Abstract
The Cath.a-differentiated (CAD) cell line is a central nervous system-derived catecholaminergic cell line originating from tyrosine hydroxylase (TH)-producing neurons located around the locus coeruleus area of the mouse brain. CAD cells have been used as an in vitro model for cellular and molecular studies due to their ability to differentiate under serum-free media conditions. However, the lack of serum-derived survival factors, limits the longevity for differentiated CAD cells to be maintained in healthy conditions; thereby, limiting their use in long-term culture studies. Here, we present a novel differentiation method that utilizes dexamethasone (Dex), a synthetic glucocorticoid receptor agonist. Specifically, we discovered that the addition of 100µM of Dex into the 1% fetal bovine serum (FBS)-supplemented media effectively induced neuronal differentiation of CAD cells, as characterized by neurite formation and elongation. Dex-differentiated CAD cells exited the cell cycle, stopped proliferating, extended the neurites, and expressed neuronal markers. These effects were dependent on the glucocorticoid receptors (GR) as they were abolished by GR knockdown. Importantly, Dex-differentiated CAD cells showed longer survival duration than serum-free differentiated CAD cells. In addition, RNA-sequencing and qPCR data demonstrate that several genes involved in proliferation, neuronal differentiation, and survival pathways were differentially expressed in the Dex-differentiated cells. This is the first study to reveal Dex as a novel differentiation methodology used to generate postmitotic neuronal CAD cells, which may be utilized as an in vitro neuronal model for cellular and molecular neurobiology research.
Highlights
Cath.a-differentiated (CAD) cells are a neuronal cell line originally established from mouse central nervous system (CNS)-derived catecholaminergic neurons located around the locus coeruleus area of mouse brain (Suri et al 1993; Qi et al 1997)
To test the ability of Dex to induce the differentiation of a neuronal cell line, we examined Dex-induced differentiation of CAD cells
We proposed a novel methodology for inducing the differentiation of a catecholaminergic CNSderived cell line, CAD cells, using the glucocorticoid receptor agonist in 1% serumcontaining medium
Summary
Cath.a-differentiated (CAD) cells are a neuronal cell line originally established from mouse central nervous system (CNS)-derived catecholaminergic neurons located around the locus coeruleus area of mouse brain (Suri et al 1993; Qi et al 1997). CAD cells have been used as an in vitro model for studying catecholaminergic neurons, largely because of their ability to be quickly switched from a proliferative phase to a CNS neuron-like differentiation phase by removing serum. CAD cells stop proliferating and differentiate into neuron-like cells. Differentiated CAD cells demonstrated a catecholaminergic phenotype, in which they show similar morphologies to primary CNS neurons and express several catecholaminergic neuronal markers (Qi et al 1997; Pasuit et al 2004; Muresan and Muresan 2006; Paris et al 2006; Arboleda et al 2007; Chesta et al 2014). CAD cells that are differentiated by serum withdrawal exit the cell cycle and morphologically differentiate into neuron-like cells; these cells cannot be effectively maintained for long durations, thereby limiting their applications as in vitro neuronal models. Alternative conditions that effectively differentiate, as well as prolong survival of, CAD cells are necessary
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
