Data Sheet 1_Calcium sulfide nanoparticles as sustained H2S donors with neuroprotective potential in ischemic stroke.docx

IntroductionAcute ischemic stroke remains a major cause of death and disability, underscoring the need for safe and effective neuroprotective strategies. Hydrogen sulfide (H₂S) exhibits dose-dependent neuroprotective effects but its therapeutic application is constrained by volatility and burst release.
MethodsWe synthesized low-solubility, slowly hydrolyzing calcium sulfide nanoparticles (CaS NPs) via a wet-chemistry route as an intrinsically slow-releasing H₂S donor. Their sustained release profile was characterized, and efficacy was evaluated in vitro using SH-SY5Y cells under oxygen-glucose deprivation/reoxygenation (OGD/R) and in BV2 microglia, and in vivo using a distal middle cerebral artery occlusion (dMCAO) mouse model.
ResultsCaS NPs demonstrated sustained H₂S release over 48 h. In vitro, they enhanced SH-SY5Y cell viability under OGD/R, decreased intracellular reactive oxygen species, suppressed TNF-α and IL-1β expression in BV2 cells, and reduced neuronal apoptosis. In the dMCAO model, CaS NPs increased cortical H₂S levels, improved 24-h neurological scores, reduced day-3 infarct area, preserved peri-infarct neurons, mitigated ROS accumulation, and attenuated astrocyte and microglia activation. Treatment consistently decreased Bax expression, increased Bcl-2 levels, and reduced pro-inflammatory cytokine expression. Short-term safety assessments indicated a favorable biosafety profile.
DiscussionCollectively, these findings provide proof-of-concept support that CaS NPs can serve as a slow-releasing H₂S donor platform for further evaluation in experimental ischemic stroke."