色素上皮衍生因子对高糖刺激下大鼠视网膜Müller细胞形态及活性的影响

Effects of PEDF on morphology and activity of rat retinal Müller cells cultured with high glucose

目的检测色素上皮衍生因子(PEDF)对高糖刺激下体外培养SD大鼠视网膜Müller细胞形态及活性的影响,探讨PEDF对Müller细胞的保护作用。方法实验分为对照组、高糖模型(HG)组、PEDF干预高糖模型(HG+PEDF)组。采用反复胰蛋白酶消化法培养纯化SD大鼠视网膜Müller细胞,免疫荧光双标鉴定纯度,应用倒置相差显微镜、透射电子显微镜、CCK-8试剂盒、SOD试剂盒及MDA试剂盒检测各组Müller细胞形态与活性的改变及超氧化物歧化酶(SOD)活力与丙二醛(MDA)含量的变化。结果对照组、HG组、PEDF+HG组细胞吸光度分别为0.67±0.03、0.54±0.03、0.63±0.03,SOD活力分别为(196.69±22.15)、(153.42±15.54)、(180.32±16.47)U/mg prot,M DA含量分别为(2.11±0.43)、(2.86±0.33)、(2.40±0.28)nmol/mg prot。与对照组相比,HG组Müller细胞形态发生明显改变,细胞活性及SOD活力降低(P<0.01),M DA含量升高(P<0.01);与HG组相比,PEDF+HG组Müller细胞形态改变较小,细胞活性升高(P<0.01),SOD活力升高(P<0.05),MDA含量降低(P<0.05)。结论外源给予PEDF可显著减轻高糖所致的Müller细胞形态、活性的改变及细胞损伤,对Müller细胞具有显著的保护作用。

Objective To investigate the effects of pigment epithelium-derived factor （PEDF） on the morphology and activity of high glucose-stimulated rat retinal muller cells in vitro, and explore the protective mechanism of PEDF on muller cells. Methods The rat retinal muller cells were divided into control group, high glucose （HG） group and high glucose coordinated PEDF （ HG ＋ PEDF） group. Retinal muller cells were cultured and purified by repeated pancreatic enzyme-degisting method. The purity of retinal muller cells was identified by double staining immunofluorescence tech-niques. The morphology was identified by inverted phase contrast microscope and transmission electron microscopy. Superoxide dismutase （SOD） activity and malondialdehyde （MDA） level were studied with CCK-8 kit, SOD kit and MDA kit in a quantitative way. Results OD values of control group, HG group and PEDF ＋ HG group were 0.67±0.03, 0.54±0.03 and 0.63±0.03, respectively. SOD activity of these groups were （ 196.69±22.15 ）, （ 153.42±15.54）, and （ 180.32±16.47 ） U/mg prot. MDA level of these groups were （ 2.11±0.43 ）, （2.86±0.33 ）, and （2.40±0.28） nmol/mg prot. When retinal muller ceils were cultured with high glucose, cells morphology changed significantly, cells activity and SOD activity decreased （P 〈 0.01 ）, while MDA level obviously increased （P 〈 0.01 ）.When PEDF were added, the change of cells morphology reduced, cells activity and SOD activity increased obviously （ P 〈 0.05 ） , while MDA level decreased （ P 〈 0.05 ）. Conclusion High glucose can damage the morphology and ac- tivity of retinal mtiller cells. PEDF can protect retinal muller cells from the damage of glucose.