Characterization of Differentiated SH-SY5Y as Neuronal Screening Model Reveals Increased Oxidative Vulnerability

Abstract

The immortalized and proliferative cell line SH-SY5Y is one of the most commonly used cell lines in neuroscience and neuroblastoma research. However, undifferentiated SH-SY5Y cells share few properties with mature neurons. In this study, we present an optimized neuronal differentiation protocol for SH-SY5Y that requires only two work steps and 6 days. After differentiation, the cells present increased levels of ATP and plasma membrane activity but reduced expression of energetic stress response genes. Differentiation results in reduced mitochondrial membrane potential and decreased robustness toward perturbations with 6-hydroxydopamine. We are convinced that the presented differentiation method will leverage genetic and chemical high-throughput screening projects targeting pathways that are involved in the selective vulnerability of neurons with high energetic stress levels.

Transcriptomic Profiling Discloses Molecular and Cellular Events Related to Neuronal Differentiation in SH-SY5Y Neuroblastoma Cells

“…This is of particular relevance considering that differentiated SH-SY5Y cells are used as in vitro model to study cellular mechanisms involved in several human neurodegenerative disorders. Exposure to differentiation media may unsettle different cellular behaviours (such as the response to oxidative stress) and, therefore, the cell properties to cope with the selected stimuli, even affecting their survival capacities and/or vulnerability to cell death (Forster et al 2016;Palomo et al 2011;Schneider et al 2011).…”

Caspase-1 Deficiency Alleviates Dopaminergic Neuronal Death via Inhibiting Caspase-7/AIF Pathway in MPTP/p Mouse Model of Parkinson’s Disease

“…The human neuroblastoma SH-SY5Y cell line has been regarded as a dopaminergic cell model for PD research [23]. Thus, we cultured SH-SY5Y cells and stimulated with 1-methyl-4-phenylpyridinium (MPP + , an active metabolite of MPTP), so as to give more mechanistic insight at the cellular level.…”

RA Differentiation Enhances Dopaminergic Features, Changes Redox Parameters, and Increases Dopamine Transporter Dependency in 6-Hydroxydopamine-Induced Neurotoxicity in SH-SY5Y Cells

“…Furthermore, different protocols also may cause changings in cell susceptibility to neurotoxins, such as 6-hydroxydopamine (6-OHDA) (Cheung et al 2009;Lopes et al 2010;Forster et al 2016). In vivo, it is widely accepted that this toxin enters into the dopaminergic neuron via DAT and causes a massive oxidative stress (Ljungdahl et al 1971).…”

“…Even though with the emergence of new, more physiologically relevant models such as iPS as in vitro models for PD (Hartfield et al 2014), it is clear that the majority of studies have been undertaken using cell lines such as SH-SY5Y due to considerations such as availability of iPS and the necessary expertise in their differentiation into dopaminergic neurons (Filograna et al 6 2015;Forster et al 2016;Lin and Tsai 2016). Hence, an understanding of the potential differences in SH-SY5Y cell line RA-differentiated and undifferentiated states and their response to 6-OHDA are imperative as this remains the most commonly-used in vitro model (Kovalevich and Langford 2013).…”