Effect of α-crystallin on lipopolysaccharide-induced retinal microglia and expression of TNF-α

Objective: As one of lentogenic factors, alpha-crystallin can promote axon regeneration after optic nerve injury. The microenvironment around retinal ganglion cells is also important. Present study was to investigate the effect of α-crystallin on the activity of rat retinal microglia and the expression of stimulated TNF-α with lipopolysaccharides (LPS). Methods: Retinal microglia cells were obtained from the newborn Long Evans rat (< 3 days) and were premarily cultivated and passaged. The 3rd generation of cells were collected and identified using CD11b with immunofluorescence and flow cytometry (FCM). 10^-6 g/L LPS was added in cultured microglia cells in LPS group, and 10^-4 g/L alpha-crystallin and 10^-6 g/L LPS was used in alpha-crystallin + LPS group. Two groups of microglia cells were detected by MTT, and expression of TNF-α mRNA and protein was assessed with ELISA kit arid RT-PGR. Results: Cultured microglia cells showed red fluorescence in cell membrane for CD11b. Purity rate of cultued microglia cells was 95.8% and 91.4% by cell immunochemistry identification and FCM of CD11b, respectively. After pretreatment with alpha-crystailin (10^-4 g/L), the total number of microglia was 1.91 ± 0.08 (OD value), indicating a obvious decrease in comparison with LPS group (2.43 ± 0.06) (t = 51.38, P = 0.001), and the expression of TNF-α mRNA and protein was significantly inhibited in alpha-crystallin + LPS group compared with LPS group (0.64 ± 0.05, 407.5 ± 50 versus 0.82 ± 0.07, 603.7 ± 78 respectively) (t = 5.74, P = 0.029; P = 45.51, P = 0.001). Conclusion: Alpha-crystallin inhibits the proliferation of microglia and expression of TNF-α by blocking the activity of microglia induced by LPS. It suggests that alpha-crystallin may have the protective effect on retinal ganglion cell.