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目的探讨葡萄糖和游离脂肪酸(FFA)联合作用对胰岛β细胞功能与凋亡的影响及病理生理机制。方法Wistar大鼠采用高脂饲料喂养制造胰岛素抵抗模型,取造模成功的肥胖胰岛素抵抗模型大鼠分为4个亚组,生理盐水组(OB-NS组,n=7)、高葡萄糖组(OB-GS组,n=9)、高游离脂肪酸组(OB-FFA组,n=8)和高糖高脂肪酸组(OBFG组,n=9)。另取5只Wistar大鼠采用普通饲料喂养作为正常对照组。采用比色法检测FFA和β-羟丁酸(β-HBA)水平,采用静脉葡萄糖耐量试验(IVGTT)评价胰岛β细胞胰岛素分泌功能,免疫组织化学方法检测β细胞胰岛素储备的能力,并应用原位末端标记法(TUNEL)检测凋亡的β细胞。结果肥胖大鼠葡萄糖输注率(GIR)低于正常对照〔(10.82±1.8mg/kg·min)vs.(25.21±1.7mg/kg·min),P<0.05〕,提示胰岛素抵抗模型成功建立。OB-FG组大鼠糖负荷后胰岛素分泌达峰时间延后且各时点胰岛素水平明显低于NC组和OB-NS组,差异有统计学意义(P<0.05)。糖负荷后5、10、15和20min时,OB-FG组血胰岛素水平也明显较OB-FFA组、OB-GS组降低,胰岛素和血糖比值及胰岛素释放指数也均低于NC组和OB-NS组,差异有统计学意义。免疫组织化学染色示,OB-FG组胰岛素储备在各高脂肥胖亚组中最低,远低于NC组和OB-NS组(P<0.01),OB-GS组和OB-FFA组胰岛素储备与OB-NS组比较也有不同程度降低(P<0.05)。OB-FG组β细胞凋亡率高于NC组和OB-NS组(P<0.01)。结论糖脂联合毒性在引起肥胖大鼠酮体生成显著增加的同时也导致胰岛细胞分泌功能障碍。
Objective To investigate the effect of glucose and free fatty acid (FFA) on the function and apoptosis of pancreatic β cells and its pathophysiological mechanism. Methods Wistar rats were fed with high-fat diet to make insulin resistance model. The obese insulin resistance model rats were divided into 4 subgroups: normal saline group (OB-NS group, n = 7), high glucose group OB-GS group, n = 9), high free fatty acid group (OB-FFA group, n = 8) and high glucose and high fatty acid group (OBFG group, n = 9). Another five Wistar rats were fed with normal diet as normal control group. The levels of FFA and β-hydroxybutyric acid (β-HBA) were measured by colorimetric assay. The insulin secretion of pancreatic β cells was evaluated by IVGTT. The ability of β-cell insulin stores was detected by immunohistochemistry. Apoptotic beta cells were detected by TUNEL. Results The glucose infusion rate (GIR) of obese rats was lower than that of the normal controls [(10.82 ± 1.8mg / kg · min) vs. (25.21 ± 1.7mg / kg · min), P <0.05〕 . The peak time of insulin secretion after glucose load in OB-FG group was delayed and the insulin level at each time point was significantly lower than that in NC group and OB-NS group (P <0.05). At 5, 10, 15 and 20 min after glucose load, the level of insulin in OB-FG group was also significantly lower than that in OB-FFA group and OB-GS group, and the insulin and blood glucose ratio and insulin release index were also lower than those in NC group and OB- NS group, the difference was statistically significant. Immunohistochemical staining showed that insulin reserve in OB-FG group was the lowest among all obesity-prone obesity groups and significantly lower than that in NC group and OB-NS group (P <0.01) OB-NS group compared with varying degrees of reduction (P <0.05). The apoptosis rate of β-cell in OB-FG group was higher than that in NC group and OB-NS group (P <0.01). Conclusion The combination of glycolipid and lipotoxicity induced a significant increase of ketone bodies in obese rats and at the same time resulted in the dysfunction of pancreatic islet cells.