Expression and localization of GPR109A (PUMA-G/HM74A) mRNA and protein in mammalian retinal pigment epithelium
Expression and localization of GPR109A (PUMA-G/HM74A) mRNA and protein in mammalian retinal pigment epithelium
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发表时间：
2009-02
期刊：
影响因子：
2.2
通讯作者：
Pamela M. Martin;S. Ananth;G. Cresci;P. Roon;Sylvia B. Smith;V. Ganapathy
中科院分区：
文献类型：
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作者：
Pamela M. Martin;S. Ananth;G. Cresci;P. Roon;Sylvia B. Smith;V. Ganapathy
目的GPR 109 A是烟酸的G蛋白偶联受体。β-羟基丁酸（β-HB）是该受体的生理配体。β-HB是血液循环中的主要酮体，在各种生理和病理条件下是神经元（包括视网膜神经元）的重要能量来源。将GPR 109 A鉴定为β-HB的受体表明该代谢产物具有其他迄今未知的功能。糖尿病患者的β-HB循环水平升高。由于视网膜病变是与糖尿病相关的严重并发症，我们研究了视网膜和不同视网膜细胞类型中的GPR 109 A表达，以确定该受体是否可能在糖尿病视网膜病变的病理生理学中发挥作用。方法应用RT-PCR、荧光原位杂交和免疫荧光技术检测GPR 109 A在小鼠视网膜及3种转化细胞系ARPE-19（RPE）、RGC-5（ganglion）和rMC-1（Müller）中的表达。通过cAMP测定证实了烟酸和β-HB在ARPE-19细胞中激活GPR 109 A。结果利用小鼠视网膜组织进行的研究表明，GPR 109 A在视网膜中表达，其表达仅限于RPE，在那里它差异极化到基底外侧膜。这些结果用细胞系证实，其证明GPR 109 A在ARPE-19中表达，但在rMC-1和RGC-5细胞中不表达。小鼠RPE的原代培养物也显示GPR 109 A的稳健表达。cAMP测定表明在RPE中表达的GPR 109 A是功能性的。结论本研究首次报道了GPR 109 A在视网膜中的表达。GPR 109 A在视网膜色素上皮基底外侧膜的特异性表达，与血液中β-HB的通路密切相关，可能在糖尿病视网膜病变中具有重要的生物学意义。
Purpose GPR109A has been identified as a G-protein-coupled receptor for niacin. β-hydroxybutyrate (β-HB) is a physiologic ligand for the receptor. β-HB, the predominate ketone body in circulation, is an important energy source for neurons, including retinal neurons, under various physiologic and pathologic conditions. The identification of GPR109A as the receptor for β-HB suggests additional, hitherto unknown, functions for this metabolite. The circulating levels of β-HB increase in diabetes. Since retinopathy is a serious complication associated with diabetes, we investigated GPR109A expression in retina and in different retinal cell types to determine if the receptor may have a role in the pathophysiology of diabetic retinopathy. Methods RT–PCR, fluorescent in situ hybridization, and immunofluorescent techniques were used to analyze GPR109A expression in mouse retina and in three transformed retinal cell lines: ARPE-19 (RPE), RGC-5 (ganglion), and rMC-1 (Müller). Activation of GPR109A by niacin and β-HB was demonstrated in ARPE-19 cells by cAMP assay. Results Studies conducted using mouse retinal tissues demonstrated that GPR109A is expressed in retina with its expression restricted to RPE, where it differentially polarizes to the basolateral membrane. These results were confirmed with cell lines, which demonstrated GPR109A expression in ARPE-19, but not in rMC-1 and RGC-5 cells. Primary cultures of mouse RPE also showed robust expression of GPR109A. cAMP assay demonstrated that GPR109A expressed in RPE is functional. Conclusions These data represent the first report on GPR109A expression in retina. The exclusive expression of GPR109A in RPE basolateral membrane, which has access to β-HB in blood, may have biologic importance in diabetic retinopathy.