Neuroprotective Effect of Chitosan Nanoparticle–Delivered Orientin Against Hypoxia-Induced Brain Injury in Rats

Reduced brain oxygenation rapidly amplifies oxidative and inflammatory injury, creating a self-perpetuating cascade that can end in neuronal loss. Orientin can activate Nrf2/ARE-dependent antioxidant responses and suppress NF-κB–mediated inflammation, but poor solubility and limited bioavailability weaken its therapeutic impact. Here, we used chitosan nanoparticles (CNPs) to improve Orientin delivery and evaluated the protective effects of CNPs/Orientin on oxidative stress, inflammation, and apoptosis in hypoxia-induced brain injury. Forty-eight male Wistar albino rats were split into eight groups: (Control, Orientin, CNPs, CNPs/Orientin, Hypoxia, Hypoxia+Orientin, Hypoxia+CNPs, Hypoxia+CNPs/Orientin). The hypoxia model was applied with an 8% O₂+92% N₂ gas mixture for 8 h per day for 7 days. Orientin (40 mg/kg/day, i.p.) was administered simultaneously with the hypoxia period. MDA, SOD, GSH, BDNF, HIF-1α, iNOS, IL-1β, and TNF-α levels were determined by ELISA; gene expressions of Caspase-3, Bcl-2, HO-1, NRF2, NF-κB, and TNF-α were quantified by RT-PCR. Histological evaluations were performed in the cortex and hippocampus regions. CNPs/Orientin had a mean diameter of ~245 nm, a positive zeta potential (+29 mV), 81.3% encapsulation efficiency, and sustained release (79.6% over 72 h). Hypoxia significantly decreased SOD, GSH, and BDNF, while increasing MDA, HIF-1α, iNOS, and Caspase-3 levels (p < 0.05). All treatment groups attenuated the hypoxia-induced increase in IL-1β and TNF-α; however, cytokine levels did not differ significantly among the treatment arms. In the Hypoxia+CNPs/Orientin group, oxidative/nitrosative markers and neuronal histopathological findings were improved compared with hypoxia. Orientin delivered via chitosan nanoparticles demonstrated a significant neuroprotective effect, reducing oxidative stress, inflammation, and apoptosis while preserving neuronal structural integrity in hypoxia-induced brain injury. These results suggest that the nano-carrier form of Orientin may be a potential therapeutic agent in neurodegenerative processes.