胰腺β细胞KCa3.1在2型糖尿病发病中的作用与调节机制

In pancreatic β cells, activation of NF-κB inhibits secretion of insulin but enhances release of proinflammatory chemokine such as CCL2 and CCL20. Interestingly, activation of intermediate-conductance Ca2+-activated K+ channels (KCa3.1) has the similar effect as NF-κB on insulin secretion and chemokine release. However, the relationship between NF-κB and KCa3.1 in the development and progression of type 2 diabetes mellitus (T2DM) remains unknown. Primal study from our group indicated that inhibition of KCa3.1 significantly improved fasting glucose and tolerance of glucose, enhanced secretion of postprandial insulin level and reduced disappear of β cell mass. Furthermore, the release of proinflammatory chemokine induced by high glucose or palmitic acid was effectively suppressed after inhibition of KCa3.1 in β cells. Therefore, we hypothesize that KCa3.1 would cause inflammatory response in diabetic pancreas by regulating chemokine release and insulin secretion in pancreatic β-cells via NF-κB pathway. The present project is designed to evalute the effect of KCa3.1 on pancreatic inflammation and the progression of T2DM in two kinds of animal models: KCa3.1 knockout mice fed with high fat diet and injected with streptozotocin, and the diabetic db/db mouse strains, both of which are commonly used as rodent models of T2DM. We also assess the effect of high glucose or palmitic acid on NF-κB activation, KCa3.1 expression, inflammatory chemokine release and insulin secretion in mouse and rat pancreatic β cells, respectively. The results will elucidate the role and mechanism of KCa3.1 channels in the progression of T2DM, and also provide valuable insights for therapeutic approaches of T2DM.

在胰腺β细胞，NF-κB活化抑制胰岛素分泌而促进趋化因子释放，中电导钙激活K+通道(KCa3.1)激活产生类似作用。但二者在糖尿病发生中的关系未见报道。预实验发现抑制KCa3.1可降低2型糖尿病小鼠空腹血糖，改善葡萄糖耐量，增加餐后胰岛素水平，减缓β细胞消亡；阻断KCa3.1可抑制高糖或软脂酸诱导的β细胞趋化因子的释放。假设：NF-κB活化介导β细胞KCa3.1上调，协同调节炎性细胞因子和胰岛素分泌，促发胰岛炎症和β细胞功能障碍，导致2型糖尿病。拟应用db/db小鼠和高脂饮食加链脲霉素注射建立2型糖尿病模型，观察阻断或敲除KCa3.1对糖尿病发病及胰腺炎症的影响；培养小鼠和大鼠胰腺β细胞，观察高糖或软脂酸对NF-κB信号、KCa3.1表达、胰岛素分泌和炎性细胞因子释放的影响，并研究NF-κB激活对β细胞KCa3.1的调节机制。研究结果将明确KCa3.1在2型糖尿病发病中的作用与调节机制。

2型糖尿病 (type 2 diabetes mellitus) 已逐步发展为人类高患病率和高死亡率的疾病之一。预计到2030年，全世界2型糖尿病患者将达到5.1亿。因此，针对糖尿病发生的分子机制和新型疗法的研究已成为世界性重大研究课题。然而，2型糖尿病的发病机制至今未完全阐明。近年来，慢性炎症对胰脏β细胞功能的损害在2型糖尿病发病中的作用受到重视。其中，炎症过程的主要调节者-核因子κB (Nuclear factor-κB, NF-κB)被认为是介导胰岛炎症和β细胞功能障碍的主要转录因子。钙激活K+通道亚家族中的中电导钙激活K+通道(intermediate-conductance Ca2+-activated K+ channel, KCa3.1)可促使T淋巴细胞、B淋巴细胞和单核-巨噬细胞等释放炎性趋化因子和细胞因子。敲除KCa3.1通道基因能有效改善小鼠葡萄糖耐量。这提示KCa3.1通道有可能成为治疗糖尿病的潜在靶点。本课题实验结果提示：在体预防性给予KCa3.1抑制剂TRAM-34（120 mg/kg）能显著改善葡萄糖耐量水平，增加餐后胰岛素分泌，降低db/db小鼠随机血糖，CCL2, CCL20 和IL-1β的分泌水平，但对胰岛素敏感性和体重无影响。 在db/db小鼠胰腺组织，TRAM-34治疗可减缓2型糖尿病引起的胰腺β细胞消亡、抑制胰岛炎性因子和KCa3.1蛋白的表达以及单核细胞的浸润。在体外培养的小鼠胰岛细胞，高糖（25 mM）或软脂酸（200 μM）刺激可使KCa3.1蛋白表达水平显著上调，CCL2和CCL20的分泌增加，应用TRAM-34 (200 µM) 和NF-κB抑制剂（PDTC, 100 µM）预处理可有效抑制高糖或软脂酸诱导的上述变化。给予IL-1β激活NF-κB的活性可显著上调KCa3.1转录和翻译水平的表达。结论：NF-κB活化介导β细胞KCa3.1上调，协同调节炎性细胞因子和胰岛素的分泌，促发胰岛炎症和β细胞功能障碍，导致2型糖尿病；KCa3.1有望成为控制2型糖尿病的治疗靶点