Effects of Collagen Coating, Fetal Bovine Serum Concentration, Differentiation Agents, and Neurotoxin Application on In Vitro Modeling of Parkinson's Disease Using SH-SY5Y Cell Culture

Abstract

Aim: This study aims to optimize SH-SY5Y culture conditions to develop precise in vitro disease models for Parkinson's disease (PD) research. It seeks to investigate the effects of various factors such as collagen coating, fetal bovine serum concentration, differentiation agents, and neurotoxin treatments on cellular behavior and disease modeling. Materials and Methods: The human neuroblastoma SH-SY5Y cell line was cultured in DMEM/F12 supplemented with heat-inactivated fetal bovine serum (FBS), penicillin-streptomycin, and L-glutamine. Collagen coating was applied to assess its impact on cell differentiation, while the ideal cell density and serum ratio for generating neurite-like cells were determined through experimentation. The MTT assay was employed to evaluate the cytotoxic effects of paraquat, while dopamine levels were quantified using ELISA. Gene expression was analyzed via real-time qPCR. Immunofluorescence staining and neurite length measurements were conducted to validate the PD model and assess cellular morphology. Results: Cells cultured at a density of 5x103 cells/cm2 with collagen and 2% FBS exhibited characteristics of dopaminergic neurons upon exposure to retinoic acid. Conversely, paraquat treatment induced neurotoxicity, resulting in decreased dopamine levels and impaired neurite outgrowth. Conclusion: This study investigated the optimization of SH-SY5Y cell culture conditions for PD modeling. Key findings include optimal cell density, FBS concentration, and beneficial effects of collagen coating. Additionally, an effective paraquat neurotoxicity protocol has been established, providing a solid framework for future research on neuronal differentiation and degeneration.