Sancho Veronica, Daniele Giuseppe, Lucchesi Daniela, Lupi Roberto, Ciccarone Annamaria, Penno Giuseppe, Bianchi Cristina, Dardano Angela, Miccoli Roberto, Del Prato Stefano
Department of Clinical and Experimental Medicine, Section of Diabetes and Metabolic Diseases, University of Pisa - Cisanello Hospital, Pisa, Italy.
Section of Diabetes and Metabolic Diseases, Azienda Ospedaliero-Universitaria Pisana, Cisanello Hospital, Pisa, Italy.
PLoS One. 2017 Nov 9;12(11):e0187836. doi: 10.1371/journal.pone.0187836. eCollection 2017.
An intra-islet incretin system has been recently suggested to operate through modulation of the expression and activity of proconvertase 1/3 and 2 (PC1/3, PC2) in pancreatic alpha-cell accounting for local release of GLP-1. Little is known, whether this alpha-cell activity can be affected by the metabolic alterations occurring in type 2 diabetes, such as hyperglycemia, hyperlipidemia or hyperglucagonemia.
AlphaTC1/6 cells from a mice pancreatic cell line were incubated in the presence of two glucose (G) concentration (5.5 and 16.7 mM) for 16 h with or without free fatty acid, IL6 or glucagon. GLP-1 secretion was measured by ELISA and expression of PC1/3 and PC2 by RT-PCR and western blot; cell viability was determined by MTT method, Reactive Oxygen Species generation (ROS) by H2DCFDA fluorescence and apoptosis by Annexin staining and Propidium Iodine (PI) fluorescence.
Upon 16.7G incubation, GLP-1 secretion (total and active) was significantly increased in parallel with a significant increment in PC1/3 expression, a slight increase in cell viability and ROS generation and by a decrement in PC2 expression with no change in cell apoptosis. When cells were incubated at 5.5mM glucose with FFA, also an increment in GLP-1 secretion and PC1/3 expression was observed together an increment in ROS generation, a decrement in cell viability, and a modest increment in apoptosis. When incubated with 16.7mM glucose with FFA, the increment in GLP-1 secretion was reduced to basal, accompanied by an increment in apoptosis and ROS generation. This was also observed with IL-6, but in this case, no modification in ROS generation or apoptosis was observed when compared to 16.7mM glucose. The presence of glucagon did not modify any of the parameters studied.
These data suggest that under hyperglycemic, hyperlipidemia or inflammatory conditions, alpha cells can increase expression PC1/3 and activate GLP-1 secretion, which may contribute protecting both alpha and beta-cells from glucose and lipotoxicity, while this effect seems to be lost in the presence of both pathophysiological conditions.
最近有研究表明，胰岛内肠促胰岛素系统通过调节胰腺α细胞中前转换酶1/3和2（PC1/3、PC2）的表达及活性来发挥作用，从而导致胰高血糖素样肽-1（GLP-1）的局部释放。目前对于2型糖尿病中出现的代谢改变，如高血糖、高血脂或高胰高血糖素血症，是否会影响这种α细胞活性，我们所知甚少。
将来自小鼠胰腺细胞系的AlphaTC1/6细胞，在两种葡萄糖（G）浓度（5.5和16.7 mM）存在的情况下孵育16小时，分别添加或不添加游离脂肪酸、白细胞介素6（IL6）或胰高血糖素。通过酶联免疫吸附测定法（ELISA）检测GLP-1分泌，通过逆转录聚合酶链反应（RT-PCR）和蛋白质免疫印迹法检测PC1/3和PC2的表达；采用噻唑蓝（MTT）法测定细胞活力，通过二氯荧光素二乙酸酯（H2DCFDA）荧光检测活性氧（ROS）生成，通过膜联蛋白染色和碘化丙啶（PI）荧光检测细胞凋亡。
在16.7 mM葡萄糖孵育后，GLP-1分泌（总量和活性形式）显著增加，同时PC1/3表达显著增加，细胞活力和ROS生成略有增加，PC2表达减少，细胞凋亡无变化。当细胞在5.5 mM葡萄糖与游离脂肪酸共同孵育时，也观察到GLP-1分泌和PC1/3表达增加，同时ROS生成增加，细胞活力下降，凋亡略有增加。当与16.7 mM葡萄糖和游离脂肪酸共同孵育时，GLP-1分泌增加幅度降至基础水平，同时凋亡和ROS生成增加。白细胞介素6（IL-6）处理时也观察到类似情况，但与16.7 mM葡萄糖处理相比，此时ROS生成或凋亡无变化。胰高血糖素的存在未改变所研究的任何参数。
这些数据表明，在高血糖、高血脂或炎症条件下，α细胞可增加PC1/3表达并激活GLP-1分泌，这可能有助于保护α细胞和β细胞免受葡萄糖和脂毒性影响，而在两种病理生理条件同时存在时，这种作用似乎消失。
