title: The influence of oxygen concentration on the extracellular matrix production of human nucleus pulposus cells during isolation-expansion process.
abstract: Nucleus pulposus (NP) cells locate in the center of avascular intervertebral discs, and thus have presumably adapted to a hypoxic environment. The purpose of this study was to investigate the influences of hypoxic condition, during isolation-expansion of human NP cells, on the cellular proliferation and extracellular matrix (ECM) synthesis in later three-dimensional cultures. Human NP tissues were obtained from patients who underwent lumbar disc surgeries. Immediately after retrieval, NP tissues from each patient were divided into two aliquots for in vitro cultivation either under classical normoxic (21% O2) or hypoxic (3.5% O2) condition. After isolation-expansion processes, microtissues of NP cells were formed and the analysis was performed after one-week culture. Experiments of pretreatment with TGF-β1 or lovastatin were designed to investigate if the isolation-expansion conditions affect the responsiveness to later exogenous treatments. Hypoxic isolation-expansion stimulated NP cell proliferation during monolayer culture. Hypoxia also upregulated mRNA levels of SOX9 and HIF-1α but downregulated type X collagen as well as improved aggrecan and type II collagen synthesis. Although TGF-β1 had no substantial effect, lovastatin pretreatment showed a greater enhancement on type II collagen expression in hypoxic group. Normoxia negatively affected the biochemical composition of regenerated ECM attributable to downregulation of SOX9 and HIF-1α, while hypoxia enhanced cellular proliferation, improved matrix production, and maintained a functional phenotype of NP cells. Hypoxic isolation-expansion of human NP cells is important to achieve better regenerative cells for later cultivation or cell transplantation. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1575-1582, 2017.
authors.affiliation: Department of Orthopedics, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, 10002, Taiwan.
authors.forename: Shu-Hua
authors.initials: SH
authors.lastname: Yang
authors.affiliation: Department of Orthopedics, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, 10002, Taiwan.|Institute of Biomedical Engineering, College of Medicine and Engineering, National Taiwan University, Taipei, 10031, Taiwan.
authors.forename: Ming-Hsiao
authors.initials: MH
authors.lastname: Hu
authors.affiliation: Department of Orthopedics, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, 10002, Taiwan.
authors.forename: Wan-Yu
authors.initials: WY
authors.lastname: Lo
authors.affiliation: Department of Orthopedics, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, 10002, Taiwan.
authors.forename: Yuan-Hui
authors.initials: YH
authors.lastname: Sun
authors.affiliation: Department of Orthopedics, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, 10002, Taiwan.|Department of Biomedical Engineering, Yuanpei University of Medical Technology, Hsinchu, 30015, Taiwan.|Department of Orthopedics, En Chu Kong Hospital, New Taipei City, 23702, Taiwan.
authors.forename: Chang-Chin
authors.initials: CC
authors.lastname: Wu
authors.affiliation: School of Dental Technology, College of Oral Medicine, Taipei Medical University, Taipei, 11031, Taiwan.
authors.forename: Kai-Chiang
authors.initials: KC
authors.lastname: Yang
mesh_terms: D000328:Adult; D018929:Cell Culture Techniques / Q000379:methods; D015687:Cell Hypoxia; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D002478:Cells, Cultured; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D005109:Extracellular Matrix / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D006909:Hydroxyproline / Q000235:genetics / Q000378:metabolism; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism; D000070614:Nucleus Pulposus / Q000166:cytology* / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism*; D055815:Young Adult
chemical_list: D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D003094:Collagen; D006909:Hydroxyproline; D010100:Oxygen
keywords: extracellular matrix; hypoxia; hypoxia inducible factor 1 alpha; isolation and expansion; nucleus pulposus cells