Identification of sequential events and factors associated with microglial activation, migration, and cytotoxicity in retinal degeneration in rd mice

PURPOSE. To elucidate the role of activated microglia in the photoreceptor apoptosis of rd mice by identifying sequential events and factors associated with microglial activation, migration, and cytotoxicity during retinal degeneration. METHODS. Photoreceptor apoptosis in rd mice at postnatal days ( P) 8, 10, 12, 14, 16, and 18 was detected by terminal dUTP transferase nick end labeling ( TUNEL). Retinal microglia were identified by CD11b antibody. Expression of chemokine mRNA, including monocyte chemoattractant protein (MCP)-1, MCP-3, macrophage inflammatory protein (MIP)-1 alpha, MIP-1 beta, regulated on activation normal T-cell expressed and secreted (RANTES), interferon-gamma-inducible 10-kDa protein (IP-10), and fractalkine in the retina were examined by reverse transcription-polymerase chain reaction (RT-PCR) assay. Production of tumor necrosis factor (TNF)-alpha in the dystrophic retina was studied by enzyme-linked immunosorbent assay ( ELISA) and immunohistochemistry analysis. Microglial expression of TNF-alpha was determined by double immunolabeling. RESULTS. Whereas photoreceptor apoptosis in the rd mice started at P10 and reached a peak at P16, activation and migration of microglial cells were observed at P10 and peaked at P14. The expression of MCP-1, MCP-3, MIP-1 alpha, MIP-1 beta, and RANTES transcripts were noted at P8 and reached a peak at P12. Production of TNF-alpha was noted in the outer nuclear layer (ONL) of the rd mice at P8 and reached a peak at P12. At the peak of microglial activity, TNF-alpha was predominantly expressed in the activated microglial cells in the ONL. CONCLUSIONS. Activation of microglia, as well as expression of their signaling molecules ( chemokines) and microglia-derived toxic factor (TNF-alpha), coincides with or precedes the occurrence of photoreceptor apoptosis, suggesting activated microglia play a major role in retinal degeneration in rd mice. The chemokines MCP-1, MCP-3, MIP-1 alpha, MIP-1 beta, and RANTES are involved in activation and recruitment of the microglia to the degenerating photoreceptor cell layer. TNF-alpha, produced by the activated microglia, may accentuate the photoreceptor cell death.