miR-153 promotes LPS-induced myocardial H9C2 cell injury by targeting SORBS2

AIM: To study the effects of microRNA-153 (miR-153) on inflammatory factors, cell viability and apoptosis of embryonic rat H9C2 cardiomyocytes induced by lipopolysaccharide (LPS), and to explore its mechanism. METHODS: The injury model of H9C2 cells was established by LPS stimulation. The H9C2 cells were divided into anti-miR-Con group (transfected with anti-miR-Con), anti-miR-153 group (transfected with anti-miR-153), pcDNA group (transfected with pcDNA), pcDNA-SORBS2 group (transfected with pcDNA-SORBS2), anti-miR-153+si-Con group (co-transfected with anti-miR-153 and si-Con) and anti-miR-153+si-SORBS2 group (co-transfected with anti-miR-153 and si-SORBS2), and treated with LPS after transfection. The expression of miR-153 and SORBS2 mRNA in the cells was detected by RT-qPCR. The viability of H9C2 cells was measured by MTT assay. The protein expression of SORBS2 in the H9C2 cells was determined by Western blot. The contents of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were detected by ELISA. The apoptosis of the H9C2 cells was analyzed by flow cytometry. The targeting relationship between miR-153 and SORBS2 was verified by dual-luciferase reporter assay. RESULTS: The LPS-induced H9C2 cell injury model was successfully constructed. Compared with PBS group, the expression of miR-153 was significantly increased and the expression of SORBS2 was significantly decreased in the H9C2 cells treated with LPS. The inhibition of miR-153 and over-expression of SORBS2 decreased the contents of TNF-α and IL-6 and the level of apoptosis, but increased the cell viability. miR-153 inhibited the luciferase activity of the H9C2 cells containing wild-type SORBS2. Inhibition of SORBS2 reversibly inhi-bited the anti-inflammatory effects of miR-153 on LPS-induced H9C2 cells and increased the viability of the cells. CONCLUSION: miR-153 promotes the secretion of inflammatory factors, induces apoptosis, and inhibits the viability of H9C2 cells induced by LPS, thus enhancing the damage. Its mechanism may be related to targeting SORBS2, which will provide new targets for the treatment of myocardial injury.     

Effects of resveratrol on viability,invasion and autophagy of osteosarcoma MG-63 cells by up-regulating miR-34a

AIM: To investigate the effects of resveratrol on the viability, invasion and autophagy of osteosarcoma MG-63 cells and the regulatory effect of microRNA-34a (miR-34a). METHODS: MG-63 cells were divided into 6 groups:control group and resveratrol treatment groups at doses of 10, 20, 40, 60 and 80 μmol/L. MTT assay, Transwell chamber method and Western blot were used to detect the effects of resveratrol on the viability, invasion ability and expression of autophagy-related proteins in the osteosarcoma cells. The effect of resveratrol at different concentrations on the expression of miR-34a in the osteosarcoma cells was detected by RT-qPCR. The effects of miR-34a mimic and miR-34a mimic negative control (miR-34a NC) transfection on the viability and invasion ability of osteosarcoma cells after treated with resveratrol at different concentrations were analyzed. The effects of miR-34a mimic transfection on autophagy-related proteins LC3-I, LC3-Ⅱ and beclin-1 were determined by Western blot. RESULTS: Compared with control group, resveratrol inhibited the viability and invasion ability of the MG-63 cells and promoted autophagy (P<0.05). Resveratrol up-regulated the expression of miR-34a in the MG-63 cells in a dose-dependent manner (P<0.05). In addition, the ratio of LC3-Ⅱ/LC3-I was increased, and beclin-1 was up-regulated (P<0.05). Co-treatment with miR-34a mimic and resveratrol increased inhibitory effects of resveratrol on the viability and invasion ability and invasion of the MG-63 cells and also promoted autophagy. CONCLUSION: Resveratrol inhibits the viability and invasion of osteosarcoma MG-63 cells and promotes auto-phagy by up-regulating miR-34a expression.     

Molecular mechanism of miR-1246 enhancing radiosensitivity of cervical cancer cells

AIM: To investigate the molecular mechanism of microRNA-1246(miR-1246) enhancing radiosensitivity of cervical cancer cells. METHODS: Cervical cancer lines HeLa, CaSki, C33A and SiHa were transfected with miR-1246 mimic and negative control mimic (NC-mimic) using Lipofectamine 2000 kit, and the expression level of miR-1246 in cervical cancer tissue, normal tissue, cervical cancer cell lines and endometrial epithelium cell line ESC was detected by real-time PCR. The transfected cells were exposed to X-ray radiation. The cell viability and migration rate were measured respectively by MTT assay and Transwell method. The protein levels of γH2AX, ATM, p-ATM and p-p53 were monitored by immunofluorescence and Western blot. RESUITS: Higher miR-1246 level was found in normal tissue and ESC cells, while lower miR-1246 level was found in HeLa, SiHa, C33A and Caski cells and cervical cancer tissues. The expression level of miR-1246 in the cells transfected with miR-1246 mimic was significantly higher than that in the cells transfected with NC-mimic (P<0.05). The cell viability and migration rate of the cervical cancer cells with miR-1246 over-expression were notably lower than those of the cells transfected with NC-mimic (P<0.05) under the same conditions. The results of immunofluorescence indicated that the protein expression level of γH2AX significantly increased in the cervical cancer cells with miR-1246 over-expression exposed to radiation compared with the negative control (P<0.05). The protein expression level of γH2AX was significantly increased in the cervical cancer cells with miR-1246 over-expression, while the protein levels of p-ATM and p-p53 were significantly decreased as compared with the negative control group (P<0.05). CONCLUSION: miR-1246 is highly expressed in normal tissue and normal endometrial epithelial cells, while is low expressed in the cervical cancer tissues or cells. miR-1246 over-expression inhibits growth and migration, and significantly enhances radiosensitivity of cervical cancer cells. The molecular mechanism is possibly related to inhibiting ATM pathway and DNA damage repair.     

IL-17 promotes viability and migration ability of endometrial carcinoma cells by inhibiting miR-195-5p expression

AIM:To investigate the potential mechanism of interleukin-17 (IL-17) promoting the viability, migration and invasion of human endometrial carcinoma cells. METHODS:The expression of IL-17 and microRNA-195-5p (miR-195-5p) in the human endometrial carcinoma and benign uterine lesion samples were detected by RT-qPCR. The expression of miR-195-5p in human endometrial carcinoma HEC-1-B cells after treatment with IL-17 at different concentrations for 48 h was detected by RT-qPCR. The viability, migration and invasion of HEC-1-B cells after treatment with IL-17 at 100 μg/L or transfection of miR-195-5p mimics were detected by MTT assay and Transwell assays. The viability, migration and invasion of HEC-1-B cells after over-expression of miR-195-5p combined with 100 μg/L IL-17 intervention were also observed. RESULTS:The expression of IL-17 was increased while the expression of miR-195-5p was decreased in the human endometrial carcinoma samples (P<0.05). The expression of miR-195-5p in the HEC-1-B cells after treatment with IL-17 at 10,100 and 300 μg/L for 48 h was significantly decreased (P<0.05). The results of MTT assay and Transwell experiments indicated that IL-17 at 100 μg/L enhanced the viability, migration and invasion of HEC-1-B cells, while over-expression of miR-195-5p resulted in the opposite effect. CONCLUSION:Over-expression of miR-195-5p inhibits the enhancing effects of IL-17 on the viability, invasion and migration of HEC-1-B cells.     

miR-181a regulates CSE-induced autophagy dysfunction and releases of pro-inflammatory factors in NR8383 alveolar macrophages

AIM:To investigate the effect of microRNA-181a (miR-181a) on cigarette smoke extract (CSE)-induced autophagy disorder and releases of pro-inflammatory factors in NR8383 rat alveolar macrophages. METHODS:The NR8383 cells were treatment with 5%,10% and 20% CSE. The release levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-8 were measured by ELISA. The level of miR-181a was detected by RT-qPCR. The numbers of autophagosomes were observed by Cyto-ID staining. The expression levels of LC3-Ⅱ, beclin-1 and p62 were determined by Western blot. NR8383 cells were pretreated with autophagy inhibitor 3-methyladenine (3-MA) or autophagy agonist rapamycin (Rapa) before treatment with 20% CSE, and the release levels of TNF-α, IL-6 and IL-8 were measured by ELISA. Furthermore, NR8383 cells were transfected with miR-181a mimic or miR-181a inhibitor before treatment with 20% CSE, and the release levels of TNF-α, IL-6 and IL-8, and the expression of LC3-Ⅱ, beclin-1 and p62 were detected by ELISA and Western blot, respectively. RESULTS:CSE increased release levels of pro-inflammatory factors and autophagy disorder in a concentration-dependent manner in the NR8383 cells (P<0.05). 3-MA increased CSE-induced releases of pro-inflammatory factors. However, Rapa partially reversed CSE-induced releases of pro-inflammatory factors. Additionally, miR-181a mimic inhibited CSE-induced releases of pro-inflammatory factors and promoted autophagy. However, miR-181a inhibitor increased CSE-induced releases of pro-inflammatory factors and autophagy disorder. CONCLUSION:miR-181a regulates CSE-induced releases of pro-inflammatory factor in the NR8383 cells, which may be related to the regulatory role of miR-181a in autophagy disorder.     

杂交水稻制种应用穗萌抑制剂试验初报

[目的]筛选出切实有效的穗萌抑制剂。[方法]通过用4种药剂(乙烯、多效唑、穗芽克、脱落酸),在4个地点(凤台、合肥、巢湖和舒城)进行试验。[结果]4种药剂对抑制穗上发芽都有一定的作用,但在不同地点不同药剂效果不尽一致。这些药剂在2007年使用中对种子发芽率没有影响。[结论]4种药剂中,以脱落酸效果最好,但是其价格昂贵,在杂交水稻制种上无法应用。     

多氯联苯(PCB153)对湛江叉鞭金藻生理生态和超微结构的影响

为研究多氯联苯对海洋微藻的生理生态毒性,以湛江叉鞭金藻(Dicrateria zhanjiangensis)为研究对象,进行7 d的六氯联苯(PCB153)胁迫实验,比较其生长、光合色素含量、谷胱甘肽S-转移酶(GST)、超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量以及藻细胞超微结构的变化情况。结果显示,PCB153胁迫对湛江叉鞭金藻的生长、光合色素含量及抗氧化和解毒相关指标均有显著影响(P<0.05)。随着PCB153浓度的增大,其对湛江叉鞭金藻生长抑制作用不断增大,250 μg/L胁迫组藻细胞在第5天全部死亡。PCB153胁迫后,湛江叉鞭金藻叶绿素a、叶绿素c、类胡萝卜素和总光合色素均显著下降(P<0.05),且随着PCB153浓度的增加,各实验组光合色素含量下降比例增大。PCB153胁迫后,各胁迫组藻细胞MDA含量显著增加;低浓度PCB153 (< 25 μg/L)胁迫显著诱导SOD和GST活性的提高;而高浓度PCB153(> 25 μg/L)胁迫则显著抑制2种酶活性。短期低浓度PCB153胁迫会改变金藻细胞超微结构,使细胞器形态改变、聚缩;高浓度PCB153胁迫则会直接破坏细胞膜的完整性,使细胞破裂,导致细胞自溶死亡。研究表明,PCB153抑制叉鞭金藻的生长和光合色素合成,低浓度PCB153激活抗氧化和解毒系统,提高其自我保护水平,高浓度PCB153加剧脂质过氧化,破坏抗氧化和解毒系统正常功能,导致细胞破裂死亡。     

miR-193b enhances cytotoxicity of doxorubicin by targeting Mcl-1 in breast cancer

AIM: To investigate the effect of microRNA(miR)-193b on doxorubicin therapy in breast cancer in vitro.METHODS: miR-193b level in plasma was detected by real-time PCR in the patients with breast cancer or the healthy controls. MTT assay was performed to measure the inhibitory effect of miR-193b plus doxorubicin on the growth of MDA-MB-231 cells. Bioinformatics, real-time PCR and Western blot were performed to determine whether the expression of Mcl-1 was regulated by miR-193b. Mcl-1 expression vector was constructed, and the role of Mcl-1 vector toward miR-193b plus doxorubicin-induced cytotoxicity in MDA-MB-231 cells was observed by MTT assay.RESULTS: Down-regulation of miR-193b was found in breast cancer patients. The miR-193b plus doxorubicin group showed a higher growth inhibition than cisplation group in MDA-MB-231 cells. The expression of Mcl-1 at both mRNA and protein levels was down-regulated after miR-193b transfection. The growth inhibition of MDA-MB-231 cells treated with miR-193b plus doxorubicin was significantly decreased after the transfection of Mcl-1 expression vector.CONCLUSION: miR-193b sensitizes doxorubicin-induced cytotoxicity by targeting Mcl-1 in breast cancer.     

Anti-atherosclerosis effect of astragaloside is related to miR-33a/ABCA1 signaling

AIM:To study whether astragaloside affects the expression of ATP binding cassette transporter A1 (ABCA1) by regulating miR-33a and promotes the outflow of cholesterol in macrophages. METHODS:In the in vivo experiments, HE staining was used to detect the pathological damage of the cross section of aorta in the mice. The expression of ABCA1 at mRNA and protein levels in mouse aorta was determined by real-time PCR and Western blot. In the in vitro experiments, THP-1 macrophage-derived foam cells were established and then treated with astragaloside-containing serum. Real-time PCR was used to detect the expression of miR-33a. The cells were randomly divided into blank serum group, astragaloside serum group and astragaloside serum+miR-33a mimic group. The expression of ABCA1 at mRNA and protein levels was determined by real-time PCR and Western blot. Oil red O staining and high-performance liquid chromatography were used to detect intracellular lipid content. The method of[³H] incorporation was used to detect intracellular cholesterol outflow. RESULTS:In vivo experiments showed that the blood vessels of the mice in astragaloside group were structurally normal, with neat arrangement, localized small calcified particles, mild lesions, small plaques, reduced foam cells and li-pid, and basically complete elastic plates, indicating that the pathological changes were significantly lighter than those in model group. Compared with model group, the expression of miR-33a in the aorta of the mice in astragaloside group was decreased and the relative expression of ABCA1 at mRNA and protein levels was increased (P<0.05). In vitro experiments showed that astragaloside significantly up-regulated the expression of ABCA1 at mRNA and protein levels, but this effect was inhibited by the transfection of miR-33 mimic without affecting the cell viability. Astragaloside reduced the lipid accumulation in the cells, but this effect was attenuated by miR-33 mimic. Astragaloside reduced intracellular cholesterol accumulation in relation to its promotion of intracellular cholesterol efflux, and the transfection of miR-33a mimic in the cells inhibited cholesterol efflux. CONCLUSION:Astragaloside inhibits the production of miR-33a to increase the expression of ABCA1 and promote the outflow of cholesterol in macrophages. This may be one of the molecular mechanisms of astragaloside in preventing atherosclerosis.