Abstract
Neuroscreen-1 (NS-1) a sub-clone of pheochromocytoma (PC12) cell is gaining broad acceptance as in vitro neuronal model for biochemical and phenotypic assays due to robust growth and differentiation profiles. However, the molecular characteristics of the cell remains to be documented. In this study, we performed comparative analysis for expression of neuronal marker genes in undifferentiated and nerve growth factor (NGF) differentiated NS-1 and PC12 by qPCR and immunoblot assays. We show that differentiation of NS-1 occurred under low concentrations of NGF relative to PC12. Cell growth also occurred more rapidly in NS-1. Transcriptional analysis of neuronal marker genes showed comparable expression of tyrosine receptor kinases (Ntrk1, Ntrk2, NGFR/p75NTR) and muscarinic acetylcholine (Chrm1, Chrm2, Chrm3, Chrm4) receptors in unspecialized cells. Ntrk2, adenosine receptors (Adora1, Adora2A) and choline acetyltransferase (ChAT) were altered in undifferentiated NS-1. In contrast, Ntrk1, Ntrk2, Chrm2 transcripts were vastly increased in NS-1 with NGF exposure, while Ntrk3, Adora1 and Adora2A transcripts were reduced. In differentiated PC12, Chrm4 and ChAT were markedly upregulated. Our data suggests that differences in morphological and phenotypic characteristics that distinguish NS-1 from PC12 is likely the product of altered gene expression. Furthermore, expression of neuron type genes in NS-1 support its use as an alternative model to PC12.
Highlights
Development of in vitro disease model that closely mimics in vivo condition has become significant with the emergence of new disease and pathological conditions
NS-1 is a clonal line of the pheochromocytoma cell PC12, which has the ability to differentiate into neuronal-like cells in presence of nerve growth factor (NGF)
NS-1 and PC12 cells exhibit phenotypic and functional characteristics associated with NGF differentiation in altered timescales
Summary
Development of in vitro disease model that closely mimics in vivo condition has become significant with the emergence of new disease and pathological conditions. Basic research for neurodegenerative disease largely relies on cell line studies, which act as the primary platform for drug screening in translational medicine. Complex nutritional requirements, slow growth properties, and expression of cell-type specific markers limit the choice of cell lines used in research. In addition to the neuroprotective and neuro-restorative properties of NGF, dysregulation in NGF signaling has been positively correlated with neurodegenerative disease including Alzheimer’s disease (AD) [7,8], epilepsy [9] and cancer [1012]. NGF regulates proliferation and differentiation of neuronal cells via activation of activation of tyrosine protein kinase (TrkA) receptor, and downstream signaling molecules that include Ras/MAP kinase cascade, IP3-dependent Ca2+ release, and PI3K/Akt pathways [13]. NGF increases expression of ChAT and VAChT, two cholinergic specific markers that are required for cholinergic neurotransmission [15,16]. Decrease in ChAT and VAChT is suggested to play a role in the progression of AD [18,19]
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