Abstract
Purpose: :
Pituitary adenylate cyclase-activating polypeptide (PACAP) is a pleiotropic neuropeptide that belongs to the secretin/glucagon/vasoactive intestinal peptide superfamily. Previously, we have revealed that PACAP has a neuroprotective effect against N-methyl-D-aspartic acid (NMDA)-induced retinal injury. It is recently reported that increment of the number of retinal microglia/macrophage (MG/Mϕ) induced by retinal degenerations contribute to neuroprotection, and one of the microglial activation states, "acquired deactivation" ameliorates neuroinflammation. However, relation between MG/Mϕ and neuroprotective effect of PACAP is still unknown. The aim of this study is to clarify the relationship between the retinoprotective effect of PACAP and the activation of retinal MG/Mϕ on NMDA-induced retinal injury.
Methods: :
Adult male C57/BL6 mice were received 40 nM NMDA with/without PACAP injection into their vitreous chamber. At 3 days after the injection, eyes were enucleated and prepared for cryosections for immunohistochemical analysis, and separated retinal samples for real time-PCR analysis. By histological observation, retinal MG/Mϕ labeled by anti-Iba1 antibody and retinal ganglion cells in the retina were counted.
Results: :
In NMDA and PACAP co-treated retina, the number of the retinal MG/Mϕ and surviving retinal ganglion cells were significantly higher than that in NMDA-treated retina. The numbers of retinal MG/Mϕ and retinal ganglion cells were significantly correlated. The mRNA levels of several cytokines related to MG/Mϕ activation such as IL-6, IL-10, IFN-γ, TNF-α and TGF-β1 were evaluated by real time PCR method. PACAP co-treatment did not affect pro-inflammatory cytokines such as IL-6, IFN-γ and TNF-α level, but elevated anti-inflammatory cytokines such as TGF-β1 and IL-10 mRNA level significantly in the retina. By double immunostaining, the TGF-β1 immunoreactivity was co-existed with MG/Mϕ marker, CD11b immunoreactivity. The PACAP-induced increment of retinal MG/Mϕ and protection of RGCs attenuate in IL-10 KO mouse.
Conclusions: :
These results suggest that PACAP enhances the proliferation or infiltration of retinal MG/Mϕ and modulates activation state to "acquired deactivation" that is known to have a neuroprotective potent in the injured retina.
Keywords: neuroprotection • neuropeptides • microglia