嘌呤受体在视网膜色素上皮细胞功能的作用

发布时间：2018-06-03 12:37

本文选题：视网膜色素上皮细胞 + SK通道　；参考：《延边大学》2012年硕士论文

【摘要】：目的：观察并了解嘌呤受体在视网膜色素上皮细胞功能的作用。方法：应用FURA/2荧光寿命成像显微术检测ARPE-19细胞内的钙离子浓度。并应用RT-PCR实验测定BK,SK,IK通道的存在,然后使用内向外式膜片实验进一步鉴定BK,SK,IK通道的存在及电流密度和开放概率。最后,通过使用一种网格蛋白或小窝蛋白(caveolin-1)为介质的运输机制,检测ATP是否通过Gaq减少了细胞表面的Kir4.1丰富度,并采用western blot蛋白印记分析,来分析小窝蛋白和网格蛋白重链。结果：实验结果表明,ATP的应用能够通过激活嘌呤受体增加RPE细胞内的钙离子浓度。增加的细胞内钙离子浓度能够打开钙离子激活钾离子通道。许多组织中的SK通道是在P2Y受体与SK通道和P2Y受体的嘌呤刺激下激活的。P2Y受体与SK通道激活的偶联,是通过蛋白质依赖性磷脂酶c的激活和IP3生产进行的。IP3受体操纵储备物引起的局部钙离子释放看来是SK的主要刺激。在这项研究里,能受体的激活不仅能引起BK通道开放概率的增加,还会引起SK通道开放概率的增加。SK通道的激活导致向外电流,降低细胞膜兴奋性。在此我们还得出,通过一种小窝蛋白内吞作用参与的机制,长期在ATP中培育会导致内向整流钾离子通道的电流密度降低。结论：无论是对钙离子激活的钾通道,还是内向整流钾离子钾离子通道,ATP都扮演一个很重要的角色。嘌呤激活可以增加SK和BK通道的开放概率,并能通过一种小窝蛋白内吞作用参与的机制降低内向整流钾离子通道的电流密度。
[Abstract]:Objective: to investigate the function of purine receptor in retinal pigment epithelium (RPE) cells. Methods: calcium concentration in ARPE-19 cells was detected by FURA/2 fluorescence lifetime imaging microscopy. The existence of IK channel was determined by RT-PCR experiment, and the existence, current density and open probability of IK channel were further identified by internal and external diaphragm experiments. Finally, by using a transport mechanism of griddle protein or nest protein caveolin-1, we detected whether ATP reduced the abundance of Kir4.1 on cell surface through Gaq, and analyzed the heavy chain of nest protein and griddle protein by western blot protein imprinting. Results: the results showed that ATP could increase calcium concentration in RPE cells by activating purine receptors. Increased intracellular calcium concentration can open the calcium-activated potassium channel. SK channels in many tissues are coupled with SK channels activated by P2Y receptors and by purine stimulation of SK channels and P2Y receptors. The local calcium release induced by the manipulation of the. IP3 receptor reserve through the activation of protein-dependent phospholipase c and the production of IP3 appears to be the main stimulation of SK. In this study, activation of energy receptors not only increased the opening probability of BK channels, but also increased the opening probability of SK channels. The activation of SK channels led to outward currents and decreased membrane excitability. We also found that long-term cultivation in ATP resulted in a decrease in the current density of inward rectifier potassium channels through a mechanism involving endocytosis of fossa proteins. Conclusion: both calcium-activated potassium channel and inward rectifier potassium channel ATP play an important role. Purine activation can increase the opening probability of SK and BK channels, and decrease the current density of inward rectifier potassium channels through a mechanism involved in the endocytosis of nest proteins.
【学位授予单位】：延边大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R774.1

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