[Effects of hypoxic preconditioning on the migration of bone marrow derived mesenchymal stem cells].
OBJECTIVE
To explore the effects and probable mechanism of CoCl2-induced hypoxic preconditioning on the migration of bone marrow derived mesenchymal stem cells (BMSC).


METHODS
BMSC were cultured by whole bone marrow adherence and identified by surface markers (CD29, CD90 and CD45) with flow cytometry (FCM). The methods of 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and FCM were applied to establish the model of CoCl2-induced hypoxic preconditioning. The migratory capacity of BMSC with hypoxic preconditioning was analyzed by the assays of scratch wound healing and transwell migration. The protein and mRNA expressions of HIF-1α and CXCR4 of BMSC were detected by Western blot and real-time polymerase chain reaction (PCR). After silencing HIF-1α by siRNA technique and blocking CXCR4 by its antagonist AMD3100, the changes of migratory capacity of BMSC were also tested.


RESULTS
Cultured BMSC were uniformly positive for CD29 and CD90 and negative for CD 45. According to the results of MTT and FCM, 200 µmol/L CoCl2 and 24 h culture time was the ideal hypoxic preconditioning model of BMSC. The migratory capacity of BMSC in hypoxic preconditioning group was higher than the one in control group (scratch wound healing assay: (0.396 ± 0.018) mm vs (0.200 ± 0.011) mm, transwell migration assay: 21.0 ± 4.5 vs 8.5 ± 1.7, both P < 0.05). The protein and mRNA levels of HIF-1α and CXCR4 of BMSC in hypoxic preconditioning group were significantly higher than in control group. After silencing HIF-1α or blocking CXCR4 by AMD3100, the migratory capacity of BMSC in hypoxic preconditioning group decreased and had no difference with the control group.


CONCLUSIONS
Hypoxic preconditioning may enhance the migratory capacity of BMSC in vitro. And it is partially attributable to the up-regulation of HIF-1α/CXCR4 axis after preconditioning.
Department of Nephrology, Zhongshan Hospital, Fudan University, Shanghai 200032, China.
Hong
H
Liu
Xiao-fang
XF
Yu
Jie
J
Teng
Jian-zhou
JZ
Zou
Yi
Y
Fang
Xiao-qiang
XQ
Ding
D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology*; D015687:Cell Hypoxia; D002465:Cell Movement; D002478:Cells, Cultured; D005434:Flow Cytometry; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D019718:Receptors, CXCR4 / Q000378:metabolism
C486264:Cxcr4 protein, rat; C497444:Hif1a protein, rat; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D019718:Receptors, CXCR4