Biological behaviors of muscarinic receptors in mesenchymal stem cells derived from human placenta and bone marrow

Document Type: Original Article

Authors

1 Hatay Mustafa Kemal University, Department of Pharmacology and Toxicology, Hatay, Turkey

2 Alanya Alaaddin Keykubat University, Department of Medical Pharmacology, Antalya, Turkey

3 Başkent University, Faculty of Medicine, Adult Bone Marrow Transplant Center, Adana, Turkey

4 Hatay Mustafa Kemal University, Faculty of Medicine, Department of Medical Biology, Hatay, Turkey

5 Cukurova University, Faculty of Medicine, Department of Medical Pharmacology, Adana, Turkey

Abstract

Objective(s): Cells perform their functional activities by communicating with each other through endogenous substances and receptors. Post-translation, stem cells function properly in new host tissue by carrying specific cell surface receptors. We aimed to characterize muscarinic receptor subtypes in mesenchymal stem cells (MSCs) together with osteogenic and adipogenic differentiation markers.
Materials and Methods: mRNA levels of 5 muscarinic receptor subtypes (CHRM1 to 5), BMP-6, and PPARγ during osteogenic and adipogenic differentiation, under the effect of atropine blockade, were measured in MSCs obtained from human fetal membrane (FM) and bone marrow (BM). Additionally, the effect of atropine on differentiation in the 1st, 2nd, and 3rd passages of MSCs, obtained from human FM and BM, were analyzed by RT-qPCR.
Results: CHRM1 mRNA levels increased in the FM group, while decreasing in the BM group. We found significant decreases in CHRM3 and CHRM5 mRNA levels in FM and BM groups, respectively. Atropine had variable effects based on cell source and receptor type. BMP-6 mRNA levels in differentiated osteogenic cells increased significantly compared to undifferentiated cells in both FM and BM groups. In MSCs derived from both sources, PPARγ mRNA levels in differentiated adipogenic cells increased significantly. Atropine showed no effect on MSCs differentiation.
Conclusion: These results indicate that expressions of muscarinic receptors in MSCs derived from BM and FM can vary and these cells keep the potential of osteogenic and adipogenic differentiation in vitro. Besides, atropine had no effect on adipogenic and osteogenic differentiation of MSCs.

Keywords


1. Friedenstein AJ, Deriglasova UF, Kulagina NN, Panasuk AF, Rudakowa SF, Luria EA, et al. Precursors for fibroblasts in different populations of hematopoietic cells as detected by the in vitro colony assay method. Exp Hematol 1974; 2:83-92.
2. Javanmard F, Azadbakht M, Pourmoradi M. The effect of hydrostatic pressure on staurosporine-induced neural differentiation in mouse bone marrowderived mesenchymal stem cells. Bratisl Lek Listy 2016; 117:283-289.
3. Branch MJ, Hashmani K, Dhillon P, Jones DR, Dua HS, Hopkinson A. Mesenchymal stem cells in the human corneal limbal stroma. Invest Ophthalmol Vis Sci 2012; 53:5109-5116.
4. Chong PP, Selvaratnam L, Abbas AA, Kamarul T. Human peripheral blood derived mesenchymal stem cells demonstrate similar characteristics and chondrogenic differentiation potential to bone marrow derived mesenchymal stem cells. J Orthop Res 2012; 30:634-642.
5. Wang S, Qu X, Zhao RC. Clinical applications of mesenchymal stem cells. J Hematol Oncol 2012; 5:19.
6. Horwitz EM, Le Blanc K, Dominici M, Mueller I, Slaper-Cortenbach I, Marini FC, et al. Clarification of the nomenclature for MSC: The International Society for Cellular Therapy position statement. Cytotherapy 2005; 7:393-395.
7. Pittenger MF, Mackay AM, Beck SC, Jaiswal RK, Douglas R, Mosca JD, et al. Multilineage potential of adult human mesenchymal stem cells. Science 1999; 284:143-147.
8. Matuskova M, Durinikova E, Altaner C, Kucerova L. Genetically engineered mesenchymal stromal cells in cancer gene therapy. Bratisl Lek Listy 2018; 119:221-223.
9. Benavente CA, Sierralta WD, Conget PA, Minguell JJ. Subcellular distribution and mitogenic effect of basic fibroblast growth factor in mesenchymal uncommitted stem cells. Growth Factors 2003; 21:87-94.
10. Boland GM, Perkins G, Hall DJ, Tuan RS. Wnt 3a promotes proliferation and suppresses osteogenic differentiation of adult human mesenchymal stem cells. J Cell Biochem 2004; 93:1210-1230.
11. Diefenderfer DL, Osyczka AM, Reilly GC, Leboy PS. BMP responsiveness in human mesenchymal stem cells. Connect Tissue Res 2003; 44 Suppl 1:305-311.
12. Li H, Yu B, Zhang Y, Pan Z, Xu W, Li H. Jagged1 protein enhances the differentiation of mesenchymal stem cells into cardiomyocytes. Biochem Biophys Res Commun 2006; 341:320-325.
13. Krampera M, Pasini A, Rigo A, Scupoli MT, Tecchio C, Malpeli G, et al. HB-EGF/HER-1 signaling in bone marrow mesenchymal stem cells: inducing cell expansion and reversibly preventing multilineage differentiation. Blood 2005; 106:59-66.
14. Erices A, Conget P, Rojas C, Minguell JJ. Gp130 activation by soluble interleukin-6 receptor/interleukin-6 enhances osteoblastic differentiation of human bone marrow-derived mesenchymal stem cells. Exp Cell Res 2002; 280:24-32.
15. Paraoanu LE, Steinert G, Koehler A, Wessler I, Layer PG. Expression and possible functions of the cholinergic system in a murine embryonic stem cell line. Life Sci 2007; 80:2375-2379.
16. Serobyan N, Jagannathan S, Orlovskaya I, Schraufstatter I, Skok M, Loring J, et al. The cholinergic system is involved in regulation of the development of the hematopoietic system. Life Sci 2007; 80:2352-2360.
17. Cui QL, Fogle E, Almazan G. Muscarinic acetylcholine receptors mediate oligodendrocyte progenitor survival through Src-like tyrosine kinases and PI3K/Akt pathways. Neurochem Int 2006; 48:383-393.
18. Ma W, Li BS, Zhang L, Pant HC. Signaling cascades implicated in muscarinic regulation of proliferation of neural stem and progenitor cells. Drug News Perspect 2004; 17:258-266.
19. Hoogduijn MJ, Cheng A, Genever PG. Functional nicotinic and muscarinic receptors on mesenchymal stem cells. Stem Cells Dev 2009; 18:103-112.
20. Kawano S, Shoji S, Ichinose S, Yamagata K, Tagami M, Hiraoka M. Characterization of Ca(2+) signaling pathways in human mesenchymal stem cells. Cell Calcium 2002; 32:165-174.
21. Schraufstatter IU, DiScipio RG, Khaldoyanidi SK. Alpha 7 subunit of nAChR regulates migration of human mesenchymal stem cells. J Stem Cells 2009; 4:203-215.
22. Grassi F, Pagani F, Spinelli G, De Angelis L, Cossu G, Eusebi F. Fusion-independent expression of functional ACh receptors in mouse mesoangioblast stem cells contacting muscle cells. J Physiol 2004; 560:479-489.
23. Hosey MM. Diversity of structure, signaling and regulation within the family of muscarinic cholinergic receptors. FASEB J 1992; 6:845-852.
24. Danielyan L, Schafer R, Schulz A, Ladewig T, Lourhmati A, Buadze M, et al. Survival, neuron-like differentiation and functionality of mesenchymal stem cells in the neurotoxic environment: the critical role of erythropoietin. Cell Death Differ 2009; 16:1599-1614.
25. Kim MH, Kim MO, Heo JS, Kim JS, Han HJ. Acetylcholine inhibits long-term hypoxia-induced apoptosis by suppressing the oxidative stress-mediated MAPKs activation as well as regulation of Bcl-2, c-IAPs, and caspase-3 in mouse embryonic stem cells. Apoptosis 2008; 13:295-304.
26. Boss A, Oppitz M, Lippert G, Drews U. Muscarinic cholinergic receptors in the human melanoma cell line SK-Mel 28: modulation of chemotaxis. Clin Exp Dermatol 2005; 30:557-564.
27. Chernyavsky AI, Arredondo J, Wess J, Karlsson E, Grando SA. Novel signaling pathways mediating reciprocal control of keratinocyte migration and wound epithelialization through M3 and M4 muscarinic receptors. J Cell Biol 2004; 166:261-272.
28. Yang K, Song Y, Tang YB, Xu ZP, Zhou W, Hou LN, et al. mAChRs activation induces epithelial-mesenchymal transition on lung epithelial cells. BMC Pulm Med 2014; 14:53.
29. Hakuno D, Fukuda K, Makino S, Konishi F, Tomita Y, Manabe T, et al. Bone marrow-derived regenerated cardiomyocytes (CMG Cells) express functional adrenergic and muscarinic receptors. Circulation 2002; 105:380-386.
30. Koch TG, Thomsen PD, Betts DH. Improved isolation protocol for equine cord blood-derived mesenchymal stromal cells. Cytotherapy 2009; 11:443-447.
31. Li H, Fong C, Chen Y, Cai G, Yang M. Beta-adrenergic signals regulate adipogenesis of mouse mesenchymal stem cells via the cAMP/PKA pathway. Mol Cell Endocrinol 2010; 323:201-207.
32. Zachos TA, Shields KM, Bertone AL. Gene-mediated osteogenic differentiation of stem cells by bone morphogenetic proteins-2 or -6. J Orthop Res 2006; 24:1279-1291.