Vitamin C and Methylprednisolone Modulate Apoptotic and Transcript Responses in a B-35 Scratch Screening Model

In vitro mechanical injury platforms using proliferative neuronal cell lines provide accessible systems for preliminary pharmacological screening before progression to differentiated, primary, or in vivo models. We investigated the effects of vitamin C and methylprednisolone, alone or in combination, on cell viability, Annexin V/PI-defined apoptotic cell death, scratch-gap closure, and selected transcript responses in the B-35 rat neuroblastoma cell line. Mechanical injury was induced in confluent B-35 monolayers using a standardized scratch method. Vitamin C, methylprednisolone, or their combination was applied either immediately after injury (0 h) or after a 24 h delay. Scratch-gap closure was quantified by image-based analysis, and IL-4, IL-10, and TUBB3 expression was evaluated by quantitative real-time PCR. Early combined exposure to vitamin C and methylprednisolone was associated with low apoptotic cell fractions, faster scratch-gap closure, and higher IL-4, IL-10, and TUBB3 mRNA levels than either single treatment, whereas delayed treatment showed attenuated responses. These findings describe treatment-associated changes in a simplified B-35 cell-line screening model. They should not be interpreted as evidence of regeneration, pharmacological synergy, neuroprotection, or spinal cord injury therapy. Protein-level validation, proliferation control, and confirmation in differentiated neuronal cultures, primary neuronglia co-cultures, organoid or slice models, and in vivo injury systems are required before mechanistic or translational conclusions can be drawn.