α-Crystallin downregulates the expression of TNF-α and iNOS by activated rat retinal microglia in vitro and in vivo

Purpose: Inhibition of microglial activation has become an important strategy to attenuate neurotoxic damage to the central nervous system. We evaluated the effects of α-crystallin on the production of cytokines in lipopolysaccharides (LPS) and optic nerve injury-activated retinal microglia. Methods: Microglia were collected from retinas of newborn rats, cultured and treated with LPS in vitro. Microglia were also activated by an optic nerve crush in vivo. Pretreatments with and without α-crystallin were performed in cultured cells, and by intravitreal injection in adult rats. Expression of tumor necrosis factor-α (TNF-α), nitric oxide (NO) and inducible NOS synthase (iNOS) were measured by RT-PCR, ELISA, Western blot and the nitrate reductase method. Results: Activated microglia significantly upregulated TNF-α and iNOS mRNA expression and protein production in vitro. An optic nerve crush also increased expression of retinal iNOS and TNF-α protein. Treatment with α-crystallin in vitro and in vivo downregulated their expression. Conclusion: The protective effect of α-crystallin may be due to its effect on microglia via a downregulation in the expression and release of 2 key immune regulatory and inflammatory molecules: TNF-α and iNOS.