3-1细胞比较,SW579和CGTHW-1细胞中miR-197-3p相对表达量升高(P<0.05);抑制miR-197-3p表达后,SW579细胞的增殖、迁移和侵袭能力明显受到抑制,细胞中cyclinD1蛋白和MMP-2蛋白表达降低而p21蛋白和E-cadherin蛋白表达升高;SW579细胞中miR-197-3p靶向负调控DMBT1的表达;过表达DMBT1明显抑制SW579细胞增殖、迁移和侵袭,而抑制DMBT1则能逆转miR-197-3p对SW579细胞增殖、迁移和侵袭的影响。结论miR-197-3p通过靶向调控DMBT1的表达,抑制甲状腺癌细胞增殖、迁移和侵袭。     

Low expression of miR-99b promotes progression of clear cell renal cell carcinoma by up-regulating IGF1R/Akt/mTOR signaling

Objective: Dysfunctions of microRNAs have been implicated in the progression of clear cell renal cell carcinoma (ccRCC). Here, we investigated the roles of miR-99b and miR-99b* in ccRCC development. Methods: The expression levels of miR-99b and miR-99b* in tumor and tumor-adjacent tissues from ccRCC patients were quantified by quantitative Real-Time PCR (qRT-PCR). MicroRNA mimics and inhibitors were employed to evaluate the functions of miR-99b and miR-99b*. The effects of miR-99b on the proliferation and migration of ccRCC cells were analyzed by MTT and wound-healing assays, respectively. The effect of miR-99b on the expression of its target gene IGF1R and mTOR was determined by western blotting and qRT-PCR. Results: The abundances of miR-99b and miR-99b* were lower in ccRCC tissues than in the tumor-adjacent tissues from patients. Similarly, the expression of these two microRNAs was higher in the normal kidney HK-2 cells than in the ccRCC cell lines. Moreover, miR-99b and miR-99b* inhibited the proliferation and migration of ccRCC cells. MiR-99b was found to down-regulate IGF1R and mTOR expression, likely through targeting their mRNAs to induce degradation. Consistently, the mRNA levels of IGF1R and mTOR were higher in ccRCC tissues than in the tumor-adjacent tissues, and Akt, a downstream factor of IGF1R, was highly activated correspondingly in ccRCC tissues. Conclusion: The low expression of miR-99b and miR-99b* contributes to ccRCC development and miR-99b acts as an onco-suppressor by suppressing IGF1R and mTOR expression to down-regulate IGF1R/AKT/mTOR signaling.