Effects of immunosuppressants, FK506 and cyclosporin A, on the osteogenic differentiation of rat mesenchymal stem cells
2
조회
0
다운로드
1
인용
0
참고문헌
키워드
초록
Purpose: The purpose of this study was to investigate the effects of the immunosuppressants FK506 and cyclosporin A (CsA)on the osteogenic differentiation of rat mesenchymal stem cells (MSCs). Methods: The effect of FK506 and CsA on rat MSCs was assessed in vitro. The MTT assay was used to determine the deleterious effect of immunosuppressants on stem cell proliferation at 1, 3, and 7 days. Alkaline phosphatase (ALP) activity was analyzed on days 3, 7, and 14. Alizarin red S staining was done on day 21 to check mineralization nodule formation. Real-time polymerase chain reaction (RT-PCR) was also performed to detect the expressions of bone tissue-specific genes on days 1 and 7. Results: Cell proliferation was promoted more in the FK506 groups than the control or CsA groups on days 3 and 7. The FK506groups showed increased ALP activity compared to the other groups during the experimental period. The ALP activity of the CsA groups did not differ from the control group in any of the assessments. Mineralization nodule formation was most prominent in the FK506 groups at 21 days. RT-PCR results of the FK506 groups showed that several bone-related genes-osteopontin,osteonectin, and type I collagen (Col-I)-were expressed more than the control in the beginning, but the intensity of expression decreased over time. Runx2 and Dlx5 gene expression were up-regulated on day 7. The effects of 50 nM CsA on osteonectin and Col-I were similar to those of the FK506 groups, but in the 500 nM CsA group, most of the genes were less expressed compared to the control. Conclusions: These results suggest that FK506 enhances the osteoblastic differentiation of rat MSCs. Therefore, FK506 might have a beneficial effect on bone regeneration when immunosuppressants are needed in xenogenic or allogenic stem cell transplantation to treat bone defects.
Abstract
Purpose: The purpose of this study was to investigate the effects of the immunosuppressants FK506 and cyclosporin A (CsA)on the osteogenic differentiation of rat mesenchymal stem cells (MSCs). Methods: The effect of FK506 and CsA on rat MSCs was assessed in vitro. The MTT assay was used to determine the deleterious effect of immunosuppressants on stem cell proliferation at 1, 3, and 7 days. Alkaline phosphatase (ALP) activity was analyzed on days 3, 7, and 14. Alizarin red S staining was done on day 21 to check mineralization nodule formation. Real-time polymerase chain reaction (RT-PCR) was also performed to detect the expressions of bone tissue-specific genes on days 1 and 7. Results: Cell proliferation was promoted more in the FK506 groups than the control or CsA groups on days 3 and 7. The FK506groups showed increased ALP activity compared to the other groups during the experimental period. The ALP activity of the CsA groups did not differ from the control group in any of the assessments. Mineralization nodule formation was most prominent in the FK506 groups at 21 days. RT-PCR results of the FK506 groups showed that several bone-related genes-osteopontin,osteonectin, and type I collagen (Col-I)-were expressed more than the control in the beginning, but the intensity of expression decreased over time. Runx2 and Dlx5 gene expression were up-regulated on day 7. The effects of 50 nM CsA on osteonectin and Col-I were similar to those of the FK506 groups, but in the 500 nM CsA group, most of the genes were less expressed compared to the control. Conclusions: These results suggest that FK506 enhances the osteoblastic differentiation of rat MSCs. Therefore, FK506 might have a beneficial effect on bone regeneration when immunosuppressants are needed in xenogenic or allogenic stem cell transplantation to treat bone defects.
참고문헌 (23)
- Otto WR, "Mesenchymal stem cells: from experiment to clinic", Fibrogenesis Tissue Repair, 4, pp.20~ (2011)
- Caplan AI, "Adult mesenchymal stem cells for tissue engineering versus regenerative medicine", J Cell Physiol, 213, pp.341~347 (2007)
- Nauta AJ, "Donor-derived mesenchymal stem cells are immunogenic in an allogeneic host and stimulate donor graft rejection in a nonmyeloablative setting", Blood, 108, pp.2114~2120 (2006)
- Penninga L, "Tacrolimus versus cyclosporine as primary immunosuppression after heart transplantation: systematic review with meta-analyses and trial sequential analyses of randomised trials", Eur J Clin Pharmacol, 66, pp.1177~1187 (2010)
- Vincenti F, "A long-term comparison of tacrolimus (FK506) and cyclosporine in kidney transplantation: evidence for improved allograft survival at five years", Transplantation, 73, pp.775~782 (2002)
- Epstein S, "Post-transplantation bone disease: the role of immunosuppressive agents and the skeleton", J Bone Miner Res, 11, pp.1~7 (1996)
- Kino T, "FK-506, a novel immunosuppressant isolated from a Streptomyces. I. Fermentation, isolation, and physico-chemical and biological characteristics", J Antibiot (Tokyo), 40, pp.1249~1255 (1987)
- Kino T, "FK-506, a novel immunosuppressant isolated from a Streptomyces. II. Immunosuppressive effect of FK-506 in vitro", J Antibiot (Tokyo), 40, pp.1256~1265 (1987)
- Kulak CA, "Post-transplantation osteoporosis", Arq Bras Endocrinol Metabol, 54, pp.143~149 (2010)
- Rodino MA, "Osteoporosis after organ transplantation", Am J Med, 104, pp.459~469 (1998)
- Stein E, "Post-transplantation osteoporosis", Endocrinol Metab Clin North Am, 36, pp.937~963 (2007)
- Cvetkovic M, "The deleterious effects of long-term cyclosporine A, cyclosporine G, and FK506 on bone mineral metabolism in vivo", Transplantation, 57, pp.1231~1237 (1994)
- Fukunaga J, "Expression of osteoclast differentiation factor and osteoclastogenesis inhibitory factor in rat osteoporosis induced by immunosuppressant FK506", Bone, 34, pp.425~431 (2004)
- Krocker D, "Effects of tacrolimus, cyclosporin A and sirolimus on MG63 cells", Transpl Int, 19, pp.563~569 (2006)
- Lee KW, "Rapamycin promotes the osteoblastic differentiation of human embryonic stem cells by blocking the mTOR pathway and stimulating the BMP", Smad pathway, 2010, pp.19, 557~568
- Isomoto S, "Rapamycin as an inhibitor of osteogenic differentiation in bone marrow-derived mesenchymal stem cells", J Orthop Sci, 12, pp.83~88 (2007)
- Kaihara S, "Simple and effective osteoinductive gene therapy by local injection of a bone morphogenetic protein-2-expressing recombinant adenoviral vector and FK506 mixture in rats", Gene Ther, 11, pp.439~447 (2004)
- Yoshikawa T, "In vivo osteogenic capability of cultured allogeneic bone in porous hydroxyapatite: immunosuppressive and osteogenic potential of FK506 in vivo", J Bone Miner Res, 15, pp.1147~1157 (2000)
- Kugimiya F, "Mechanism of osteogenic induction by FK506 via BMP", Smad pathways, 2005, pp.338, 872~879
- Liu J, "Calcineurin is a common target of cyclophilin-cyclosporin A and FKBP-FK506 complexes", Cell, 66, pp.807~815 (1991)
- Tang L, "FK506 enhanced osteoblastic differentiation in mesenchymal cells", Cell Biol Int, 26, pp.75~84 (2002)
- Owen TA, "Progressive development of the rat osteoblast phenotype in vitro: reciprocal relationships in expression of genes associated with osteoblast proliferation and differentiation during formation of the bone extracellular matrix", J Cell Physiol, 143, pp.420~430 (1990)
- 김수환, "Ex vivo bone morphogenetic protein-2 gene delivery using gingival fibroblasts promotes bone regeneration in rats", JOURNAL OF CLINICAL PERIODONTOLOGY, 37, pp.305~311, BLACKWELL PUBLISHING