title: Optimization of culture condition of human bone marrow stromal cells in terms of purification, proliferation, and pluripotency.
abstract: Human bone marrow stromal cells (hBMSCs) possess multilineage differentiation potential and play an important role in modern tissue engineering. However, the development of culture media to maintain hBMSCs in an undifferentiated, self-renewing state during their robust proliferation remains a challenge. We developed and tested modified growth medium [medium 1: epidermal growth factor (EGF), platelet-derived growth factor (PDGF), low glucose, 2% fetal calf serum (FCS)] on hBMSCs by comparing primary cell isolation, multipassage expansion, culture morphology, proliferation, and cellular phenotype, and performing an expression analysis of intrinsic-regulated genes to other two media. Cell morphology, proliferation, and phenotype varied among the media, while cells cultured in medium 1 displayed small, spindle-shaped morphology with the highest rate of growth capacities and the expected phenotype. RT-PCR analysis showed that medium 1 displayed the lowest expression levels of osteogenic genes, chondrogenic genes (osteonectin, runt-related transcription factor 2, cartilage oligo matrix protein, and SOX9), and adipogenic genes (lipoprotein lipase). The expression of another adipogenic gene, peroxisome proliferator-activator receptor-γ2, was higher in medium 1 but did not reach significance. In addition, hBMSCs expanded in medium 1 showed the highest expression ratio of self-renewing-related genes Krüppel-like factor 2 (KLF2) and KLF5. In conclusion, medium 1 allows for better expansion and pluripotency maintenance of hBMSCs and serves as a preferred alternative to traditional serum-containing media for research applications and future clinical use.
authors.affiliation: Department of Oral and Maxillofacial Surgery, First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, 310003, People's Republic of China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhou
authors.forename: Dan
authors.initials: D
authors.lastname: Yu
authors.forename: Huiyong
authors.initials: H
authors.lastname: Zhu
mesh_terms: D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D003470:Culture Media; D005434:Flow Cytometry; D006801:Humans; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D010982:Platelet-Derived Growth Factor; D039904:Pluripotent Stem Cells / Q000166:cytology*; D016133:Polymerase Chain Reaction
chemical_list: D003470:Culture Media; D051741:Kruppel-Like Transcription Factors; D010982:Platelet-Derived Growth Factor