Peptide Fraction from Naja mandalayensis Snake Venom Showed Neuroprotection Against Oxidative Stress in Hippocampal mHippoE-18 Cells but Not in Neuronal PC12 Cells

Functional characterization of peptide fraction (PF) from snake venom has provided novel opportunities to investigate possible neuroprotective compounds relevant to pharmaceuticals. This study was performed to investigate the PF-mediated neuroprotection obtained from <i>Naja mandalayensis</i> snake venom, a member of the Elapidae family, using two neuronal cell lines, undifferentiated PC12 and differentiated mHippoE-18, in response to H₂O₂-induced oxidative stress. Cells were pre-treated for 4 h with PF (10, 1, 0.01, and 0.001 μg mL^-1), and thereafter exposed to H₂O₂ (0.5 mmol L^-1) for 20 h. Then, the oxidative stress markers and label-free differential proteome strategy were analyzed to understand the neuroprotective effects of PF. In PC12 cells, PF showed no neuroprotective effects against oxidative stress. In mHippoE-18 cells, PF at 0.01 and 0.001 μg mL^-1 increased the viability and metabolism of cells against H₂O₂-induced neurotoxicity, reducing reactive oxygen species (ROS) generation. Interestingly, PF also exhibited a substantial reduction in baseline ROS levels compared to the control, indicating that PF could have compounds with antioxidant features. The comparative proteomic profiling identified 53 proteins with differential expression related to antioxidant action, catalysis, molecular function regulators, structural molecule activity, translation regulatory activity, ATP, and binding. The PF + H₂O₂ group indicated that protein expression is 6% upregulated, 4% downregulated, and 94% unchanged compared to the H₂O₂ group. Three significant proteins upregulated in the PF + H₂O₂ group, including elongation factor 2 (P58252), proteasome subunit alpha type (E9Q0X0), and E2 ubiquitin-conjugating enzyme (A0A338P786), suggested that PF-mediated neuroprotection happens through translational regulation and the degradation of defective proteins via the proteasome complex. Additionally, differential protein expression in PF changed the metabolism, protein synthesis, synaptic activity, and intracellular transport, suggesting that PF contains the rich mixture of bioactive peptides of interest pharmacologically. Overall, this study offers new opportunities for evaluating whether PF's neuroprotective features in specific neuronal cells are maintained and to investigate neurodegenerative disease drug development processes.