Glucose Stimulation of Pancreatic -cell Lines Induces Expression and Secretion of Dynorphin
Glucose Stimulation of Pancreatic -cell Lines Induces Expression and Secretion of Dynorphin
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通讯作者：
K. Josefsen;Karsten Buschard
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作者：
K. Josefsen;Karsten Buschard
为了研究胰腺细胞对高血糖的适应性反应，采用减法克隆的方法鉴定了葡萄糖刺激诱导的基因。在代表差异表达基因的53个克隆中，20个编码内源性阿片前体prodynorn -phin。小鼠前啡肽的氨基酸序列与大鼠蛋白的氨基酸序列相同，其序列包括阿片肽，其余部分86%相同。刺激MIN6细胞使前啡肽RNA水平增加到生理葡萄糖浓度的20倍以上。在小鼠TC3和大鼠Rinm5F -细胞系中观察到相似的诱导水平。葡萄糖刺激后1小时内，促啡肽RNA的表达增加，4小时后达到最高水平，并在至少24小时内保持升高。通过RIA，发现葡萄糖刺激后MIN6细胞含有并分泌更多的促啡肽a。用KCl、福斯克林或异丁基甲基黄嘌呤处理MIN6细胞强烈诱导前啡肽RNA表达，提示诱导可能与分泌偶联信号通路有关。prodynorphin在若干个自欺欺人的细胞系中的诱导，与先前的自欺欺人细胞合成其他内源性阿片样物质（包括自欺欺人的内啡肽）的证明是一致的，这表明阿片样物质可能在调节自欺欺人细胞分泌中具有潜在的重要作用。T HE´-朗格汉斯胰岛细胞通过精确调节预成形颗粒中胰岛素的释放来调节血清葡萄糖水平。由葡萄糖磷酸化引发的一系列快速电生理事件最终导致了细胞膜的去极化和电压敏感的l型Ca ϩϩ-channels的打开(1)。由此引起的细胞内Ca ϩϩ-ion浓度的增加激活胰岛素颗粒的胞外分泌。在体内和体外，慢性葡萄糖刺激可能会改变细胞对葡萄糖的反应（参考文献2）。在体内，短期暴露于葡萄糖可以增强或抑制胰岛素分泌，以应对随后的刺激(3)。相反，长期的高血糖通常会降低细胞的反应性。有研究表明，后一种被称为葡萄糖毒性的现象，可能与非胰岛素依赖型糖尿病特征的胰岛素细胞葡萄糖反应受损有关（4 - 6）。短期葡萄糖对细胞反应性的影响可能部分通过自分泌因子的作用介导。在静息的细胞中，胰岛素储存在预先形成的颗粒中，这些颗粒也含有染色质粒蛋白A、激素原转化酶2和3 （PC2和PC3）和胰淀素（也称为胰岛淀粉样多肽，IAPP）(7)。经证实，ami -lin对体外灌注的大鼠胰岛分泌胰岛素有直接抑制作用（文献回顾8）。胰抑素，由…
To investigate adaptive responses of pancreatic -cells to hyper-glycemia, genes induced by glucose stimulation were identified by subtraction cloning. Among 53 clones representing differentially expressed genes, 20 encoded the endogenous opioid precursor, prodynor-phin. The amino acid sequence of murine prodynorphin is identical to the rat protein in sequences comprising the opioid peptides and 86% identical in the remainder of the molecule. Stimulation of MIN6 cells increased prodynorphin RNA levels to more than 20-fold in proportion to physiological glucose concentrations. Similar induction levels were observed in murine TC3 and rat Rinm5F -cell lines. Prodynorphin RNA expression increased within 1 h of glucose stimulation, achieved maximal levels by 4 h, and remained elevated for at least 24 h. By using RIA, MIN6 cells were shown to contain and secrete increased amounts of dynorphin-A following glucose stimulation. Treatment of MIN6 cells with KCl, forskolin, or isobutyl-methyl-xanthine strongly induced prodynorphin RNA expression, suggesting that induction may be related to secretion-coupled signaling pathways. The induction of prodynorphin in several -cell lines is consistent with previous demonstrations of -cell synthesis of other endogenous opioids, including -endorphin, and suggests that opioids may have a potentially significant role in regulating -cell secretion. T HE -CELLS of pancreatic islets of Langerhans modulate serum glucose levels through precisely regulated release of insulin from preformed granules. A sequence of rapid electrophysiologic events initiated by glucose phos-phorylation culminates in depolarization of -cell membranes and opening of voltage-sensitive, L-type Ca ϩϩ-channels (1). The resulting increase in intracellular Ca ϩϩ-ion concentration activates exocytosis of insulin granules. Chronic glucose stimulation may alter -cell responses to glucose in vivo and in vitro (reviewed in Ref. 2). In vivo, short-term exposure to glucose can either augment or inhibit insulin secretion in response to subsequent stimulation (3). In contrast, prolonged hyperglycemia typically reduces -cell responsiveness. It has been suggested that the latter phenomenon, referred to as glucose toxicity, may relate to the impaired -cell glucose responses that characterize non-insulin-dependent diabetes mellitus (4 – 6). Short-term glucose effects on -cell responsiveness may be mediated in part through actions of autocrine factors. In resting -cells, insulin is stored in preformed granules that also contain chromogranin A, the prohormone convertases 2 and 3 (PC2 and PC3), and amylin (also known as islet amy-loid polypeptide, IAPP) (7). A direct inhibitory effect of amy-lin on insulin secretion by perfused rat islets in vitro has been demonstrated (reviewed in Ref. 8). Pancreastatin, derived by …