Circ_0021573 acts as a competing endogenous RNA to promote the malignant phenotypes of human ovarian cancer cells

Circular RNAs (circRNAs) have been reported to be implicated in the tumorigenesis and progression of ovarian cancer. Here, the study was designed to explore the activity of human circ_0021573 in ovarian cancer pathogenesis and its regulation through the competing endogenous RNA (ceRNA) crosstalk. Circ_0021573, microRNA (miR)? 936, and cullin 4B (CUL4B) were quantified by qRT-PCR and western blot. Cell proliferation ability was detected by XTT, 5-Ethynyl-2′-Deoxyuridine (EdU), and colony formation assays. Cell apoptosis, migration, and invasion were assessed by flow cytometry, wound-healing, and transwell assays, respectively. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were used to evaluate the direct relationship between miR-936 and circ_0021573 or CUL4B 3′UTR. Xenograft studies were applied to assess the role of circ_0021573 in tumor growth. Our data showed that circ_0021573 expression is enhanced in human ovarian cancer. Inhibition of circ_0021573 impedes cell proliferation, migration, and invasion and promotes apoptosis in vitro, as well as diminishes tumor growth in vivo. Mechanistically, circ_0021573 contains a miR-936 binding site, and miR-936 is a relevant mediator of circ_0021573 regulation. MiR-936 direct targets and inhibits CUL4B. MiR-936-mediated suppression of CUL4B hinders cell proliferation, migration, and invasion and accelerates apoptosis in vitro.. These data suggested that circ_0021573 might promote the malignant phenotypes of ovarian cancer cells by functioning as a ceRNA for miR-936 to induce CUL4B, which provided a promising target for the prevention and inhibition of ovarian cancer.     

外泌体miR-338对骨质疏松大鼠骨代谢水平、骨小梁微结构和骨生物力学的影响

摘要 目的：探讨外泌体miR-338对骨质疏松大鼠骨代谢水平、骨小梁微结构和骨生物力的影响。方法：采用健康成年SPF级SD雄性大鼠进行骨髓间充质干细胞（BMSCs）分离。采用双侧卵巢摘除手术方法构建了骨质疏松大鼠模型。采用qRT-PCR法检测miR-338的表达水平;检测大鼠的骨密度,骨小梁微结构和骨生物力学指标。结果：与空白对照组相比,OP模型组、OP+ ExoBMSCs、抑制组和过表达组miR-338的表达水平明显更高（P＜0.05）;抑制组的miR-338的表达水平低于OP模型组、OP+ExoBMSCs和过表达组（P＜0.05）;与空白对照组相比,OP模型组、OP+ ExoBMSCs、抑制组和过表达组OC、PINP、BALP的表达水平明显更低（P＜0.05）;抑制组的OC、PINP、BALP的表达水平明显高于OP模型组、OP+ ExoBMSCs和过表达组（P＜0.05）;与空白对照组相比,OP模型组、OP+ ExoBMSCs、抑制组和过表达组BV/TV、Th.N、Tb.Th、Conn.D水平更低,而Tb.Sp、SMI明显更高（P＜0.05）;抑制组组的BV/TV、Th.N、Tb.Th、Conn.D水平明显高于OP模型组、OP+ ExoBMSCs和过表达组,而Tb.Sp、SMI更低（P＜0.05）;与空白对照组相比,OP模型组、OP+ ExoBMSCs、抑制组和过表达组BMD、最大荷载、最大应力、最大位移、刚度水平更低（P＜0.05）;抑制组的BMD、最大荷载、最大应力、最大位移、刚度水平高于OP模型组、OP+ExoBMSCs和过表达组（P＜0.05）。结论：BMSCs源性的miR-338可影响骨质疏松大鼠骨代谢、骨小梁微结构和骨生物力学状态。     

miR-106a-5p调控 PTEN对鼻咽癌细胞SUNE2增殖和迁移的抑制作用研究

目的研究miR-106a-5p对鼻咽癌细胞SUNE2增殖和迁移的影响。 方法将体外培养的鼻咽癌细胞SUNE2分成对照组（细胞未做任何处理）、Anti-NC组（转染阴性对照抑制剂）、Anti-miR-106a-5p组（转染miR-106a-5p抑制剂）、后期实验另设Anti-miR-106a-5p-inhibitor+si-NC组（转染miR-106a-5p抑制剂、siRNA阴性对照）、Anti-miR-106a-5p-inhibitor+si-PTEN组（转染miR-106a-5p抑制剂、PTEN siRNA）,采用Realtime PCR测定miR-106a-5p表达,MTT法检测增殖,Transwell小室检测细胞侵袭和迁移能力变化,用Western blot方法测定vimentin、E-cadherin蛋白表达。在线靶基因预测软件预测miR-106a-5p的靶基因可能为PTEN,双荧光素酶报告系统鉴定miR-106a-5p和PTEN的靶向关系。两组间比较用独立样本t检验,多组间比较采用单因素方差分析,组间两两比较采用LSD-t检验。 结果与正常鼻咽上皮细胞NP69比较,鼻咽癌细胞CNE-2、HK1、SUNE2中miR-106a-5p水平（1.00±0.11比1.84±0.13、2.19±0.14、2.87±0.25）升高,差异具有统计学意义（P < 0.05）。与对照组、Anti-NC组比较,Anti-miR-106a-5p组鼻咽癌细胞miR-106a-5p水平（1.00±0.10、0.99±0.12比0.76±0.08）降低,OD值（0.56±0.05、0.57±0.04比0.32±0.02）,细胞侵袭数[（128.47±11.65）个、（129.84±10.93）个比（85.12±6.75）个],迁移数[（182.51±14.81）个、（180.68±17.64）个比（122.01±11.62）个],vimentin蛋白表达水平（0.84±0.09、0.82±0.07比0.30±0.05）降低,E-cadherin蛋白表达水平（0.29±0.04、0.28±0.05比0.76±0.08）升高,差异具有统计学意义（P均< 0.05）。与Anti-miR-106a-inhibitor+si-NC组比较,Anti-miR-106a-inhibitor+si-PTEN组细胞OD值（0.33±0.03比0.52±0.05）、侵袭数[（84.16±5.91）个比（105.79±8.63）个]、迁移数[（118.42±10.25）个比（164.28±12.05）个]、vimentin蛋白表达水平（0.34±0.06比0.68±0.05）均升高,E-cadherin蛋白表达水平（0.72±0.06比0.29±0.05）降低,差异具有统计学意义（P均< 0.05）。 结论miR-106a-5p可通过靶向调控PTEN抑制鼻咽癌细胞SUNE2增殖和迁移潜能。     

Ligustrazine inhibits the proliferation and migration of ovarian cancer cells via regulating miR-211

Ovarian cancer (OC) is a commonly diagnosed female cancer. Ligustrazine (LSZ), a natural compound, has been reported to exert anti-cancer activity, although the mechanisms underlying the anti-cancer effects are not clear. The present study investigated the impact of LSZ on cell proliferation and migration by regulating microRNA-211 (miR-211) expression using the human ovarian cancer SK-OV-3 and OVCAR-3 cell lines. OC cells were treated with 0, 0.5, 1, and 2 mM LSZ, and quantitative real-time PCR was utilized to measure miR-211 levels in SK-OV-3 and OVCAR-3 cells with different treatment. Moreover, to further confirm the roles of miR-211 in LSZ induced anti-tumor effects, miR-211 expression was inhibited by transfection of miR-211 inhibitors in SK-OV-3 cells. Cell proliferation of transfected cells was evaluated using the CCK-8 and colony formation assay. The scratch assay was employed to assess cell migration and transwell assay was performed for evaluating the cell invasion. Protein levels of epithelial–mesenchymal transition (EMT) markers were determined by Western blotting. We found that LSZ inhibited the viability, proliferation, migration and invasion ability of SK-OV-3 and OVCAR-3 cells in a dose-dependent manner; moreover, LSZ could significantly increase the expression of miR-211 in both SK-OV-3 and OVCAR-3, and knockdown of miR-211 in SK-OV-3 cells partially abrogated the anti-tumor behavior of LSZ by promoting the viability, proliferation, migration, invasion and EMT of SK-OV-3 cells. Thus, we found that LSZ can inhibit the proliferation and migration of OC cells via regulating miR-211. Our study suggests that LSZ might be a potential and effective treatment for OC.     

乏氧和辐射诱导的miR-1290 促进宫颈癌细胞EMT发生

目的:研究mi R-1290在宫颈癌Hela细胞上皮间质转化中的作用。方法:模拟肿瘤放射治疗的分割疗法,单次2 Gyγ射线连续照射宫颈癌Hela、Siha细胞诱导辐射抗性细胞株;采用micro RNA芯片技术比较辐射抗性细胞与亲本细胞中mi RNA的表达谱差异;经不同条件乏氧和辐射处理宫颈癌Hela细胞后检测mi R-1290的表达水平;借助mi R-1290及mi R-1290-inhibition慢病毒表达载体,调变mi R-1290在Hela细胞的表达;调变mi R-1290后,划痕实验及Transwell侵袭实验比较Hela细胞的侵袭和转移能力;蛋白质印迹法检测细胞中E钙黏素(E-cadherin)和N钙黏素(N-cadherin)的表达。结果:在宫颈癌辐射抗性细胞中mi R-1290表达水平显著升高(P0.05);乏氧和辐射可诱导mi R-1290在宫颈癌Hela细胞中表达(P0.05);上调表达mi R-1290,Hela细胞出现了明显的间质细胞的形态改变,细胞的侵袭和转移能力明显增强(P0.05)。在Hela细胞中上调表达mi R-1290,降低了细胞中E-cadherin的表达,同时升高了N-cadherin的表达(P0.05)。结论:乏氧和辐射可诱导mi R-1290在宫颈癌Hela细胞中表达,mi R-1290通过调控E-cadherin和N-cadherin的表达促进宫颈癌Hela细胞发生EMT。     

Diagnostic and prognostic value of miR-200 family in breast cancer: A meta-analysis and systematic review

BackgroundBreast cancer (BC) is the most common cancer for women all over the world. Great interests have been paid to discover accurate and noninvasive methods for breast cancer diagnosis and prognosis. Although the diagnostic and prognostic value of microRNA-200 (miRNA- 200, miR-200) family has been revealed in many studies, the results were inconsistent. Thus, this meta-analysis aims to assess the overall value of miRNA-200 family in breast cancer diagnosis and prognosis.MethodRelevant studies were searched from the following databases: PubMed, PMC, EMBASE, and ScienceDirect using key words: ("miRNA-200 family" or "miR-141" or "miR-200a" or "miR-200b" or "miR-200c" or "miR-429") and (“HER2” or “Luminal A” or “Luminal B” or “TNBC”) and ("breast cancers" or "breast carcinoma" or "breast malignancy" or "breast tumor"). The sensitivity, specificity, AUC were then calculated to estimate the diagnostic accuracy of the miR-200 family. As for the prognostic value of the miR-200 family, the pooled hazard ratio (HR) was assessed. Heterogeneity among individual studies was also examined by subgroup analyses.ResultA total of 24 articles were included in the meta-analysis. The diagnostic value of miR-200s in BC was presented by the pooled sensitivity was 0.86 (95% CI: 0.83-0.88); the pooled specificity was 0.82 (95% CI: 0.72-0.89); the pooled AUC was 0.931 (95% CI: 0.919-0.942). Besides, expression of miR-200s in metastatic breast cancer has sensitivity, specificity and AUC of 0.70 (95%CI: 0.56-0.81), 0.72 (95%CI: 0.61-0.81), and 0.814 (95%CI: 0.741-0.903), respectively. The meta-analysis then revealed that high expression of miR-200 family corresponded to poor OS (HR: 1.63, 95% CI: 1.03-2.52), poor DFS (HR: 1.55, 95% CI: 0.95-2.56) in BC patients while downregulation of miRNA-200s corresponded to poor OS (HR= 0.84, 95%CI: 0.46-1.63) in TNBC patients and poor OS (HR=0.49; 95%CI: 0.27-0.88) in luminal BC patient.ConclusionThe MiR-200 family has high diagnostic accuracy and can be used as an important biomarker to prognosticate breast cancer.     

Neuroprotective effects of microRNA-211-5p on chronic stress-induced neuronal apoptosis and depression-like behaviours

Findings from recent studies have revealed that microRNAs (miRNAs) are related to numerous neurological disorders. However, whether miRNAs regulate neuronal anomalies involved in the pathogenesis of depression remain unclear. In the present study, we screened miRNA expression profiles in the CA1 hippocampus of a rat model of depression and found that a specific miRNA, microRNA-211-5p, was significantly down-regulated in depressed rats. When miR-211-5p was up-regulated in these rats, neuronal apoptosis within the CA1 area was suppressed, effects which were accompanied with an amelioration of depression-like behaviours in these rats. These neuroprotective effects of miR-211-5p in depressed rats appear to result through suppression of the Dyrk1A/ASK1/JNK signalling pathway within the CA1 area. In further support of this proposal are the findings that knock-down of miR-211-5p within the CA1 area of normal rats activated the Dyrk1A/ASK1/JNK pathway, resulting in the promotion of neuronal apoptosis and display of depression-like behaviours in these rats. Taken together, these results demonstrate that deficits in miR-211-5p contribute to neuronal apoptosis and thus depression-like behaviours in rats. Therefore, the miR-211-5p/Dyrk1A pathway may be critically involved in the pathogenesis of depression and serve as a potential therapeutic target for the treatment of depression.     

MiR-92a regulates brown adipocytes differentiation,mitochondrial oxidative respiration,and heat generation by targeting SMAD7

Brown adipocytes are rich in mitochondria and linked to the body's blood fat levels and obesity. MiR-92a is negatively correlated with the activity of brown adipocytes. This study aimed to explore the mechanism of miR-92a on brown adipocytes. The expression of miR-92a in C2C12 cell was detected by a quantitative real-time-polymerase chain reaction (qRT-PCR). C2C12 cells were induced to brown adipocytes. The direct target gene of miR-92a was determined using the dual-luciferase reporter assay. Brown adipocytes were treated with isoprenaline (Iso) and transfected by miR-92a inhibitor and siSMAD7. The expression of heat-producing genes and adipose differentiation genes related to brown adipocytes were detected by qRT-PCR and Western blot analysis. The expression of SMAD7, p-SMAD2, and p-SMAD3 were detected using Western blot analysis. The mitochondrial content was measured by mitotracker fluorescent staining. MiR-92a inhibitor significantly decreased the expression of miR-92a in C2C12 cells. MiR-92a inhibitor could upregulate the expression of Ucp1, Cox7a1, Elovl3, Ppargc1α, PPARγ, and FABP4, and its effect on Ucp1 was increased after the treatment of isoprenaline. Moreover, miR-92a inhibitor increased mitochondrial content, oxygen consumption rate (OCR) and the expression of SMAD7 and suppressed the expressions of p-SMAD2 and p-SMAD3, whereas miR-92a directly targeted SMAD7 to exert its inhibitory effects. SiSMAD7 reversed the effects of the inhibitor on heat-producing genes, mitochondrial content, OCR and the expressions of SMAD7, p-SMAD2, and p-SMAD3 in brown adipocytes. Blocking miR-92a might promote brown adipocytes differentiation, mitochondrial oxidative respiration, and thermogenesis by targeting SMAD7 to inhibit the expressions of p-SMAD2 and p-SMAD3.     

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。     

lncRNA RPL34-AS1通过靶向调控miR-575表达对卵巢癌SKOV3细胞增殖和凋亡的影响

目的探究RPL34-AS1对卵巢癌细胞增殖、迁移的影响及其作用机制。 方法取对数生长期SKOV3细胞用无血清培养基同步化12 h,将pcDNA、pcDNA-RPL34-AS1、si-NC、si-RPL34-AS1、anti-miR-NC、anti-miR-575转染至SKOV3细胞中,分别记为pcDNA组、pcDNA-RPL34-AS1组、si-NC组、si-RPL34-AS1组、anti-miR-NC组、anti-miR-575组;将pcDNA-RPL34-AS1与miR-NC、miR-575分别共转染至SKOV3细胞中,记为pcDNA-RPL34-AS1+miR-NC组、pcDNA-RPL34-AS1+miR-575组。实时荧光定量PCR （RT-qPCR）检测临床组织标本及转染后各组细胞中RPL34-AS1和miR-575的表达水平;双荧光素酶报告实验检测RPL34-AS1和miR-575的靶向关系;四甲基偶氮唑盐比色法（MTT）检测细胞存活率;流式细胞术检测细胞凋亡;蛋白质印迹（Western blot）法检测细胞周期蛋白D1（Cyclin D1）、细胞周期蛋白依赖性激酶抑制剂1A （p21）、B细胞淋巴瘤/白血病-2 （Bcl-2）、Bcl-2相关X蛋白（Bax）蛋白表达水平。两组间比较采用独立样本t检验,多组间比较采用单因素方差分析,组间两两比较采用LSD-t检验。 结果与癌旁组织相比,卵巢癌组织中RPL34-AS1表达水平降低（1.00±0.08比0.47±0.05）,miR-575表达水平升高（1.01±0.07比3.12±0.28）（P < 0.05）。转染si-RPL34-AS1后,细胞活性升高（48 h：0.68±0.06比0.55±0.05;72 h：0.99±0.08比0.71±0.06）,G1期细胞所占比例降低（13.42±1.38比32.15±2.11）,S期细胞所占比例升高（53.75±5.22比34.69±3.41）,细胞凋亡率降低（4.31±0.42比9.25±0.91）,CyclinD1、Bcl-2表达水平升高（0.92±0.08比0.71±0.07;0.86±0.07比0.61±0.06）,p21、Bax表达水平降低（0.13±0.02比0.29±0.03;0.19±0.02比0.31±0.03） （P均< 0.05）。RPL34-AS1靶向调控miR-575,过表达RPL34-AS1或抑制miR-575后可抑制细胞活性,阻滞细胞周期和促进细胞凋亡。miR-575过表达逆转了RPL34-AS1过表达对卵巢癌SKOV3细胞增殖抑制和凋亡促进的作用。 结论过表达RPL34-AS1可抑制卵巢癌SKOV3细胞增殖,促进细胞凋亡,其机制可能与下调miR-575有关。