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目的:观察高浓度胰淀素对胰岛细胞内活性氧(ROS)生成的影响。方法:以体外培养胰岛单层细胞为模型,应用ROS特异性荧光染料2′,7′-二氯二氢荧光素双醋酸盐(H_2 DCF)进行标记,以粘附细胞仪570型,测定10μmol/L胰淀素作用2h后胰岛细胞内DCF荧光强度变化。结果:10μmol/L胰淀素作用2h,胰岛细胞内DCF荧光强度在16.7mmol/L葡萄糖刺激后,迅速急骤升高进入平台期,升高速度为28×10~(-3)/s,而正常细胞DCF荧光强度无明显变化。光镜下可见部分细胞胞体缩小,胞质发生空泡变性,少数细胞核固缩、裂解,胞膜突起。结论:10μmol/L胰淀素作用后的胰岛细胞内ROS生成增多,细胞形态受损,这些活性氧基的生成可以反映胰淀素对胰岛细胞损害程度,推测活性氧可作为胰岛细胞毒性损伤指标之一。
Objective: To observe the effect of high concentration amylin on reactive oxygen species (ROS) production in islet cells. Methods: The monolayer cells were cultured in vitro and labeled with ROS specific fluorescent dye 2 ’, 7’-dichlorodihydrofluorescein diacetate (H 2 DCF) Fluorescence intensity changes of DCF in islet cells after treated with 10μmol / L amylin for 2h. Results: After treated with 10μmol / L amylin for 2h, the fluorescence intensity of DCF in islet cells rapidly increased rapidly to plateau after the stimulation with 16.7mmol / L glucose, the rate of increase was 28 × 10 -3 / s, DCF fluorescence intensity of normal cells showed no significant change. Light microscope shows that part of the cell body shrinks, vacuolar degeneration of the cytoplasm, a small number of nuclear pyknosis, lysis, membrane protrusion. CONCLUSION: The production of ROS in islet cells is increased after 10μmol / L amylin treatment, and the cell morphology is impaired. The generation of these reactive oxygen species can reflect the degree of damage to pancreatic islet cells induced by amylin, suggesting that ROS can be used as an indicator of cytotoxicity of islet cells one.