title: [HYPOXIA INDUCIBLE FACTOR lα/2α GENES EXPRESSION IN CHONDROGENIC DIFFERENTIATION OF HUMAN BONE MARROW MESENCHYMAL STEM CELLS].
abstract: OBJECTIVE
  To observe the genes expression of hypoxia inducible factor lα (HIF-1α) and HIF- 2α by inducing chondrogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) so as to provide a fundamental basis for HIF involving in the mechanism of chondrogenesis.
  
  
  METHODS
  High density pellet of hBMSCs was obtained by centrifugation and cultured with H-DMEM medium containing 2% fetal bovine serum (control group) and with chondrogenic medium (chondrogenic induction group) under hypoxia (2% O2) for 3 weeks. Immunohistochemistry staining was utilized to identify extracellular proteoglycan and collagen type II at 3 weeks after culture. Western blot was applied for measuring HIF-1α and HIF-2α protein levels at 1 week after culture. Real-time quantitative PCR was performed to detect the genes expressions of HIF-1α, HIF-2α, Sox-9, collagen type II, collagen type X, and Aggrecan at 1, 2, and 3 weeks after culture.
  
  
  RESULTS
  Toluidine blue staining showed sparse nucleus in the control group, and dense nucleus in the chondrogenic induction group; extracellular matrix staining was deeper in the chondrogenic induction group than the control group. Immunohistochemical staining for collagen type II was positive in cytoplasm; when compared with the chondrogenic induction group, the control group showed sparse and light-coloured nucleus. At 1 week after culture, the protein expression levels of HIF-1α and HIF-2α in the chondrogenic induction group were significantly lower than those in the control group (t = 8.345, P = 0.001; t = 7.683, P = 0.002). When compared with control group, the HIF-1α mRNA expression was significantly down-regulated at 1 week and significantly up-regulated at 2 weeks in chondrogenic induction group (P < 0.05), but no significant difference was found at 3 weeks between the 2 groups (P > 0.05). And the mRNA expression of HIF-2α was significantly down-regulated and mRNA expression of Sox-9 was significantly up-regulated after chondrogenic differentiation when compared with the control group (P < 0.01). The mRNA expressions of collagen type II and collagen type X were significantly up-regulated at 2 and 3 weeks after chondrogenic differentiation when compared with the control group (P < 0.05). And the mRNA expression of Aggrecan was significantly up-regulated at each time point after chondrogenic differentiation (P < 0.05).
  
  
  CONCLUSIONS
  HIF-1α may involve the hBMSCs chondrogenic differentiation under hypoxia, while HIF-2α expression is depressed throughout the period and may have negative effect on differentiation.
authors.forename: Ming
authors.initials: M
authors.lastname: Gong
authors.forename: Sheng
authors.initials: S
authors.lastname: Huang
authors.forename: Jiaquan
authors.initials: J
authors.lastname: Luo
authors.forename: Baoding
authors.initials: B
authors.lastname: Huang
authors.forename: Zhiyu
authors.initials: Z
authors.lastname: Zhou
authors.forename: Xuejun
authors.initials: X
authors.lastname: Dai
authors.forename: Manman
authors.initials: M
authors.lastname: Gao
authors.forename: Liangping
authors.initials: L
authors.lastname: Li
authors.forename: Xuenong
authors.initials: X
authors.lastname: Zou
mesh_terms: D053674:Aggrecans / Q000235:genetics / Q000378:metabolism; D001854:Bone Marrow Cells / Q000166:cytology* / Q000378:metabolism; D002454:Cell Differentiation; D015687:Cell Hypoxia; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000502:physiology*; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D024121:Collagen Type X / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006412:Hematopoietic Stem Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism*; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism; D011509:Proteoglycans
chemical_list: D053674:Aggrecans; D024043:Collagen Type II; D024121:Collagen Type X; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D011509:Proteoglycans