Expression analysis of human adipose-derived stem cells during in vitro differentiation to an adipocyte lineage

BMC Medical Genomics

J. Peter Rubin⁴  Sandeep Kathju⁴  William A. LaFramboise¹  Kacey G. Marra⁴  Sudheer Ravuri³  Brian J. Philips³  Fang Liu³  Shelley Des Etages²  Phillip H. Gallo³  J. Michael Krill-Burger¹  Latha Satish⁴ 

[1] Department of Pathology, University of Pittsburgh, Pittsburgh, PA, USA;Connecticut College, Department of Biology, New London, CT, USA;Department of Plastic Surgery, University of Pittsburgh Medical Center, 3550 Terrace Street, 6B Scaife Hall, Pittsburgh, 15261, PA, USA;McGowan Institute for Regenerative Medicine, Pittsburgh, PA, USA

关键词: Adipogenesis;    Stromal vascular fraction;    Gene expression;    Transcriptome;    Adipose-derived stem cells;    Microarray;   

Others  :  1223016
DOI  :  10.1186/s12920-015-0119-8

 received in 2014-09-11, accepted in 2015-07-10,  发布年份 2015

【 摘 要 】

Background

Adipose tissue-derived stromal stem cells (ASCs) represent a promising regenerative resource for soft tissue reconstruction. Although autologous grafting of whole fat has long been practiced, a major clinical limitation of this technique is inconsistent long-term graft retention. To understand the changes in cell function during the transition of ASCs into fully mature fat cells, we compared the transcriptome profiles of cultured undifferentiated human primary ASCs under conditions leading to acquisition of a mature adipocyte phenotype.

Methods

Microarray analysis was performed on total RNA extracted from separate ACS isolates of six human adult females before and after 7 days (7 days: early stage) and 21 days (21 days: late stage) of adipocyte differentiation in vitro. Differential gene expression profiles were determined using Partek Genomics Suite Version 6.4 for analysis of variance (ANOVA) based on time in culture. We also performed unsupervised hierarchical clustering to test for gene expression patterns among the three cell populations. Ingenuity Pathway Analysis was used to determine biologically significant networks and canonical pathways relevant to adipogenesis.

Results

Cells at each stage showed remarkable intra-group consistency of expression profiles while abundant differences were detected across stages and groups. More than 14,000 transcripts were significantly altered during differentiation while ~6000 transcripts were affected between 7 days and 21 days cultures. Setting a cutoff of +/-two-fold change, 1350 transcripts were elevated while 2929 genes were significantly decreased by 7 days. Comparison of early and late stage cultures revealed increased expression of 1107 transcripts while 606 genes showed significantly reduced expression. In addition to confirming differential expression of known markers of adipogenesis (e.g., FABP4, ADIPOQ, PLIN4), multiple genes and signaling pathways not previously known to be involved in regulating adipogenesis were identified (e.g. POSTN, PPP1R1A, FGF11) as potential novel mediators of adipogenesis. Quantitative RT-PCR validated the microarray results.

Conclusions

ASC maturation into an adipocyte phenotype proceeds from a gene expression program that involves thousands of genes. This is the first study to compare mRNA expression profiles during early and late stage adipogenesis using cultured human primary ASCs from multiple patients.

【 授权许可】

   
2015 Satish et al.

【 图 表 】

Fig. 1.

Fig. 2.

Fig. 3.

Fig. 4.

【 参考文献 】

• [1]Zuk PA, Zhu M, Mizuno H, Huang J, Futrell JW, Katz AJ, Benhaim P, Lorenz HP, Hedrick MH: Multilineage cells from human adipose tissue: implications for cell-based therapies. Tissue Eng 2001, 7:211-228.
• [2]Yoshimura K, Sato K, Aoi N, Kurita M, Inoue K, Suga H, Eto H, Kato H, Hirohi T, Harii K: Cell-assisted lipotransfer for facial lipoatrophy: efficacy of clinical use of adipose-derived stem cells. Dermatologic Surg 2008, 34:1178-1185.
• [3]Yoshimura K, Asano Y, Aoi N, Kurita M, Oshima Y, Sato K, Inoue K, Suga H, Eto H, Kato H, Harii K: Progenitor-enriched adipose tissue transplantation as rescue for breast implant complications. Breast J 2010, 16:169-175.
• [4]Matsumoto D, Sato K, Gonda K, Takaki Y, Shigeura T, Sato T, Aiba-Kojima E, Lizuka F, Inoue K, Suga H, Yoshimura K: Cell-assisted lipotransfer: Supportive use of human adipose-derived cells for soft tissue augmentation with lipoinjection. Tissue Eng 2006, 12:3375-3382.
• [5]Masuda T, Furue M, Matsuda T: Novel strategy for soft tissue augmentation based on transplantation of fragmented omentum and preadipocytes. Tissue Eng 2004, 10:1672-1683.
• [6]Moseley TA, Zhu M, Hedrick MH: Adipose-derived stem and progenitor cells as fillers in plastic and reconstructive surgery. Plast Reconstr Surg 2006, 118:121S-128S.
• [7]Todaro GJ, Green H: Quantitative studies of the growth of mouse embryo cells in culture and their development into established lines. J Cell Biol 1963, 17:299-313.
• [8]Burton GR, Guan Y, Nagarajan R, McGehee RE Jr: Microarray analysis of gene expression during early adipocyte differentiation. Gene 2002, 293:21-31.
• [9]Gerhold DL, Liu F, Jiang G, Li Z, Xu J, Lu M, Sachs JR, Bagchi A, Fridman A, Holdert DJ, Doebber TW, Berger J, Elbrecht A, Moller DE, Zhang BB: Gene expression profile of adipocyte differentiation and its regulation by peroxisome proliferator-activated receptor-gamma agonists. Endocrinology 2002, 143:2106-2118.
• [10]Ntambi JM, Young-Cheul K: Adipocyte differentiation and gene expression. J Nutr 2000, 130:3122S-3126S.
• [11]Christy RJ, Yang VW, Ntambi JM, Geiman DE, Landschulz WH, Friedman AD, Nakabeppu Y, Kelly TJ, Lane MD: Differentiation-induced gene expression in 3 T3-L1 preadipocytes: CCAAT/enhancer binding protein interacts with and activates the promoters of two adipocyte-specific genes. Genes Dev 1989, 3:1323-1335.
• [12]Soukas A, Socci ND, Saatkamp BD, Novelli S, Friedman JM: Distinct transcriptional profiles of adipogenesis in vivo and in vitro. J Biol Chem 2001, 276:34167-34174.
• [13]Lopez IP, Marti A, Milagro FI, Zulet MA, Moreno-Aliaga MJ, Martinez JA, De Miguel C: DNA microarray analysis of gene differentially expressed in diet-induced (Cafeteria) obese rats. Obes Res 2003, 11:188-194.
• [14]Labrecque B, Mathieu O, Bordignon V, Murphy BD, Palin MF: Identification of differentially expressed genes in a porcine in vivo model of adipogenesis using suppression subtractive hybridization. Comp Biochem Physiol Part D genomics Proteomics 2009, 4:32-44.
• [15]Zuk PA, Zhu M, Ashjian P, De Ugarte DA, Huang JI, Mizuno H, Alfonso ZC, Frazer JK, Benhaim P, Hedrick MH: Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell 2002, 13:4279-4295.
• [16]Urs S, Smith C, Campbell B, Saxton AM, Taylor J, Zhang B, Snoddy J, Jones Voy B, Moustaid-Moussa N: Gene expression profiling in human preadipocytes and adipocytes by microarray analysis. J Nutr 2004, 134:762-770.
• [17]DeLany JP, Floyd ZE, Zvonic S, Smith A, Gravois A, Reiners E, Wu X, Kilroy G, Lefevre M, Gimble JM: Proteomic analysis of primary cultures of human adipose-derived stem cells: modulation by adipogenesis. Mol Cell Proteomics 2005, 4:731-740.
• [18]Monaco E, Bionaz M, Rodriguez-Zas S, Hurley WL, Wheeler MB: Transcriptomics comparison between porcine adipose and bone marrow mesenchymal stem cells during in vitro osteogenic and adipogenic differentiation. PLoS One 2012, 7(3):e32481.
• [19]Yu G, Wu X, Dietrich MA, Polk P, Scott LK, Ptitsyn AA, Gimble JM: Yield and characterization of subcutaneous human adipose-derived stem cells by flow cytometric and adipogenic mRNA analyzes. Cytotherapy 2010, 12:538-546.
• [20]Archer KJ, Dumur CI, Joel SE, Ramakrishnan V: Assessing quality of hydridized RNA in Affymetrix Gene Chip experiments. Biostatistics 2006, 7:198-212.
• [21]Satish L, LaFramboise WA, Johnson S, Vi L, Njarlangattil A, Raykha C, Krill-Burger JM, Gallo PH, O’gorman DB, Gan BS, Baratz ME, Ehrlich GD, Kathju S: Fibroblasts from phenotypically normal palmar fascia exhibit molecular profiles highly similar to fibroblasts from active disease in Dupuytren’s contracture. BMC Med Genomics 2012, 5:15. BioMed Central Full Text
• [22]Li H, Zimmerlin L, Marra KG, Donnenberg VS, Donnenberg AD, Rubin JP: Adipogenic potential of adipose stem cell subpopulations. Plast Reconstr Surg 2011, 128:663-672.
• [23]Parker GE, Pederson BA, Obayashi M, Schroeder JM, Harris RA, Roach PJ: Gene expression profiling of mice with genetically modified muscle glycogen content. Biochem J 2006, 395:137-145.
• [24]Jiang L, Brackeva B, Ling Z, Kramer G, Aerts JM, Schuit F, Keymeulen B, Pipeleers D, Gorus F, Martens GA: Potential of protein phosphatase inhibitor 1 as biomarker of pancreatic β-cell injury in vitro and in vivo. Diabetes 2013, 62:2683-2688.
• [25]Hutley L, Shurety W, Newell F, McGeary R, Pelton N, Grant J, Herington A, Cameron D, Whitehead J, Prins J: Fibroblast growth factor 1: a key regulator of human adipogenesis. Diabetes 2004, 53:3097-3106.
• [26]Jonker JW, Suh JM, Atkins AR, Ahmadian M, Li P, Whyte J, He M, Juguilon H, Yin Y-Q, Phillips CT, Yu RT, Olefsky JM, Henry RR, Downes M, Evans RM: A PPARγ-FGF1 axis is required for adaptive adipose remodeling and metabolic homeostasis. Nature 2012, 485:391-394.
• [27]Luo Y, Cai J, Liu Y, Xue H, Chrest FJ, Wersto RP, Rao M: Microarray analysis of selected genes in neural stem and progenitor cells. J Neurochem 2002, 83:1481-1497.
• [28]Hennessey JA, Wei EQ, Pitt GS: Fibroblast growth factor homologous factors modulate cardiac calcium channels. Circ Res 2013, 113:381-388.
• [29]Zhuang Z, Jian P, Longjiang L, Bo H, Wenlin X: Oral cancer cells with different potential of lymphatic metastasis displayed distinct biologic behaviors and gene expression profiles. J Oral Pathol Med 2010, 39:168-175.
• [30]Wang X, Hinson ER, Cresswell P: The interferon-inducible protein viperin inhibits influenza virus release by perturbing lipid rafts. Cell Host Microbe 2007, 2:96-105.
• [31]Hinson ER, Cresswell P: The antiviral protein, viperin, localizes to lipid droplets via its N-terminal amphipathic alpha-helix. Proc Natl Acad Sci U S A 2009, 106:20452-20457.
• [32]Dogan A, Lasch P, Neuschl C, Millrose MK, Alberts R, Schughart K, Naumann D, Brockmann GA: ATR-FTIR spectroscopy reveals genomic loci regulating the tissue response in high fat diet fed BXD recombinant inbred mouse strains. BMC Genomics 2013, 14:386. BioMed Central Full Text
• [33]Kudo A: Periostin in fibrillogenesis for tissue regeneration: periostin actions inside and outside the cell. Cell Mol Life Sci 2011, 68:3201-3207.
• [34]Norris RA, Moreno-Rodriguez R, Hoffman S, Markwald RR: The many facets of the matricellular protein periostin during cardiac development, remodeling, and pathophysiology. J Cell Commun Signal 2009, 3:275-286.
• [35]Ruan K, Bao S, Ouyang G: The multifaceted role of periostin in tumorigenesis. Cell Mol Life Sci 2009, 66:2219-2230.
• [36]Vi L, Feng L, Zhu RD, Wu Y, Satish L, Gan BS, O'Gorman DB: Periostin differentially induces proliferation, contraction and apoptosis of primary Dupuytren's disease and adjacent palmar fascia cells. Exp Cell Res 2009, 315:3574-3586.