Relationship between expression of Smac and Survivin and apoptosis of primary hepatocellular carcinoma

BACKGROUND: T he second mitochondria-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pI (Smac/DIABLO) was recently identified as a protein that is released from mitochondria in response to apoptotic stimuli and promotes apoptosis by antagonizing inhibitor of apoptosis proteins. Furthermore, Smac/DIABLO plays an important regulatory role in the sensitization of cancer cells to both immune- and drug-induced apoptosis. However, little is known about the clinical significance of Smac/DIABLO in various cancers including hepatocellular carcinoma (HCC). This study was undertaken to investigate the expression of Smac and Survivin and their relationship with the apoptosis in primary HCC. METHODS: The expression of Smac and Survivin proteins was evaluated by immunohistochemistry. The mRNA expression of Smac and Survivin was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) in HCC tissues of 50 patients, para-carcinoma tissues of 20 patients, and normal liver tissues of 15 patients. RESULTS: Smac mRNA was detected by RT-PCR in HCC tissues of 21 (42.0%) of the 50 patients, para-carcinoma tissues of 19 (95.0%) of the 20 patients, and normal liver tissues of 15 (100%) of the 15 patients. Survivin mRNA was found in HCC tissues of 46 of the 50 patients, para- carcinoma tissues of 2 of the 20 patients, and normal liver tissues of 0 of 15 patients. Immunohistochemistry revealed Smac protein in HCC tissues of 20 patients (40.0%), in para-carcinoma tissues of 18 patients (90.0%), and normal liver tissues of 15 patients (100.0%). The expression of Smac was significantly different in HCC tissues and non- HCC tissues. Survivin protein was found in HCC tissuesin 45 patients, para-carcinoma tissues in 2 patients, and normal liver tissues in none of the patients. The expression of Survivin was significantly different in HCC tissues and non-HCC tissues. CONCLUSION: Smac inhibits apoptosis of HCC cells by suppression of Survivin, and the two genes probably form an important link in the signal pathway of HCC cells.

The Inhibition of Apoptosis of Hepatoma Cells Induced by HBx Is Mediated by Up-regulation of Survivin Expression

To investigate the effect of HBx on expression of survivin in hepatoma cells and mechanisms of inhibition of apoptosis on hepatoma cells induced by HBx, the expression plasmid pHA HBx encoding full length of HBx was transfected into HepG2 cells and the transformed cells were identified by RT PCR. The expression of survivin both in HepG2 cells and HBx transfected cells was examined with RT PCR. The nude mice model of hepatoma was established by injecting HepG2 cells and HBx transfected cells into the flank of nude mice subcutaneously. The expression level of survivin both in HepG2 formed tumors and HBx transfected cell formed tumors in nude mice was examined with Western blot. The TUNEL assay was used to detect the apoptotic cells of tumor tissues in nude mice after intraabdominal chemotherapy with adriamycin. The results indicated that the amplification of survivin in HBx transfected HepG2 cells was up regulated when compared with that in non transfected cells. Western blot showed that the tumor cells expressing HBx in nude mice had a positive band of survivin expression and the tumor cells without HBx expression had no positive band. The result of TUNEL assay showed that there were less apoptotic cells in tumor tissues expressing HBx than that in control group cells. It was concluded that HBx could up regulate the expression of survivin in hepatic carcinoma cells, which can inhibit apoptosis of hepatic carcinoma cells induced by adriamycin.

miR-145-5p调控Survivin通过凋亡通路对食管癌生物学行为的影响

目的:探讨miR-145-5p靶向调控Survivin及凋亡信号通路对食管癌凋亡、侵袭和迁移的影响。方法:采用实时荧光定量聚合酶链反应(RT-qPCR)检测食管癌组织和细胞系中miR-145-5p、Survivin mRNA相对表达量。双荧光素酶报告基因分析验证miR-145-5p和Survivin的靶向调控关系。通过RT-qPCR法测定细胞转染后miR-145-5p和Survivin mRNA的相对表达量。Western blot检测凋亡通路中BCL2、MDM2和Caspase-3蛋白的表达。通过MTT、流式细胞术、划痕愈合实验及Transwell小室实验检测TE-1细胞存活能力、凋亡率、迁移率及侵袭能力。结果:与癌旁组织相比,miR-145-5p在癌组织中表达低,而Survivin表达高(P<0.05)。与正常食管黏膜上皮HEEC细胞相比,食管癌TE-1细胞中miR-145-5p低表达,Survivin高表达(P<0.05)。双荧光素酶报告基因实验证实,miR-145-5p和Survivin间存在靶向调控关系。与未转染和转染miR-145-5p对照物的TE-1细胞相比,转染miR-145-5p模拟物后可显著增加miR-145-5p的表达,抑制Survivin mRNA的表达,同时BCL2和MDM2蛋白的表达下降,Caspase-3表达上升,促进细胞凋亡,抑制了细胞活性、侵袭和迁移能力(P<0.05)。结论:通过miR-145-5p的负向调控,Survivin表达被抑制,凋亡通路被激活,从而促进细胞凋亡,抑制食管癌细胞存活,侵袭和迁移能力。

Expression of a novel apoptosis inhibitor-survivin in colorectal carcinoma

AIM: To investigate the role of survivin expression in the pathogenesis of colorectal carcinoma.METHODS: Immunohistochemistry S-P method and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) were used to detect the expression of survivin and apoptotic cell in situ in colorectal cancerous tissues, para-cancerous tissues and normal tissues of 48 cases of colorectal carcinoma.RESULTS: The survivin positive unit (PU) was higher in cancerous tissues (38.76±5.14)than in para-cancerous (25.17±7.26) or normal tissues (0.57±0.03) (P<0.05).The apoptosis index (AI) of para-cancerous tissues was(7.51±2.63%) higher than cancerous tissues (4.65±1.76%).The expression of survivin was associated with pathological grade, lymph node metastasis and Dukes stage of colorectal carcinoma.CONCLUSION: Survivin expression may play an important role in carcinogenesis of colorectal carcinoma and may be associated with malignant biological behaviors of colorectal carcinoma.

Survivin antisense compound inhibits proliferation and promotes apoptosis in liver cancer cells

AIM: To evaluate the effects of survivin on cell proliferation and apoptosis in liver cancer.METHODS: MTT assay was used to generate and optimize phosphorothioate antisense oligonucleotides (ODNs)LipofectamineTM2000 (LiP) compound by varying ODNs (μg):LiP (μL) ratios from 1:0.5 to 1:5. Then, liver cancer cells (HepG2) were transfected with the compound. By using RT-PCR and Western blot, the expression levels of survivin mRNA and proteins were detected in HepG2 cells treated with antisense compounds (ODNs:LiP = 1:4), and compared with those treated with sense compounds (1:4) as control.MTT assay was applied to the determination of cell proliferation in HepG2 cells. Active caspase-3 was evaluated by flow cytometric analysis. The morphological changes were assessed by electron microscopy. Laser scanning confocal microscopy was performed to detect the subcellular localization of survivin proteins in treated and untreated cells.RESULTS: Antisense compounds (1:4) down-regulated survivin expression (mRNA and protein) in a dose-dependent manner with an IC50 of 250 nmol/L. Its maximum effect was achieved at a concentration of 500 nmol/L, at whichmRNA and protein levels were down-regulated by 80%.The similar results were found in MTT assay. Antisense compound (1:4)-treated cells revealed increased caspase3-like protease activity compared with untreated cells.Untreated cells as control were primarily negative for the presence of active-caspase-3. As shown by transmission electron microscopy, treated cells with antisense compounds (1:4) resulted in morphological changes such as blebbing and loss of microvilli, vacuolization in the cytoplasm,condensation of the cytoplasm and nuclei, and fragmented chromatin. Immunofluorescence analysis confirmed the presence of survivin protein pool inside the cytoplasm in untreated cells. Labeled-FITC immunofluorescence staining of survivin clearly showed that survivin was distributed mainly in a spotted form inside the cytoplasm. Whereas cells treated with antisense compounds were rare and weak inside the cytoplasm.CONCLUSION: Down-regulation of survivin expression induced by the antisense compounds reduces tumor growth potential, promotes apoptosis and affects the localization of survivin proteins in HepG2 cells. Furthermore, survivin protein is a key molecule associated with proliferation and apoptosis, and antisense oligonucleotides targeting survivin have a bright prospect in the therapy of liver cancer.

Antisense RNA of Survivin Gene Inhibits the Proliferation of Leukemia Cells and Sensitizes Leukemia Cell Line to Taxol-induced Apoptosis

The effects of survivin antisense RNA on proliferation of leukemia cell line HL-60 and taxol-induced chemotherapy was explored. A cDNA fragment of survivin obtained by RT-PCR was inserted into a plamid vector named pcDNA3 in the reverse direction. The vector encoding antisense RNA of survivin was confirmed by restriction enzyme digestion and DNA sequencing. The recombinant plasmid was delivered into HL-60 cells by electroporation. Growth curves were plotted based on cell counting. Trypan blue dye exclusion assay and MTT assay were carried out after the cells were incubated with taxol. DNA gel electrophoresis and nuclear staining were performed for cell apoptosis assay. The correct construction of the recombinant plasmid has been identified by restriction enzyme digestion and DNA sequencing. A stable down-regulation has been achieved in HL-60 SVVas cells after G418 selection. Compared to HL-60 cells, the proliferation of HL-60 SVVas cells was significantly inhibited （P〈0.05）. Cytotoxicity assays indicated that IC50 of HL-60 SVVas for taxol was relatively lower than controls （P〈0.01）. Apoptosis assays revealed that taxol-induced apoptosis was detected in HL-60 SVVas cells incubated with 50 ng/ml taxol for 12 h, while in HL-60 cells incubated with 100 ng/ml taxol for 72 h. It was suggested that Survivin antisense RNA could inhibit the proliferation of HL-60 cells and enhance taxol-induced apoptosis in HL-60 cells, which may lay an experimental foundation for further research on gene therapy in leukemia.

Knockdown of survivin gene expression by RNAi induces apoptosis in human hepatocellular carcinoma cell line SMMC-7721

AIM: To investigate the survivin gene expression in human hepatocellular carcinoma cell line SMMC-7721 and the effects of survivin gene RNA interference (RNAi) on cell apoptosis and biological behaviors of SMMC-7721 cells.METHODS: Eukaryotic expression vector of survivin gene RNAi and recombinant plasmid pSuppressorNeo-survivin (pSuNeo-SW), were constructed by ligating into the vector,pSupperssorNeo (pSuNeo) digested with restriction enzymes Xba I and Sa/I and the designed double-chain RNAi primers. A cell model of SMMC-7721 after treatment with RNAi was prepared by transfecting SMMC-7721 cells with the lipofectin transfection method. Strept-avidinbiotin-complex (SABC) immunohistochemical staining and RT-PCR were used to detect survivin gene expressions in SMMC-7721 cells. Flow cytometry was used for the cell cycle analysis. Transmission electron microscopy was performed to determine whether RNAi induced cell apoptosis, and the method of measuring the cell growth curve was utilized to study the growth of SMMC-7721 cells before and after treatment with RNAi.RESULTS: The eukaryotic expression vector of survivin gene RNAi and pSuNeo-SW, were constructed successfully. The expression level of survivin gene in SMMC-7721 cells was observed. After the treatment of RNAi, the expression of survivin gene in SMMC-7721 cells was almost absent,apoptosis index was increased by 15.6%, and the number of cells was decreased in G2/M phase and the cell growth was inhibited.CONCLUSION: RNAi can exert a knockdown of survivin gene expression in SMMC-7721 cells, and induce apoptosis and inhibit the growth of carcinoma cells.

Differential regulation of survivin by p53 contributes to cell cycle dependent apoptosis

Recent studies indicate that cell-cycle checkpoints are tightly correlated with the regulation of apoptosis, in which p53 plays an important role. Our present works show that the expression of E6/E7 oncogenes of human papillomavirus in HeLa cells is inhibited in the presence of anti-tumor reagent tripchlorolide (TC), which results in the up-regulation of p53 in HeLa cells. Interestingly, under the same TC-treatment, the cells at the early S-phase are more susceptible to apoptosis than those at the middle S-phase although p53 protein is stabilized to the same level in both situations. Significant difference is exhibited between the two specified expression profiles. Further analysis demonstrates that anti-apoptotic gene survivin is up-regulated by p53 in the TC-treated middle-S cells, whereas it is down-regulated by p53 in the TC-treated early-S cells. Taken together, the present study indicates that the differential p53-regulated expres- sion of survivin at different stages of the cell cycle results in different cellular outputs under the same apoptosis-inducer.

Knockdown of Survivin Expression by siRNA Induces Apoptosis of Hepatocellular Carcinoma Cells

Survivin, a newly identified member of lAP family, is a powerful apoptosis-inhibiting factor. It is expressed in embryonic tissues as well as in the majority of human cancers, but not in most normal adult tissues. The cancer-specific expression of survivin makes it a potential target for cancer treatment. A survivin-specific small inhibitory RNA （siRNA） was introduced into hepatocellular carcinoma cells to investigate its effect on cancer cell apoptosis, growth and sensitivity to chemotherapeutic drugs. It was found that expressions of survivin protein and proliferation index （PI） in siRNA groups were significantly decreased, the apoptosis index （AI） of siRNA groups was significantly higher than those of others groups, and the growth inhibition rate （GIR） of chemotherapeutic drugs in siRNA groups were significantly higher than those of other groups. Our study suggests that the expression of survivin may be significantly decreased in hepG2 cell after siRNA transfection. siRNA targeting survivin could induce cell apoptosis, inhibit cell proliferation and sensitize hepatocarcinoma cells to chemotherapy. Our findings provide preliminary evidence for the therapeutic use of survivin-targeted RNA interference for human tumors that express high levels of this molecule.

Survivin small interfering RNA suppresses glioblastoma growth by inducing cellular apoptosis

A survivin small interfering RNA sequence specific for a human and mouse homogenous sequenc~ was constructed. Survivin small interfering RNA could significantly inhibit glioma cell proliferation and induce apoptosis when it was transfected into either a human glioma cell line U251 or rat glioma C6 cells in vitro. In addition, treatment of rat orthotopic glioma models with survivin small interfering demonstrated the inhibition of glioma growth in vivo. Our experimental findings suggest that the use of RNA interference techniques to target the survivin sequence may be useful in the treatment of glioma.

Effects of shRNA Targeting Survivin on Apoptosis of Human Retino-blastoma Cell Line Hxo-rb44 in vitro

In order to construct a recombinant plasmid containing short hairpin RNA （shRNA） targeting survivin and to investigate its effect on survivin expression and cell apoptosis of human retinoblastoma cell line Hxo-rb44 in vitro, RNA interference plasmid pSIRENS that can express shRNA of survivin was designed, constructed, and transfected into human retinoblastoma cell line Hxo-rb44, Survivin and c-Myc expression was detected by reverse transcription polymerase chain reaction （RT-PCR） and Western blot. Apoptosis of Hxo-rb44 cells was assayed by Honchest33258 staining and cell growth curve was drawn. The results showed that the oligonucleotide targeting survivin was identified in pSIRENS plasmid. After pSIRENS plasmid transfected, survivin and c-Myc expression in Hxo-rb44 cells was decreased significantly. Apoptotic rate of cells was up-regulated from （3.5±1.29） % to （36.1±19.66） %. The proliferation ability of Hxo-rb44 cells was inhibited. No significant effects on survivin expression and apoptosis of the cells were found when negative control plasmid was transfected. In conclusion, the plasmid containing shRNA targeting survivin was constructed successfully. It could inhibit efficiently the expression of survivin and c-Myc in human retinoblastoma cell Hxo-rb44 in vitro. The inhibition of the expression of c-Myc might be involved in the apoptosis of Hxo-rb44 cells.

Expression of Survivin, p53 and its relationship with apoptosis, proliferation in hepatocellular carcinoma(HCC)

Objective：To investigate the expression of Survivin p53 and its relationship with apoptosis, proliferation in hepatocellular carcinoma （HCC）. Methods：The expression of Survivin, p53 and the proliferation of tumor cells marked by proliferation cell nuclear antigen （PCNA） in 42 cases of HCC were assessed by immunohistochemical method. TUNEL was used to detect apoptosis. Results：Survivin protein was expressed in 30 of 42 cases of HCC（71.4%） and in 4 of 34 cases of adjacent cirrhosis tissues（11.8%）. Expression of Survivin protein was negative in 10 cases of normal tissues. Survivin protein positive expression rate in HCC was significantly higher than adjacent cirrhosis tissues and normal tissues（P 〈 0.001）. The increased Survivin protein expression in cancer was significantly associated with histological grade（P = 0.003）, p53 protein（P = 0.013）, survival time（P 〈 0.05） and the ratio of proliferative index to apoptotic index（P〈 0.01）, but was not significantly correlated with age, sex, clinical stage, tumor size, metastasis, AFP, HBs-Ag（P 〉 0.05）. Conclusion：There is a marked increased expression of Survivin in HCC, which may play an important role in breaking the balance of proliferation and apoptosis of HCC cells. The correlation between Survivin and p53 expression in HCC indicates that cooperation between Survivin and p53 plays a certain role in occurrence and/or development of HCC.

Impact of resveratrol on the expression of apoptosis related gene survivin and bax in human cancer cells

客观我们由 resveratrol 在人的食道的癌症 Eca109 房间探索了 apoptosis 的机制。Eca109 房间增长上的 resveratrol 的镇压比率被 MTT 比色的试金和形态学评估的方法被传播电子显微镜观察。survivin 和 bax 的表示被 RT-PCR 和流动 Cytometry (FCM ) 分析。结果 Resveratrol 在剂量禁止了 Eca109 房间的生长 -- 并且时间依赖者举止，和镇压比率在 76.42% 点到了。在与 resveratrol 对待以后，词法 apoptosis 能被观察。一些对待药的房间的体积变得小，原子染色质变得压缩并且有边缘。RT-PCR 和 FCM 决定的结果显示出那 resveratrol 能下面调整幸存，当时起来调整 bax。结论 Resveratrol 能导致人的食道的癌症 Eca109 房间的 apoptosis，并且它的可能的分子的机制可能与调整有关是 survivin 和 bax 的表示。

Influence of Ginkgo biloba extract on the proliferation,apoptosis of ACC-2 cell and Survivin gene expression in adenoid cystic carcinoma of lacrimal gland

Objective:To explore the influence of extract of Ginkgo biloba（EGB） on the proliferation, apoptosis of ACC-2 cell and Survivin gene expression in adenoid cystic carcinoma（ACC） of lacrimal gland.Methods:ACC-2 cell in human with ACC of lacrimal gland was in vitro cultured. MTT method was used for cell proliferation detection.Annexin V/PI double-staining flow cytometer was used to detect cell apoptosis and cell cycle.Survivin gene expression was analyzed by RT-PCR and Western blotting.Results:EGB had inhibitory effect on the proliferation of ACC-2 cell with significant dose-effect relationship,and there was statistical difference when compared with the control group（P【0.01）.The inhibitory concentration 50%(IC50) is 88 mg/L. The flow cytometer test indicated that CGB can gradually increase ACC-2 cell in G0-G1 stage and decrease it in G2-M and S stage.With the increase of dose,the apoptosis rate of ACC-2 cell was obviously increased（P【0.05 or P【0.01）.EGB had certain inhibitor)’ effect on Survivin gene expression of ACC-2 cell,and Survivin gene expression was decreased with the increasing of the EGB concentration（P【0.01）.Conclusions:EGB can effectively inhibit Survivin gene expression of ACC-2 cell in human with ACC of lacrimal gland,induce the apoptosis of ACC-2 cell and inhibit tumor cell proliferation.

Experimental Study on Induction of Renal Cell Carcinoma Apoptosis by Antisense Oligonucleotide Targeting Survivin

The effect of antisense oligonucleotide (ASODN) targeting survivin on renal cell carcinoma (RCC) apoptosis, proliferation and sensitivity to chemotherapeutic drugs was investigated. Antisense oligonucleotide targeting survivin was designed and composed. The cDNA of survivin was transfected into RCC cell lines ACHN and 769-P, respectively. They were both cultured in normal condition. After transfection, ACHN and 769-P cell lines were cultured in a 6-well culture plate. The cultured cells were divided into 6 groups. Twenty-four h after transfection, survivin protein expression was detected by Western blot. Apoptotic index (AI) and proliferative index (PI) were examined by flow cytometry. Rate of inhibition (IR) by chemotherapeutic drugs was determined by the colorimetric MTT cell viability/proliferation assay. The results showed AI and IR of chemotherapeutic drugs in ASODN groups were significantly higher than those in the groups without ASODN. There were statistical differences between ASODN groups and control groups (P< 0.05), but there was no difference among control groups (P>05). The PI in the ASODN groups was significantly lower than that in the control groups with the difference being statistically significant (P<0.05). In addition, there were no differences among control groups. It was concluded that the expression of survivin could be down-regulated in 2 cell lines after ASODN transfection. Antisense oligonucleotide targeting survivin could induce RCC apoptosis, inhibit cell proliferation and sensitize RCC to chemotherapy.

Down Regulation of Survivin Gene and Up Regulation of p53 Gene expression by siRNA Induces Apoptosis in human Hepatocellular Carcinoma cell Line HepG2

Survivin gene may be a good target for cancer gene therapy because it is over expressed in a variety of human tumors including human hepatocellular carcinoma but not in differen- tiated adult tissues. To explore the effects of the siRNA of survivin gene inducing apoptosis in human hepatocellular cancer cells, three siRNAs cpusiRNA1, cpusiRNA2 and cpusiRNA3 were designed and transferred into human hepatocellular carcinoma cell line HepG2 (HepG2) by lipofection. MTT test showed that the growth of HepG2 decreased when it was transfected with 25nM, 50nM, 100nM, 150nM, 200nM, 400nM siRNA respectively after 48 hours. And the change of mRNA and protein of survivin gene and p53 gene had been detected by RT-PCR and Western blot. Cells presented an increase in apoptosis index was assayed by flow cytometry. Small interfering RNA can exert a knockdown of survivin gene expression and up regulation of p53 gene to induce apoptosis and to inhibit the growth of HepG2.

Study on the apoptosis of Raji cell line induced by arsenic trioxide and its correlation with Survivin gene

Objective: To investigate the apoptosis induction by arsenic trioxide (As2O3) in Raji cells and its correlation with cell cycle arrest and expression of the Survivin gene. Methods: After Raji cells were treated with As2O3 in different concentrations (1, 2, 4 and 8 μM), for 24, 48 and 72 h, respectively, and cell proliferation was tested by MTT assay. Apoptosis was observed with electron microscope and DNA electrophoresis. The distribution of cell cycles and cell apoptosis were detected by flow cytometry. Expression of the Survivin gene was determined by real-time quantitative RT-PCR. Results: As2O3 (1–8 μM) inhibited Raji cells growth effectively in a dose- and time-dependent manner. As2O3 at 2–8 μM could induce cell apoptosis and cell cycle arrest. However, As2O3 (1 μM) inhibited Raji proliferation only by cell cycle arrest, without any symptoms of cell apoptosis. At the same time, Survivin gene expression was down-regulated after the treatment. Conclusion: As2O3 could induce substantial proliferation inhibition, cell cycle arrest and apoptosis in Raji cell. Cell cycle arrest might be a reason why apoptosis occurs. As2O3 can markedly down-regulate expression of the Survivin gene in a dose- and timedependent manner. The down-regulated Survivin gene might be leading to cell apoptosis by As2O3.

Modulating effects of survivin antisense oligonucleotide on changes of apoptosis and cell cycle of human hepatocellular carcinoma cell line SMMC-7721

To investigate the modulating effects of survivn antisense oligonucletode (ASODN) on the cell cycle and apoptosis of human hepatocellular carcinoma (HCC) cell line SMMC-7721 and explore its mechanism.Methods Survivin ASODN was transfected into SMMC-7721 cells mediated by DOTAP liposomal reagent.Electron microscopy,flow cytometry and RT-PCR were used to detect the changes in cell ultrastructure,apoptosis,cell cycle and the expression of cyclinB1 mRNA,respectively.Results After transfection of survivin ASODN,the expression of cyclinB1 mRNA in the cells significantly increased and increase in G2-M arrest and apoptosis appeared.Meanwhile,the cell ultrastructure had apoptotic changes such as chromatin condensation and apoptotic body formation.Conclusion Survivin ASODN can induce the expression of cyclinB1 that may result in G2-M arrest.Consequently,apoptosis is triggered.Survivin ASODN transfection might be an improtant new treatment for HCC.14 refs,2 figs,1 tab.

miR-26a-3p靶向调控Survivin表达对H9c2心肌细胞缺氧/复氧损伤的影响

目的探讨miR-26a-3p对缺氧/复氧(H/R)诱导的大鼠心肌细胞(H9c2)损伤的影响及其机制。方法取对数生长期H9c2心肌细胞进行缺氧(1%O 2)6 h,复氧不同时间(2、4、8、12 h)建立细胞H/R模型,另设常氧组,细胞计数试剂盒(CCK-8)检测细胞增殖活性;比色法检测细胞上清液中乳酸脱氢酶(LDH)水平;实时荧光定量PCR(qRT-PCR)检测细胞中miR-26a-3p和生存素(Survivin)mRNA表达水平;Western blot检测细胞中Survivin蛋白表达水平。将miR-26a-3p inhibitor及其阴性对照inhibitor NC、Survivin基因siRNA干扰质粒(si-Survivin)及其阴性对照si-NC共转染至H9c2细胞中,随后进行H/R干预,CCK-8检测各组细胞增殖水平;比色法检测各组细胞中超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量及上清液中LDH水平;流式细胞术检测各组细胞凋亡水平;Western blot检测各组细胞中Bcl-2相关X蛋白(Bax)、B淋巴细胞瘤-2(Bcl-2)、活化型半胱天冬酶-3(cleaved caspase-3)和Survivin蛋白表达水平;双荧光素酶测定miR-26a-3p和Survivin基因的靶向关系。结果与常氧组细胞比较,随着复氧时间的延长,H9c2细胞增殖活性、Survivin mRNA和蛋白表达水平逐渐降低(P<0.05),而LDH及miR-26a-3p表达水平逐渐升高(P<0.05)。下调miR-26a-3p表达可提高H/R暴露下H9c2细胞增殖活性、SOD活性、Bcl-2蛋白表达水平(P<0.05),同时降低MDA含量、LDH释放量、凋亡率、Bax和cleaved caspase-3蛋白表达水平(P<0.05);而Survivin缺失可明显逆转miR-26a-3p inhibitor对H/R诱导下H9c2细胞的保护作用。双荧光素酶报告基因实验表明Survivin是miR-93-5p的靶基因。结论miR-26a-3p在H/R诱导的心肌细胞损伤中高表达,抑制miR-26a-3p表达可通过靶向上调Survivin表达抑制H/R诱导的心肌细胞凋亡和氧化应激。