The inflammatory microenvironment in the retina plays a vital role in the pathogenesis and progression of retinitis pigmentosa (RP). Microglial inflammatory cytokines production leads to gliosis and apoptosis of retinal neurons, and ultimately, visual loss. Cell-based therapies using grafted olfactory ensheathing cells (OECs) have demonstrated modulation of degenerative microenvironments in the central nervous system (CNS), in a number of animal models. However, mechanisms by which grafted OECs can reduce degeneration in the retina are not well understood. In the present study, we set up an in vitro OEC/BV2 microglia co-culture system, and an in vivo royal college of surgeons (RCS) rat model, used cell transplantation, immunohistochemistry, RT-PCR, western blot to explore the mechanisms by which OECs affect expression of pro- or anti-inflammatory cytokines and polarization of M(IL-6) and M(Arg1) type microglial activation in the retina. We found that compared with the LPS (Lipopolysaccharide) and olfactory nerve fibroblast (ONF), the OEC and BV2 co-culture group modulate microglial cytokines releasing toward the anti-inflammation, and away from the pro-inflammation, which was followed by higher IL-4 and IL-10 and lower TNF-a and IL-6 in their expression levels. In vivo , the transplantation group significantly reduced activated resident microglia/infiltrated macrophage, and expression of pro-inflammatory cytokines in RCS rats retina, increased anti-inflammatory cytokines in transplantation area. Additionally, we found that OECs expressed SOCS3 and down-regulated the JAK2/STAT3 (Janus Kinase 2/Signal Transducer and Activator of Transcription 3) pathway. Thirdly, OEC transplantation reduced Caspase-3 expression, protected inner retinal neurons and photoreceptors and therefore, delayed the visual function degeneration. In conclusion, our data suggest that OECs delay retinal degeneration in RP, at least in part through immunomodulation of microglia via the JAK/STAT pathway.

Retinal regeneration and self-repair, whether in response to injury or degenerative disease, are severely impeded by glial scar formation by Müller cells (specialized retinal macroglia). We have previously demonstrated that the activation of Müller cells and gliosis in the degenerative retina are significantly suppressed by the subretinal transplantation of a mixture of olfactory ensheathing cells (OECs) and olfactory nerve fibroblasts. However, the underlying molecular mechanism has remained elusive. Here we transplanted purified rat OECs into the subretinal space of pigmented Royal College of Surgeons (RCS) rats, a classic rodent model of retinal degeneration. Using behavioral testing and electroretinography, we confirmed that the grafted OECs preserved the visual function of rats for 8 weeks, relative to vehicle controls (phosphate-buffered saline). Histological evaluation of outer nuclear layer thickness and composition demonstrated that more photoreceptors and ON-bipolar cells were preserved in the retinas of OEC-treated RCS rats than in controls. The grafted OECs migrated into the outer plexiform layer, inner nuclear layer, and inner plexiform layer. They interacted directly with Müller cells in the retina of RCS rats, in three distinct patterns, and secreted matrix metalloproteinases 2 and 3. Previous studies have demonstrated that rat OECs express delta-like ligand (DLL), while Müller cells express Notch3, the receptor for DLL. Here we found that the grafted OECs significantly decreased the expression, by retinal cells, of Notch signaling pathway components (including Notch3, Notch4, DLL1, DLL4, Jagged1, Hes1, and Hes5) 2 weeks after the cell transplantation and that this effect persisted for a further 2 weeks. Based on these findings, we suggest that transplanted OECs inhibit the activation of Müller cells and the associated gliosis, at least partly through suppression of the Notch pathway.