摘要
目的 :探讨过氧化物酶体增殖物激活受体γ(peroxisome proliferator-activated receptorγ,PPARγ)激动剂匹格列酮(pioglitazone,Pio)对Müller细胞活化及炎症因子分泌的影响。方法:第2代原代培养的大鼠Müller细胞鉴定后分为6组,对照组:不加任何药物;脂多糖(lipopolysaccharides,LPS)组:加入1μg/m L LPS;Pio组(共3组):分别加入0.1、1.0、10.0μmol/L Pio处理后再加入1μg/m L LPS;拮抗剂组:加入1.0μmol/L Pio及1μmol/L GW9662(PPARγ拮抗剂)处理后再加入1μg/m L LPS。将各组Müller细胞分别与视网膜神经节细胞(retinal ganglion cells,RGCs)(RGC-5细胞株)共培养,流式细胞术检测RGC-5细胞的凋亡情况;免疫荧光及Western Blot测定Müller细胞中胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)及核因子(nuclear factorκB,NF-κB)P65的表达变化;酶联免疫吸附法(enzyme-linked immunoabsorbent assay,ELISA)检测细胞上清液中炎症因子的分泌。结果 :(1)LPS刺激后Müller细胞中GFAP表达较对照组明显上升(P<0.05),Pio组GFAP的表达较LPS组均有下降(P<0.05)。(2)Pio组PPARγ蛋白表达较LPS组明显增加(P<0.05),拮抗剂组PPARγ蛋白表达较Pio组显著降低(P<0.05)。(3)Pio组NF-κB P65的表达较LPS组明显下降(P<0.05),而拮抗剂组较Pio组明显上升(P<0.05)。(4)Pio组肿瘤坏死因子α(tumor necrosis factorα,TNF-α)、白细胞介素6(interleukin 6,IL-6)的水平较LPS组明显下降(P<0.05)。拮抗剂组TNF-α、IL-6较Pio组有明显上升(P<0.05)。(5)不同处理组的Müller细胞与RGC-5细胞共培养,Pio组RGC-5细胞凋亡较LPS组明显降低(P>0.05),拮抗剂组较Pio组明显上升(P>0.05)。结论 :PPARγ激动剂Pio能抑制Müller细胞的激活、抑制NF-κB信号通路及炎症因子释放,从而保护视网膜神经元。
Objective： To investigate the effects of peroxisome proliferator-activated receptor γ（PPARγ） agonist pioglitazone（Pio） on Müller cell activation and secretion of inflammatory cytokines. Methods： Primary cultured rat Müller cells were identified and divided into six groups. The control： Müller cells without stimulation of lipopolysaccharides（LPS）; the LPS group： Müller cells treated with 1 μg/m L LPS; the Pio groups： Müller cells treated with different concentrations of Pio（0.1, 1.0,10.0 μmol/m L） and then treated with 1 μg/m L LPS; the antagonists group： Müller cells treated with 1.0 μmol/L Pio and 1 μmol/L GW9662 and then treated with 1 μg/m L LPS. The different treated Müller cells were co-cultured with RGC-5 cells and the apoptosis of RGC-5 was detected by flow cytometry. Immunofluorescence and Western Blot analysis were performed to detect the levels of glial fibrillary acidic protein（GFAP） and nuclear factor κB（NF-κB） P65. Inflammatory cytokines in the cell supernatants were detected by enzyme-linked immunoabsorbent assay（ELISA）. Results：（1）The expression of GFAP in LPS group was increased（P〈0.05）. Pio pre-treatment significantly decreased the increasing of GFAP induced by LPS（ P〈0.05）.（2）PPARγ protein expression was significantly increased in Pio group（ P〈0.05）, while adding PPARγ antagonist GW9662 decreased PPARγ protein expression（P〈0.05）.（3）The expression of NF-κB P65 was significantly decreased in Pio grouops and increased again when GW9662 was used（P〈0.05）.（4）The amounts of tumor necrosis factor α（TNF-α） and interleukin 6（IL-6） in Pio groups were decreased compared with LPS group（ P〈0.05）, but their amounts in the antagonists group were increased compared with Pio groups（P〈0.05）.（5）The result of flow cytometry suggested that Pio significantly decreased the apoptosis of RGC-5 cells in Pio treated groups. Conclusions： The mechanisms for protective effects of PPARγ agonist Pio on retinal neural cells may be associated with decreasing the activation of Müller cells, inhibition of NF-κB signaling pathway and the inflammatory response.
作者
顾宏卫
张少华
张俊芳
秦柏
胡楠
GU Hongwei;ZHANG Shaohua;ZHANG Junfang;QIN Bai;HU Nan(Department of Ophthalmology,the Affiliated Hospital of Nantong University,Nantong 226001)
出处
《南通大学学报（医学版）》
2018年第3期187-192,共6页
Journal of Nantong University(Medical sciences)
基金
南通市科技计划项目(MS22015002)
作者简介
顾宏卫，男，汉族，生于1978年11月，江苏省南通市人，硕士，研究方向眼表疾病。;【通信作者】胡楠，电话：13515209931，E-mail：hunaneye@hotmail.com
