TLR-4/MyD88通路激活和小胶质细胞极化在氧化低密度脂蛋白导致的小鼠视网膜损伤中的作用

目的 探讨氧化低密度脂蛋白(ox-LDL)导致的视网膜损伤是否与Toll样受体-4(TLR-4)/MyD88通路激活和活化小胶质细胞使其极化为M1表型相关。方法 雄性C57BL/6小鼠分为两组，实验组小鼠视网膜下注射1μL ox-LDL,对照组小鼠注射等体积的磷酸盐缓冲液(PBS);2周后，采用免疫荧光染色检测视网膜小胶质细胞的离子钙结合适配器分子1(Iba-1)、iNOS的表达，TUNEL染色检测光感受器细胞凋亡，OCT检测视网膜外核层(ONL)和内核层(INL)厚度，ERG检测视网膜功能，qPCR检测视网膜Iba-1、TLR-4和MyD88的mRNA和蛋白表达。体外实验使用ARPE-19细胞，ox-LDL组细胞用100 mg·L^-1 ox-LDL处理24 h, PBS组细胞使用等体积的PBS处理24 h; TUNEL染色检测细胞凋亡，Western blot和qPCR检测细胞TLR-4和MyD88的mRNA和蛋白表达。结果 qPCR和Western blot检测结果均显示，与对照组相比，实验组小鼠视网膜中TLR-4、MyD88、Iba-1的mRNA和蛋白表达水...

Effect of Toll-like receptor-4/myeloid differentiation factor 88 pathway activation and microglial polarization on retinal degeneration induced by oxidized low-density lipoprotein in mice

Objective　To investigate whether retinal degeneration caused by oxidized low-density lipoprotein (ox-LDL) is associated with the activation of Toll-like receptor-4/myeloid differentiation factor 88 (TLR-4/MyD88) pathway and the polarization of microglia to M1 phenotype. Methods　Male C57BL/6 mice were subretinally injected with 1 μL of ox-LDL in the experimental group and an equal volume of PBS in the control group. Two weeks later, the expression levels of ionized calcium-binding adaptor molecule 1 (Iba-1) and inducible nitric oxide synthase (iNOS) in retinal microglia were detected by immunofluorescence. Photoreceptor cell apoptosis was assessed by TUNEL staining. The thickness of the outer nuclear layer (ONL) and inner nuclear layer (INL) was measured by optical coherence tomography (OCT). Retinal function was evaluated by electroretinogram (ERG). The mRNA and protein levels of Iba-1, TLR-4 and MyD88 were detected by quantitative polymerase chain reaction (qPCR). In vitro, ARPE-19 cells were treated with 100 mg·L^-1 ox-LDL for 24 hours in the ox-LDL group and with 100 mg·L^-1 PBS for 24 hours in the PBS group. TUNEL staining was used to detect apoptosis, and Western blot and qPCR to detect mRNA and protein levels of TLR-4 and MyD88. Results　qPCR and Western blot results showed that compared with the control group, the mRNA and protein levels of TLR-4, MyD88 and Iba-1 in the experimental group were up-regulated (all P<0.05), and compared with the PBS group, the mRNA and protein levels of TLR-4 and MyD88 in the ox-LDL group were also up-regulated (all P<0.01). Immunofluorescence results showed that the number of Iba-1 and iNOS positive cells in the experimental group was significantly increased compared with the control group. TUNEL staining results showed that the number of TUNEL-positive cells in the experimental group was increased compared with the control group, and the number of TUNEL-positive cells in the ox-LDL group was also increased compared with the PBS group. ERG results showed that the amplitudes of a and b waves in the experimental group were decreased compared with the control group (both P<0.001). OCT results showed that the ONL/INL ratio in the experimental group was decreased compared with the control group (P<0.01). Conclusion　ox-LDL causes retinal damage, which may be related to the activation of the TLR-4/MyD88 pathway and the polarization of microglia to the M1 phenotype.