title: Retention of the stemness of mouse adipose-derived stem cells by their expansion on human bone marrow stromal cell-derived extracellular matrix.
abstract: Mesenchymal stem cells (MSCs) usually lose their stemness during in vitro expansion as they are deprived of their niche environment. Cell-extracellular matrix (ECM) interaction is known to play important roles in preserving the stemness of the cells in their stem cell niche environment. Previously, coating with bone marrow MSC (BMSC)-derived ECM was found able to maintain the differentiation potential of in vitro cultured MSCs. This study aimed to determine if this ECM coating could also maintain the stemness of cultured murine adipose-derived stem cells (ASCs) using a regular culture flask as a control. Cells were expanded in ECM-coated and ECM-noncoated flasks for two and four passages and then harvested for various analyses. The results showed that ASCs exhibited fibroblast-like spindle morphology in ECM-coated flasks, whereas ASCs gradually spread and enlarged in the ECM-noncoated flasks. After three and five passages, both groups of cells exhibited similar cytokinetics in the MSC culture medium (MesenPRO RS™ Medium). However, when cultured in Dulbecco's modified Eagles medium (DMEM) plus 10% fetal bovine serum, coating group cells exhibited more potent proliferation than control group cells with a significant difference in both passages 3 and 5 (p<0.01). When seeded at low density (500 cells/10-cm dish), coating group cells formed significantly more and larger sized cell colonies than control group cells with significant difference in cell colony numbers between two groups (p<0.05). In addition, coated colony cells were much smaller and more compactly arranged compared to control colony cells. Furthermore, ASCs expanded in coated flasks exhibited greater potentials for adipogenic, osteogenic, and chondrogenic differentiations than the cells expanded in regular flasks. Quantitatively, the Oil Red O staining area, Alizarin staining area, and Toluidine Blue staining area were all significantly larger than the respective staining areas of control cells (p<0.05). Real-time polymerase chain reaction also revealed significantly higher gene expression levels of peroxisome proliferator-activated receptor gamma (PPARγ), adipocyte protein 2 (aP2), CCAAT/enhancer-binding protein (C/EBP), Runx2, osteocalcin, Sox9, collagen II, and aggrecan in ECM-coated group cells than in control group cells (p<0.05). Collectively, these results suggest that human BMSC decellular ECM coating helps to preserve the stemness of cultured murine ASCs.
authors.affiliation: 1Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China.
authors.forename: Yao
authors.initials: Y
authors.lastname: Xiong
authors.affiliation: 2Department of Anatomy and Neurobiology, Tongji University School of Medicine, Shanghai, People's Republic of China.
authors.forename: Jing
authors.initials: J
authors.lastname: He
authors.affiliation: 1Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China.|3National Tissue Engineering Center of China, Shanghai, People's Republic of China.
authors.forename: Wenjie
authors.initials: W
authors.lastname: Zhang
authors.affiliation: 1Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China.|3National Tissue Engineering Center of China, Shanghai, People's Republic of China.
authors.forename: Guangdong
authors.initials: G
authors.lastname: Zhou
authors.affiliation: 1Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China.|3National Tissue Engineering Center of China, Shanghai, People's Republic of China.
authors.forename: Yilin
authors.initials: Y
authors.lastname: Cao
authors.affiliation: 1Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China.|3National Tissue Engineering Center of China, Shanghai, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
mesh_terms: D053687:Adult Stem Cells / Q000166:cytology* / Q000187:drug effects; D053674:Aggrecans / Q000096:biosynthesis / Q000235:genetics; D000818:Animals; D001854:Bone Marrow Cells / Q000737:chemistry; D022762:CCAAT-Enhancer-Binding Proteins / Q000096:biosynthesis / Q000235:genetics; D002452:Cell Count; D018929:Cell Culture Techniques; D048430:Cell Shape; D020219:Chondrogenesis / Q000187:drug effects; D024043:Collagen Type II / Q000096:biosynthesis / Q000235:genetics; D003114:Colony-Forming Units Assay; D050659:Core Binding Factor Alpha 1 Subunit / Q000096:biosynthesis / Q000235:genetics; D003470:Culture Media / Q000494:pharmacology; D005109:Extracellular Matrix; D050556:Fatty Acid-Binding Proteins / Q000096:biosynthesis / Q000235:genetics; D005260:Female; D020869:Gene Expression Profiling; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects; D051379:Mice; D008810:Mice, Inbred C57BL; D039902:Multipotent Stem Cells / Q000166:cytology* / Q000187:drug effects; D015675:Osteocalcin / Q000096:biosynthesis / Q000235:genetics; D010012:Osteogenesis / Q000187:drug effects; D047495:PPAR gamma / Q000096:biosynthesis / Q000235:genetics; D055758:SOX9 Transcription Factor / Q000096:biosynthesis / Q000235:genetics; D017154:Stromal Cells / Q000737:chemistry*; D050151:Subcutaneous Fat / Q000166:cytology*
chemical_list: C508320:Acan protein, mouse; D053674:Aggrecans; D022762:CCAAT-Enhancer-Binding Proteins; C530718:CEBPA protein, mouse; D024043:Collagen Type II; D050659:Core Binding Factor Alpha 1 Subunit; D003470:Culture Media; C493411:Fabp4 protein, mouse; D050556:Fatty Acid-Binding Proteins; D047495:PPAR gamma; C493716:Runx2 protein, mouse; D055758:SOX9 Transcription Factor; C527933:Sox9 protein, mouse; D015675:Osteocalcin