title: HIF-1A and C/EBPs transcriptionally regulate adipogenic differentiation of bone marrow-derived MSCs in hypoxia.
abstract: BACKGROUND
  Bone marrow-derived mesenchymal stem cells (BMSCs, also known as bone marrow-derived mesenchymal stromal cells) are known to be a component of the tumor microenvironment. BMSCs are multipotent stromal cells that can differentiate into a variety of cell types, including osteocytes, chondrocytes, adipocytes, epithelial cells and endothelial cells. Stem cells found in niches or transplanted into injured tissues constantly encounter hypoxic stress. Areas with very low to no oxygen pressure exist in solid tumors. The differentiation capacity of BMSCs under hypoxic conditions remains controversial.
  
  
  METHODS
  In this study, a hypoxic workstation, set at an oxygen concentration of 0.2% was used to mimic the hypoxic microenvironment of cancer in vivo. Oil red O staining and alkaline phosphatase staining were used to examine the adipogenic or osteogenic differentiation, respectively, of BMSCs. Real-time PCR was performed to explore the expression of adipocyte- or osteocyte-specific genes. An RT2 Profiler™ PCR Array was used to screen a panel of 84 genes associated with human adipogenesis in BMSCs under normal and hypoxic conditions. A dual-luciferase reporter assay and chromatin immunoprecipitation (ChIP) were applied to analyze promoter activity to evaluate the possible regulatory mechanism of adipocyte-specific gene expression.
  
  
  RESULTS
  We found that this extreme hypoxia impaired osteogenic differentiation as indicated by the attenuation of alkaline phosphatase (ALP) activity and the reduced expression of osteogenic markers osteocalcin and osteopontin. Moreover, extreme hypoxia enhanced adipogenic differentiation, as indicated by the accumulation of lipid droplets and the expression of the adipocyte-specific genes leptin, LPL, CFD, PGAR and HIG2. In the extreme hypoxic conditions (0.2% oxygen), the overexpression of CCAAT enhancer-binding proteins (C/EBPs), especially C/EBPδ, and HIF-1A upregulated the promoter activities of adipocyte-specific genes such as leptin, CFD, HIG2, LPL, PGAR. In the present study, peroxisome proliferator-activated receptor-gamma (PPARγ) exerted a negative effect on the differentiation of BMSCs into adipocytes.
  
  
  CONCLUSIONS
  In view of these findings, extreme hypoxia induced the adipogenic differentiation of BMSCs through HIF-1A and C/EBPs. These findings might provide clues regarding the roles of BMSCs in the cancer microenvironment.
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. jiangchen3701@163.com.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. jiangchen3701@163.com.|Hunan Key Laboratory of Nonresolving Inflammation and Cancer, Changsha, Hunan, 410013, China. jiangchen3701@163.com.
authors.forename: Chen
authors.initials: C
authors.lastname: Jiang
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. sunjun0828@csu.edu.cn.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. sunjun0828@csu.edu.cn.|Hunan Key Laboratory of Nonresolving Inflammation and Cancer, Changsha, Hunan, 410013, China. sunjun0828@csu.edu.cn.
authors.forename: Jun
authors.initials: J
authors.lastname: Sun
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. daiyafei136@126.com.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. daiyafei136@126.com.|Hunan Key Laboratory of Nonresolving Inflammation and Cancer, Changsha, Hunan, 410013, China. daiyafei136@126.com.
authors.forename: Yafei
authors.initials: Y
authors.lastname: Dai
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. caopengfei66@163.com.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. caopengfei66@163.com.|Hunan Key Laboratory of Nonresolving Inflammation and Cancer, Changsha, Hunan, 410013, China. caopengfei66@163.com.
authors.forename: Pengfei
authors.initials: P
authors.lastname: Cao
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. 124769836@qq.com.
authors.forename: Liyang
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. shuping@csu.edu.cn.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. shuping@csu.edu.cn.
authors.forename: Shuping
authors.initials: S
authors.lastname: Peng
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. zhouyanhong2004@hotmail.com.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. zhouyanhong2004@hotmail.com.
authors.forename: Yanhong
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. lgy@csu.edu.cn.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. lgy@csu.edu.cn.|Hunan Key Laboratory of Nonresolving Inflammation and Cancer, Changsha, Hunan, 410013, China. lgy@csu.edu.cn.
authors.forename: Guiyuan
authors.initials: G
authors.lastname: Li
authors.affiliation: Department of Thoracic Surgery, the Second Xiangya Hospital of Central South University, Changsha, Hunan, 410011, China. jingquntang@hotmail.com.
authors.forename: Jingqun
authors.initials: J
authors.lastname: Tang
authors.affiliation: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Key Laboratory of Carcinogenesis of Ministry of Health, Central South University, 110 Xiangya Road, Changsha, Hunan, 410078, China. xiangjj@csu.edu.cn.|Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, Hunan, 410013, China. xiangjj@csu.edu.cn.|Hunan Key Laboratory of Nonresolving Inflammation and Cancer, Changsha, Hunan, 410013, China. xiangjj@csu.edu.cn.
authors.forename: Juanjuan
authors.initials: J
authors.lastname: Xiang
mesh_terms: D017667:Adipocytes / Q000166:cytology / Q000378:metabolism; D050156:Adipogenesis; D000469:Alkaline Phosphatase / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism; D001854:Bone Marrow Cells / Q000166:cytology*; D051743:CCAAT-Enhancer-Binding Protein-delta / Q000235:genetics / Q000378:metabolism; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015687:Cell Hypoxia; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism; D000071063:Endoglin; D005786:Gene Expression Regulation; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism; D010012:Osteogenesis; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism
chemical_list: D015703:Antigens, CD; D022762:CCAAT-Enhancer-Binding Proteins; C497252:CEBPD protein, human; C064206:ENG protein, human; D000071063:Endoglin; C497442:HIF1A protein, human; D018960:Hyaluronan Receptors; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D034741:RNA, Small Interfering; D011956:Receptors, Cell Surface; D051743:CCAAT-Enhancer-Binding Protein-delta; D000469:Alkaline Phosphatase