Zhou Yi-Jun, Song Yu-Ling, Zhou Hui, Li Yan
Department of Endocrinology and Metabolism, Fourth Affiliated Hospital, China Medical University, Shenyang 110034, China.
Chin Med Sci J. 2012 Mar;27(1):18-23. doi: 10.1016/s1001-9294(12)60017-0.
To elucidate GPR40/FFA1 and its downstream signaling pathways in regulating insulin secretion.
GPR40/FFA1 expression was detected by immunofluorescence imaging. We employed linoleic acid (LA), a free fatty acid that has a high affinity to the rat GPR40, and examined its effect on cytosolic free calcium concentration ([Ca2+]i) in primary rat beta-cells by Fluo-3 intensity under confocal microscopy recording. Downregulation of GPR40/FFA1 expression by antisense oligonucleotides was performed in pancreatic beta-cells, and insulin secretion was assessed by enzyme-linked immunosorbent assay.
LA acutely stimulated insulin secretion from primary cultured rat pancreatic islets. LA induced significant increase of [Ca2+]i in the presence of 5.6 mmol/L and 11.1 mmol/L glucose, which was reflected by increased Fluo-3 intensity under confocal microscopy recording. LA-stimulated increase in [Ca2+]i and insulin secretion were blocked by inhibition of GPR40/FFA1 expression in beta-cells after GPR40/FFA1-specific antisense treatment. In addition, the inhibition of phospholipase C (PLC) activity by U73122, PLC inhibitor, also markedly inhibited the LA-induced [Ca2+]i increase.
LA activates GPR40/FFA1 and PLC to stimulate Ca2+ release, resulting in an increase in [Ca2+]i and insulin secretion in rat islet beta-cells.
阐明GPR40/FFA1及其下游信号通路在调节胰岛素分泌中的作用。
通过免疫荧光成像检测GPR40/FFA1的表达。我们使用对大鼠GPR40具有高亲和力的游离脂肪酸亚油酸（LA），并在共聚焦显微镜记录下通过Fluo-3强度检测其对原代大鼠β细胞胞质游离钙浓度（[Ca2+]i）的影响。在胰腺β细胞中用反义寡核苷酸下调GPR40/FFA1的表达，并通过酶联免疫吸附测定法评估胰岛素分泌。
LA急性刺激原代培养的大鼠胰岛分泌胰岛素。在5.6 mmol/L和11.1 mmol/L葡萄糖存在的情况下，LA诱导[Ca2+]i显著增加，这在共聚焦显微镜记录下表现为Fluo-3强度增加。在GPR40/FFA1特异性反义处理后，β细胞中GPR40/FFA1表达的抑制阻断了LA刺激的[Ca2+]i增加和胰岛素分泌。此外，PLC抑制剂U73122对磷脂酶C（PLC）活性的抑制也显著抑制了LA诱导的[Ca2+]i增加。
LA激活GPR40/FFA1和PLC以刺激Ca2+释放，导致大鼠胰岛β细胞中[Ca2+]i增加和胰岛素分泌增加。
