RESUMO
Objective To observe the effects of low dose irradiation (LDR) on proliferation,adipogenesis and osteogenic potential of human umbilical cord mesenchymal stem cells (hucMSCs).Methods hucMSCs were isolated from Wharton's jelly tissue of human umbilical cord by modified tissuepiece inoculation,and flow cytometry was used to detect the expression of specific marker in the hucMSCs.The hucMSCs were randomly divided into two groups:irradiation group undergoing irradiation with the doses 50,100,or 200 mGy respectively,and control group without irradiation.MTT method was applied to evaluate the proliferation of the hucMSCs at different time points with various doses irradiation.The third passage hucMSCs were randomly divided into two groups:irradiation group undergoing low dose irradiation of 200 mGy,and control group without irradiation,and then underwent induction by adipocytic and oesteocytic differentiation induction fluids respectively so as to differentiate into adipocytes and osteoblasts.Oil red O staining was used to detect the activity of alkaline phophatase (ALP),and RT-PCR was used to detect the mRNA expression of core binding factor alpha 1 in human osteoblast.Results After 9-12days,fibroblasts began to swim out of the tissue piece with a confluence rate of 80% 2 weeks later.Within 7 days the absorption values of the hucMSCs undergoing different irradiation doses 2,3,4,5,and 6 days later were all significantly higher than those of the control group(F = 159.17,448.81,265.15,183.93,and 181.83 ,all P <0.01),with the proliferation rates of the 100 mGy subgroup being the highest.After being induced liquid,vacuoles were observed in the irradiated group 2 days later.21 days later,the adipogenic rates of irradiated group was significantly higher than that of the control group (t = 28.25,P <0.01).The ALP activity increased in the irradiated group compared with control group (t=16.87,P <0.01) .The expression level of Cbf-α1 mRNA was up-regulated obviously (t = 14.16,P＜0.01).Conclusions LDR promotes the proliferation of hucMSCs,and accelerates the hucMSCs' differentiation into adipocytes and osteoblasts.
RESUMO
Objective To investigate the effects of icarrin on the activity and protein expression of core binding factor otl(Cbfa1) in rat osteoblasts cultured in vitro,and to explore whether mitogen-activated protein kinase (MAPK) pathway is involved in this process.Methods Calvarial osteoblasts were obtained from newborn (<24 h) SD rats by trypsin-coUagenase digestion method.The second generation osteoblasts were cultured in the medium containing icariin (10 ng/ml) or estradiol (10-8 mol/L) with or without extracellular-signal regulated kinase (ERK) inhibitor (UO126) or p38MAPK inhibitor (SB203580).Nuclear protein was extracted from osteoblasts.And then the activity of Cbfa1 was detected by ELISA.The amounts of Cbfa1 protein were detected by Western blot.Results Calvarial osteoblasts were obtained successfully and were used in this study after indentified by alkaline phosphatase and mineralized nodus staining.Cbfa1 expression and the activity in osteoblasts were up-regulated by both icariin and estradiol (P<0.05).The effects were partly inhibited by addition of U0126or SB203580 (P<0.05).Conclusions Either icarrin or estradiol can stimulate the proliferation and maturation of cultured osteoblasts in vitro via up-regulating the activity and expression of Cbfal.The MAPK signal pathway inhibitor seems to partly decrease Cbfa1 activity.It suggests that MAPK pathway may be involved in the transduction of icariin's impact on proliferation and mineralization of osteoblasts.