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71.
ROCK promotes high glucose-induced cardiomyocyte apoptosis by inhi-biting PI3K/Akt signaling pathway
AIMTo investigate whether Rho-associated coiled-coil kinase (ROCK) is involved in high glucose-induced apoptosis of primary cardiomyocytes by regulating PI3K/Akt signaling pathway. METHODSPrimary Wistar rat cardiomyocytes were cultured and identified by α-sarcomeric actin (α-SCA) immunohistochemistry. Cardiomyocytes were treated with 5.5, 33 and 40 mmol/L glucose for 48 h. The cell viability was measured by MTT assay, and the mRNA expression of ROCK1 and ROCK2 in the cardiomyocytes was detected by RT-qPCR. Flow cytometry was used to analyze the apoptosis of the cardiomyocytes. The protein levels of ROCK1, ROCK2, cleaved caspase-3, Bcl-2, PI3K, Akt and p-Akt were determined by Western blot. In order to confirm the regulatory effect of ROCKs on PI3K/Akt signaling pathway, the cells were divided into control group (5.5 mmol/L glucose), high glucose group (33 mmol/L glucose) and high glucose+Y27632 (ROCK inhibitor) group. Western blot was used to detect the protein levels of ROCK1, ROCK2, PI3K, Akt and p-Akt. RESULTSAfter 48 h of high glucose exposure, the values of relative cell viability in 33 and 40 mmol/L glucose groups were (79.71±2.43)% and (68.41±7.49)%, respectively, both of which were significantly decreased compared with normal control group (P <0.05). After 48 h of high glucose exposure, the relative mRNA levels of ROCK1 and ROCK2 in 33 and 40 mmol/L glucose groups were significantly increased compared with normal control group (P <0.05). Compared with normal control group, the apoptotic rate in 33 and 40 mmol/L glucose groups was increased significantly (P <0.05). Compared with normal control group, the protein expression of ROCK1, ROCK2 and cleaved caspase-3 in 33 and 40 mmol/L glucose groups was increased (P <0.05), while the protein expression of Bcl-2 was decreased (P <0.05). No significant difference in the protein levels of PI3K and Akt among the 3 groups was observed, while the protein level of p-Akt in 33 and 40 mmol/L glucose groups was decreased compared with normal control group (P <0.05). Compared with high glucose group, the expression of ROCK1 and ROCK2 was decreased in high glucose+Y27632 group. No significant difference in the protein levels of PI3K and Akt among the 3 groups was observed. Compared with normal control group, the protein level of p-Akt in high glucose group was decreased, and the protein level of p-Akt in high glucose+Y27632 group was increased significantly compared with high glucose group. CONCLUSION Under high glucose environment, ROCK may reduce the level of p-Akt by inhibiting the PI3K/Akt signaling pathway, thus promoting the apoptosis of cardiomyocytes. 相似文献
72.
AIM: To explore the effect of inositol 1, 4, 5-trisposphate receptor (IP3R) in luteinizing hormone-epidermal growth factor receptor (LH-EGFR)-induced oocyte meiotic resumption. METHODS: Models of mouse cumulus-oocyte complexs (COCs) culture and follicle culture in vitro were generated to study the effects of 2-aminoethyl diphenyl borate (2-APB) and heparin (IP3R specific inhibitors) on LH/EGF-induced oocyte meiotic resumption and EGF-induced cumulus cell expansion. Real-time PCR was used to detect the mRNA expression of cumulus expansion-related factors. The changes of the intracellular calcium level were monitored using Fluo 3-AM, and the cGMP level was measured by ELISA. RESULTS: The inhibitors of IP3R, 2-APB and heparin, dramatically reversed EGF-induced oocyte maturation (P <0.05) and decreased cGMP levels in COCs (P <0.05). In addition, 2-APB and heparin reversed EGF-induced cumulus expansion, and significantly inhibited EGF-induced cumulus expansion-related factor expression (P <0.05). The activation of IP3R increased intracellular calcium level, and the study found that 2-APB and heparin dramatically reversed EGF-induced elevation of calcium level in cumulus cells (P <0.05). Follicular culture in vitro showed that 2-APB and heparin significantly reversed the LH-induced oocyte maturation (P <0.05). CONCLUSION: LH-EGFR signaling pathway increases calcium level in cumulus cells through IP3R, resulting in meiotic resumption. 相似文献
73.
AIM: To observe the effect of histone deacetylase inhibitor (HDACi) Belinostat on the viability of osteosarcoma cells and to study the underlying mechanism. METHODS: Osteosarcoma cell lines SAOS-2 and U2OS were incubated with Belinostat at different concentrations in vitro . The viability of the cells was measured by MTT assay. The activity of caspase-3/-7 and the DNA fragmentation were detected by fluorescence probe and ELISA, respectively. Western blot was used to detect the levels of histone acetylation, expression of PTEN, caspase-3, Bcl-xL and Akt, and phosphorylation of glycogen synthetase kinase 3β (GSK-3β) and Akt. Finally, the cells were incubated with Belinostat and doxorubicin at different concentrations, and then the combination index (CI) was calculated by MTT. RESULTS: Belinostat at 0.5, 1, 2.5 and 5 μmol/L inhibited the viability of U2OS cells and SAOS-2 cells in a dose-dependent manner, induced DNA fragmentation, enhanced caspase-3/-7 activity, and promoted the activation of caspase-3. At the same time, in the SAOS-2 cells, the expression of Bcl-xL was reduced, and the acetylation of histones H3 and H4 was increased. The results of Western blot showed that phosphorylation levels of Akt and GSK-3β in U2OS cells and SAOS-2 cells were decreased significantly after treatment with Belinostat (P <0.05). MTT results showed that combination of Belinostat and doxorubicin further reduced the viability of U2OS and SAOS-2 cells (CI<1). CONCLUSION: Belinostat inhibits the viability of osteosarcoma cells treated with doxorubicin, and the mechanism may be related to the inhibition of Akt signaling pathway. 相似文献
74.
由于人口不断增长,人们要快速得到高产高质粮食的要求迫切,大量使用化肥,导致了有害物质残留,土壤或水污染,土壤板结或某些营养元素相对匮乏等一系列环境问题。丛枝菌根(Arbuscular mycorrhiza, AM)是土壤内常见的共生结构,由AM真菌(AMF)与土壤根系形成。已有研究表明其可通过分泌代谢物,增大根系与土壤接触面积,调节某些土壤元素存在形式等多种途径,影响植物对土壤元素的吸收转运。硫是维持植物生长发育的必需元素之一,可由于植物对S的需要并不如N,P,K大量,现代农业在对土壤进行施肥过程中往往将其忽略,因此土壤缺S正逐渐成为中国农业发展的限制因素。为了解决以上问题,本文将主要对AMF影响植物吸收土壤元素的途径及生理机制进行总结分析。并根据其作用方式特点进一步分析AM共生对植物吸收转运硫素的影响,指出AMF作为生物化肥的可行性,以期为解决现代化肥的替代问题以及土壤缺硫问题提供新的思路。 相似文献
75.
花粉管通道法在玉米自交系改良中的应用 总被引:5,自引:0,他引:5
就几年来花粉管通道法在玉米自交系改良中的应用情况加以概括,提出了导入方法的适宜细则,对后代变异情况加以论述,并对今后该项工作的开展提出几点建议. 相似文献
76.
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78.
细胞自噬是哺乳动物细胞物质代谢的一个重要机制,与细胞凋亡共同参与卵巢卵泡的发育和闭锁,并发挥重要的作用。近年研究发现,磷脂酰肌醇3-激酶/蛋白激酶B(phosphatidylinositol 3-kinase/protein kinase B,PI3K/AKT)信号通路参与卵巢疾病的发生。PI3K和AKT的过度激活可使原始卵泡过早发育以及卵泡过快凋亡,卵巢颗粒细胞作为卵泡发育重要的支持细胞,其功能的减退或凋亡很可能引发一系列女性内分泌方面的疾病。FOXO3a转录因子是PI3K/AKT信号通路下游的重要靶蛋白之一,参与抗增殖和凋亡。本文就关于卵巢颗粒细胞自噬与PI3K/AKT/FOXO3a信号通路的相关进展加以综述。 相似文献
79.
80.
【目的】克隆鸭维甲酸诱导基因I(retinoic acid inducible gene I,RIG-I),分析其不同结构域的功能。【方法】根据GenBank上公布的鸭RIG-I序列设计引物,利用RT-PCR克隆鸭RIG-I基因CDS(coding sequence)区,根据保守结构域预测结果,构建携带6*his组氨酸标签的不同结构域缺失突变体的真核表载体(RIG-I-Full、RIG-I-N和RIG-I-C),转染鸡胚成纤维细胞DF1,经RT-PCR、间接免疫荧光方法鉴定重组质粒在细胞中转录与表达;同时,利用RT-qPCR检测RLR抗病毒信号通路中的IFN-β、Mx1和PKR等下游基因的表达变化。【结果】鸭RIG-I基因CDS区全长2 802 bp,共编码933个氨基酸;不同结构域缺失突变体的真核表载体转染DF1细胞后,重组蛋白均在DF1细胞中表达;RT-qPCR结果显示,N端能显著激活RLR通路上IFN-β、Mx1及PKR基因的表达上调。【结论】 duRIG-I及不同区段均能在DF1细胞中表达,其中N端在调节RLR抗病毒信号通路下游基因的表达过程中发挥了重要作用。 相似文献