Platelet factor 4 attenuates inflammation of microglia and protects retinal ganglion cells in retinal excitotoxicity

Glaucoma, a progressive optic neuropathy characterized by RGC degeneration and irreversible vision loss, currently affects approximately 76 million individuals globally. Despite conventional therapeutic strategies primarily targeting IOP reduction, the ongoing progression of vision loss in normotensive patients highlights an urgent need for alternative neuroprotective interventions. We employed a comprehensive experimental paradigm that integrated both in vivo and in vitro approaches. The in vivo component was utilized by NMDA-induced excitotoxicity involving Sprague-Dawley rats. In vitro analyses were conducted using R28 and BV2 cells. Quantitative assessments encompassed electroretinography, RGC survival, axonal integrity measurements, inflammatory marker profiles, flow cytometry, as well as molecular pathway analyses through immunofluorescence microscopy, Western blot analysis. Administration of PF4 (500 ng/ml) exhibited significant neuroprotective efficacy via multiple cellular mechanisms. Quantitative analyses indicated substantial preservation of RGC density (p < 0.001) alongside maintenance of inner plexiform layer thickness(p < 0.05) within the NMDA-induced model. PF4 treatment markedly attenuated microglial activation (p < 0.01) while modulating the inflammatory response-characterized by reduced expression of pro-inflammatory cytokines coupled with enhanced production of anti-inflammatory mediators. CTB tracing confirmed the preservation of both RGC axons and their projections. Molecular analyses revealed that PF4 may exerted its effects on RGC through different mechanisms: suppression of the Galectin-3/NLRP3-inflammasome/Caspase-1 pathway in microglia and enhancement of the CaMKII/CREB/BDNF neuroprotective cascade within RGCs; these protective effects can attenuate necroptosis independent from IOP modulation in retinal excitotoxicity. Our findings suggest that PF4 can protect RGCs through activate CaMKII/CREB/BDNF pathway induced by excitotoxicity. Moreover, it attenuates NLRP3 inflammasome activation via mediating Galectin-3 and thus decreasing necroptosis of RGCs. This study demonstrates that PF4 may possesses neuroprotective properties through simultaneous modulation across multiple cellular pathways in glaucomatous neurodegeneration, and emphasized the significance of immune-mediated mechanisms.