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Objective:To screen 17-AAG-M-induced differentially expressed miRNAs in human non-small cell lung cancer (NSCLC) A549 cells under X-ray and evaluate its effect on radio-sensitivity.Methods:A549 cells were treated with 17-AAG-M and 4 Gy. Total RNA was extracted for microarray screening. The expression of the miRNAs of interest in the tumor was observed by public database. The target miRNAs were analyzed by using GO and KEGG pathways, and verified by qPCR. The effect of target miRNAs on the survival rate and proliferation of A549 cells under X-ray was evaluated by MTT and clone formation assays. The radio-sensitivity of the target miRNAs was analyzed by the single-hit multi-target model formula.Results:20 differentially expressed miRNAs were screened. The down-regulated hsa-miR-30a-3p showed a close correlation with lung cancer in the database. It was involved with 50 biological processes including cell proliferation and affected the MAPK signaling pathway, cancer-related pathways and cell cycle, etc. Compared with the 17-AAG-M group, the relative expression level of hsa-miR-30a-3p under the action of 17-AAG-M and X-ray was down-regulated from 2.42 to 0.16. hsa-miR-30a-3p inhibited the survival rate of A549 cells (survival rate: 78.52%) and further decreased to 69.00% under X-ray. Up-regulation of hsa-miR-30a-3p expression inhibited the proliferation of tumor cells and increased the radio-sensitivity of A549 cells. The radio-sensitization ratio was 1.18. The above performance became more obvious under the action of 17-AAG-M.Conclusions:In A549 cells, hsa-miR-30a-3p is differentially expressed under the action of 17-AAG-M and X-ray. Moreover, up-regulation of the expression level of hsa-miR-30a-3p in A549 cells can reduce the viability and proliferation of tumor cells, and increase the radio-sensitivity of tumor cells. The inhibition effect of X-ray combined with 17-AAG-M upon tumors can be strengthened.
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@#[摘 要] 胃癌（gastric carcinoma，GC）是最常见的消化系统恶性肿瘤之一，其病因及发病机制等都尚未明了，诊断、治疗及预后仍然面临着严峻的问题。微小RNA（microRNA，miRNA）作为一类重要的基因表达调节剂，能够与其相对应的靶基因相结合，从而影响基因表达，以此在GC发生发展中发挥重要的作用。miR-30家族中5个高度保守且成熟的成员——miR-30a、miR-30b、miR-30c、miR-30d和miR-30e位于不同的染色体或邻近位点，这些miRNA在5'末端附近有一个共同序列，3'末端附近具有不同的补偿序列。通过靶向各自相对应的基因影响着GC细胞的增殖、转移、侵袭和对化学药物的耐药性。miR-30家族在GC中发挥着重要的抑癌作用。本文就近年来miR-30家族中5个成员在GC发生发展中作用的研究进展作一综述。
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OBJECTIVE@#To investigate the differential expression of miR-30a-5p in patients with poststroke depression and explore the possible mechanism.@*METHODS@#We obtained the target microRNAs through searching PubMed using the online software VENNY2.1. We collected the baseline demographic, clinical and radiographic data from consecutive patients with first-ever acute ischemic stroke on admission in our department from October, 2018 to March, 2019. From each patient, 5 mL peripheral venous blood was collected upon admission. Hamilton Depression Scale (HAMD-17) was used to evaluate the degree of depression at the end of the 3-month follow-up. The patients with a HAMD-17 score≥7 were diagnosed to have depression according to the diagnostic criteria of the Fourth Edition of the Diagnostic and Statistical Manual of Mental Disorders of the American Psychiatric Association (DSM-IV). The patients were divided into post-stroke depression group (PSD group, =11) and non-post-stroke depression group (non-PSD group, =25), and their plasma levels of miR-30a-5p were detected using qPCR. The STARBASE Database ENCORI miRNA-mRNA module and Comparative Toxicogenomics Database were used to predict and screen the possible target genes related to miR-30a-5p, and the possible mechanism of the target genes was further analyzed through bioinformatics.@*RESULTS@#miR-30a-5p was identified by cross-screening as the target miRNA associated with stroke and depression and showed obvious differential expression between PSD and non-PSD patients (2.462±0.326 1±0.126, < 0.0001). ROC curve analysis showed that the AUC of miR-30a-5p for predicting PSD was 0.869 (95%: 0.745-0.993, =0.0005) at the cutoff value of 1.597, with a sensitivity and specificity of 0.727 and 0.840, respectively. The target proteins of miR-30a-5p involved a wide range of biological processes, including signal transduction, intercellular communication, regulation of nucleobase, nucleoside, nucleotide and nucleic acid metabolism. KEGG pathway enrichment analysis showed that the target proteins affected mainly the neural nutrient signaling pathway, axon guidance signaling pathway and insulin signaling system. We also identified the top 20 HUB genes that might be associated with post-stroke depression.@*CONCLUSIONS@#Plasma miR-30a-5p is differentially expressed in PSD and can serve as a new blood marker for diagnosis and also a therapeutic target of PSD.
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Humans , Brain Ischemia , Computational Biology , Depression , Gene Expression Regulation, Neoplastic , MicroRNAs , StrokeABSTRACT
microRNAs play an important regulative role in body's growth and development,and the development of the disease process.Much microRNAs can maintain normal kidney function and regulate kidney pathological process,the miR-30a has extensive effect on kidney development and progression of renal diseases.In this review,a brief overview on the role of miR-30a in renal pathology is presented.
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Objective To explore the potential role and function of miR-30 a in myocardial fibrosis after myocardi-al infarction( MI) .Methods We constructed the AAV plasmid vector which carried the miR-30 a gene of rat.The recombinant plasmid was detected by gene sequencing , enzyme digestion and PCR .Virus was packaged into HEK293 cells and virus titer was determined after extraction and purification by PCR .PBS fluid, rAAV9-miR-30 a-NC and rAAV9-miR-30 a were transmited to rat hearts from PBS group , miR-30 a-NC group and miR-30 a group respectively through transcoronary infusion before anterior descending coronary artery ligation .Sham group was set up at the same time.After 4 weeks, heart function was monitored by serial echocardiography , including fractional shortening ( FS) , and left ventricular ejection fraction ( LVEF) .Masson staining was used to calculate collagen volume fraction ( CVF) .The expression of collagen ⅠandⅢwere detected by immunohistochemistry . The mRNA level of miR-30a, TGF-β1 and CTGF were detected by real-time PCR.The protein level of TGF-β1 and CTGF were detected by western blot analysis .Results The cardiac function of miR-30 a group was improved significantly compared with PBS group and miR-30a-NC group (P<0.05).The levels of CVF,collagenⅠ,Ⅲexpression and Collagen Ⅰ/Ⅲ ratio in miR-30 a group was significantly lower than PBS group and miR-30 a-NC group ( P<0.01 ) .The mRNA and protein level of TGF-β1 and CTGF in miR-30 a group were reduced signifi-cantly than PBS group and miR-30 a-NC group ( P<0.001 ) .Conclusions The overexpression of miR-30 a after MI may reduce the mRNA and protein level of TGF-β1 and CTGF, so as to suppress myocardial fibrosis and im-prove cardiac function.
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Objective To investigate the micrometastatic lesion of tumor stem cells in the axillary swollen lymph nodes of breast cancer patients and the influence of miR30a on its invasive ability ,and to explore the feasibility of miRNAs anti‐breast cancer treatment .Methods The tumor stem cell‐like breast cancer cells were separated from the axillary swollen lymph nodes in breast cancer patients and cultured .miR30a oligonucleotide fragment was synthesized and transfected into human primary generation breast cancer cells by using adenovirus ,meanwhile the breast cancer cell line MDA‐MB‐231 was taken as the experimental control and the transfection efficiency was assessed by the fluorescence microscopy .The changes of tumor cell proliferation and invasiveness before and after transfection were detected by the Transwell chamber in vitro invasion assay .Western blot was used to detect the ALDH1 ,Vimentin and N‐Cadherin protein expression .Results The Transwell chamber in vitro invasion assay showed that the pri‐mary generation breast cancer cell had more strong invasive ability than MDA‐MB‐231 cell line ,their invasion indexes were (75 .3 ± 3 .2)% and (58 .4 ± 2 .8)% respectively ,the difference was statistically significant (P<0 .05) ,and after transfecting miR30a ,the in vitro invasiveness ability in these two kinds of cells were significantly weakened and their invasion indexes were (21 .4 ± 1 .9)% and (28 .2 ± 2 .3)% respectively ,the difference compared with the control group showed the statistical significance (P<0 .05) .Conclu‐sion The ALDH expression in partial axillary hyperplasia and swollen lymph nodes in the patients with breast cancer is increased , and the tumor micrometastasis may exist ,which should be completely cleaned in operation .miR30a inhibits the expression and inva‐sive ability of tumor stem gene .
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Objective To investigate the effect of miR-30a on human osteosarcoma cell 143B in migration,invasion andcellviability.Methods 143BcellswereinfectedortransfectedwithrecombinantadenovirusmiR-30a(Ad-miR30a) and miR-30a inhibitor respectively .Wound healing assay was performed to detect the cell healing ability ( P<0.05 ) .Cell migration and invasion ability were determined by Transwell assay ( P<0.05 ) .The cell viability was analyzed by MTT assay ( P<0.01 ) .Real-time quantitative PCR was performed to analyze the expression of RUNX2 mRNA level and confirmed the adenovirus miR-30a expressed in 143B cells.The expression of RUNX2 was analyzed by Western blot .miR-30a target to RUNX2 was verified by luciferase reported gene assay .Results The ability of migration and invasion was suppressed in osteosarcoma cell 143B by overexpression miR-30a,and the cell viability also decreased .After the endogenous miR-30 a being inhibited , the cell motility and invasion enhanced and the cell viability was promoted .The RUNX2 protein decreased after overexpression miR-30 a as compared with controlgroup.TheluciferaseactivityofRUNX2decreasedbyaddingmiR-30a.Conclusions 143Bcellmigration, invasion and viability were suppressed by miR-30a,and this process is potentially achieved via suppressing RUNX 2 protein expression .
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Objective:Mesenchymal stem cells( MSCs) have self-renewal capacity and potential to differentiate into the cells.It was reported that the expression of miR-30a changed in some immune diseases.But it remains unclear the effect of miR-30a on the im-munoregulatory functions of MSCs.Here we studied the impact of miR-30a on the phenotype,cell viability,apoptosis,cell cycle and im-munoregulatory functions of MSCs.Methods: The mixed enzyme methods were used for the isolation of human umbilical cord MSCs.Flow cytometry(FCM)was used to investigate the effect of overexpressed miR-30a on the phenotype of MSCs.CCK-8 was used to examine the cell viability of miR-30a-overexpressed MSCs.Annexin V/PI was used for the detection of apoptosis of MSCs.Q-PCR and Western blot were used to investigate the effect of miR-30a on the expression of Cyclin E2( CCNE2).CCNE2 was one putative target of miR-30a predicted by Targetscan database.The effects of miR-30a-overexpressed MSCs on the maturation of dendritic cells(DCs)were determined.Results:Overexpression of miR-30a blocked the cell cycle of MSCs in the G0/G1 phase by inhibiting the expression of CCNE2,but did not affect the phenotype, cell viability and apoptosis of MSCs.When co-cultured with DCs, although MSCs down-regulated the expression of CD40 and CD86 on DCs,overexpression of miR-30a more significantly enhanced the suppressive impact of MSCs on the maturation of DCs.Conclusion: miR-30a affects the cell cycle of MSCs and enhances its immunosuppressive effect on DCs.
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Aims To investigate the protective effects of noninvasive limb ischemic preconditioning (LIPC) on myocardial ischemia-reperfusion (I /R)injury,and to explore the mechanism.Methods Healthy male Wistar rats were divided randomly into I /R,I /R +LIPC,I /R +5-Hydroxydecanoate (5-HD)and I /R +LIPC +5-HD groups.The I /R +LIPC and I /R +LIPC-5-HD groups of rats were subjected to three cy-cles of LIPC induction per day with 5 min of reperfu-sion after occlusion for 5 min at the left hind limb for 3 days.All rats were subjected to myocardial I /R injury on the fourth day.The I /R +5-HD and I /R +LIPC-5-HD groups of rats were given the inhibitor of ATP-sen-sitive potassium channel 5-HD before and during myo-cardial I /R injury. Results Compared with I /R group,LIPC reduced myocardial infarct size (P <0.05),lowered cardiocyte apoptosis index and Fas, FasL positive cell number (P <0.01 ),increased the reduced nitric oxide (NO)/endothelin (ET)-1 ratio (P <0.05)in serum in I /R +LIPC group.5-HD a-bolished the protective effects induced by LIPC in I /R+LIPC-5-HD group.Compared with normal myocardi-al tissue,expression of mir-30a-3p was increased in I /R group (P <0.01 )and was decreased in LIPC group (P <0.01 ).Conclusion LIPC alleviates myocardial I /R injury and improves endothelial function. The mechanism may be related with the opening of ATP-sensitive potassium channel,regulating the balance be-tween NO and ET-1 and decreasing the expression of myocardial mir-30a-3p.
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Objectives To explore the relationship of serum miR-30a and cardiac function in children with congenital heart disease (CHD), and the diagnostic value of serum miR-30a to heart failure. Methods From January 2013 to June 2013, 65 children with CHD were enrolled, and divided into the HF group (patients with heart failure, n=40) and the Non-HF group (patients without heart failure, n=25) according to modified Ross score. The serum miR-30a expression level was quantified by real-time PCR. Left ventricular ejection fraction (LVEF) was tested by echocardiography. The correlation between miR-30a and modified Ross score and LVEF were evaluated with Spearman's analysis. The area under the receiver-operating characteristic (ROC) curve for miR-30a was examined. Results The expression level of miR-30a in HF group was higher than that in Non-HF group, the LVEF was lower in HF group than that in Non-HF group (all P<0.001). The serum miR-30a level was positively correlated with modified Ross score (r=0.63, P<0.01) and negatively correlated with LVEF (r=-0.32, P<0.01). According to ROC analysis, the AUC of miR-30a for detection of HF was 0.7440. Conclusion The expression level of serum miR-30a was closely correlated with the cardiac function, which can be as a biomarker for indicating the heart failure in CHD.
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Objective To study the inhibitory effect of knocking down miR-30a-5p on the U87 human glioma xenograft growth and its possible mechanism.Methods Nude mice bearing subcutaneous U87 human glioblastoma were established and separated into three groups (eight for each group) by randomized digital table method,including control group,scr-ODN treated group and AS-miR-30a-5p treated group.After relevant subcutaneous injection treatment,tumor size was measured every other day until the observation period ended.Researchers executed the animals after the treatment,stripped tumor tissues and extracted RNA and protein.Real-time PCR was conducted to detect the expression of miR-30a-5p.The histopathological characteristics and proliferation and apoptosis biological characters (including SEPT7,PCNA,cyclin D1,MMP-2,apoptosis related factor P53,bcl-2 and caspase3) were evaluated by HE and immunohistochemical staining,Westem blot analysis respectively,and the cell apoptosis was detected by TUNEL method.Results In AS-miR-30a-5p treated group,the tumor growth was delayed and the final tumor volume was smaller than that in the control and scr-ODN treated group (F =7.167,P ＜0.05),and the expression of miR-30a-5p was knocked down.The expression of PCNA,cyclin D1 were significantly downregulated while P53,SEPT7 and caspase3 up-regulated.Apoptotic index was increased significantly.Conclusion As-miR-30a-5p suppresses the growth of U87 human gliomas xenografts significantly.Malignant phenotype of tumors are reversed to a considerable degree.Therefore,miR-30a-5p can be a candidate for targeted therapy of human glioma.