M2 muscarinic receptor activation inhibits cell proliferation and migration of rat adipose‐mesenchymal stem cells

Abstract

Mesenchymal stem cells (MSCs), also known as stromal mesenchymal stem cells, are multipotent cells, which can be found in many tissues and organs as bone marrow, adipose tissue and other tissues. In particular MSCs derived from Adipose tissue (ADSCs) are the most frequently used in regenerative medicine because they are easy to source, rapidly expandable in culture and excellent differentiation potential into adipocytes, chondrocytes, and other cell types. Acetylcholine (ACh), the most important neurotransmitter in Central nervous system (CNS) and peripheral nervous system (PNS), plays important roles also in non-neural tissue, but its functions in MSCs are still not investigated. Although MSCs express muscarinic receptor subtypes, their role is completely unknown. In the present work muscarinic cholinergic effects were characterized in rat ADSCs. Analysis by RT-PCR demonstrates that ADSCs express M1-M4 muscarinic receptor subtypes, whereas M2 is one of the most expressed subtype. For this reason, our attention was focused on M2 subtype. By using the selective M2 against Arecaidine Propargyl Ester (APE) we performed cell proliferation and migration assays demonstrating that APE causes cell growth and migration inhibition without affecting cell survival. Our results indicate that ACh via M2 receptors, may contribute to the maintaining of the ADSCs quiescent status. These data are the first evidence that ACh, via muscarinic receptors, might contribute to control ADSCs physiology.

Muscarinic receptors modulate Nerve Growth Factor production in rat Schwann-like adipose-derived stem cells and in Schwann cells

“…P0 and MBP) 23 . dASCs express functional receptors for several neurotransmitters such as GABA, ATP [29][30][31] and all muscarinic receptor subtypes 32,33 . In dASCs, M2 receptor activation produces a reversible decrease of cell proliferation, reduces migration and enhances dASCs differentiation as shown by improved spindle shaped morphology accompanied by early growth factor 2 (EGR2) upregulation 33 .…”

Effects mediated by the α7 nicotinic acetylcholine receptor on cell proliferation and migration in rat adipose-derived stem cells

“…Previously, the characterization of stemness markers to confirm the stemness nature of ASCs cultures was performed. 32 The α7 nAChR expression was analysed in ASCs by RTPCR and Western blot analysis, which allowed to detect expression of both α7 transcript ( Figure 1A ) and protein, respectively ( Figure 1B ). We evaluated whether the activation of the α7 subtype could induce an autocrine regulation of its expression in ASCs.…”

“…Given the results obtained in a previous study about the M2 receptor meaningful upregulation after exposure to the M2 receptor agonist, APE, 32 we examined the putative modulation of the α7 nAChR and M2 mAChR expression upon treatment with M2 receptor agonist and ICH3. Figure 5A shows the lack of a significant modulation of the α7 receptor protein expression after treatment with the M2 receptor agonist APE.…”

“…ASCs were harvested as previously reported; 32 , 40 ASCs were isolated from adult male (3 months) Sprague-Dawley rats. Fat pads were dissected and minced using a sterile razor blade.…”

“…The M2 muscarinic receptor selective agonist arecaidine propargyl ester hydrobromide (APE, Sigma-Aldrich, Milan, Italy) was used at the final concentration of 100 μM. 32 , 45 Controls were obtained maintaining the cells in normal growth medium. Technical and experimental triplicates were performed for all experiments.…”