title: Modulation of in vitro microenvironment facilitates synovium-derived stem cell-based nucleus pulposus tissue regeneration.
abstract: METHODS
  Two experiments were conducted. Experiment 1 evaluated the effect of 3 kinds of decellularized extracellular matrices (DECMs) deposited by synovium-derived stem cells (SDSCs) and/or nucleus pulposus cells (NPCs) on SDSC expansion and NP lineage differentiation. Experiment 2 evaluated the effect of DECM deposited by SDSCs on NPC expansion and redifferentiation capacity. In both experiments, hypoxia was evaluated in DECM preparation and pellet culture.
  
  
  OBJECTIVE
  Modulating the in vitro microenvironment facilitates SDSC-based NP tissue regeneration.
  
  
  BACKGROUND
  Autologous cell therapy is a promising approach for NP regeneration. Current in vitro expansion in monolayer results in cell dedifferentiation.
  
  
  METHODS
  In Experiment 1, passage 3 SDSCs were expanded for 1 passage on DECM deposited by NPCs, SDSCs, or NPCs combined with SDSCs (50:50); DECM was prepared under either normoxia (21% O2) or hypoxia (5% O2). Expanded SDSCs were then cultured in a serum-free chondrogenic medium in hypoxia for 14 days. In Experiment 2, passage 2 NPCs were expanded for 1 passage on DECM deposited by SDSCs; DECM was prepared under either normoxia or hypoxia. Expanded NPCs were cultured in a serum-free chondrogenic medium under either hypoxia or normoxia for 14 days. Cell expansion on plastic flasks served as a control in both experiments. Fourteen-day pellets were evaluated for chondrogenesis using histology, immunostaining, biochemistry, and real-time polymerase chain reaction.
  
  
  RESULTS
  DECM deposited by NPCs combined with SDSCs effectively enhanced expanded SDSC viability and guided SDSC differentiation toward an NP lineage; this effect is comparable with DECM deposited by SDSCs but higher than that deposited by NPCs. DECM prepared under normoxia favored SDSC viability and NP lineage differentiation whereas DECM prepared under hypoxia benefited NPC viability and redifferentiation. Low oxygen in a pellet culture system enhanced NPC viability and redifferentiation.
  
  
  CONCLUSIONS
  The in vitro microenvironment can be modulated by low oxygen and tissue-specific cell-based DECM to facilitate NP tissue regeneration.
authors.affiliation: Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, WV 26506, USA. mpei@hsc.wvu.edu
authors.forename: Ming
authors.initials: M
authors.lastname: Pei
authors.forename: Mark
authors.initials: M
authors.lastname: Shoukry
authors.forename: Jingting
authors.initials: J
authors.lastname: Li
authors.forename: Scott D
authors.initials: SD
authors.lastname: Daffner
authors.forename: John C
authors.initials: JC
authors.lastname: France
authors.forename: Sanford E
authors.initials: SE
authors.lastname: Emery
mesh_terms: D000818:Animals; D054337:Cell Dedifferentiation / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D015687:Cell Hypoxia; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D060833:Cellular Microenvironment / Q000187:drug effects; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D006025:Glycosaminoglycans / Q000378:metabolism; D007150:Immunohistochemistry; D007403:Intervertebral Disc / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D010100:Oxygen / Q000378:metabolism / Q000494:pharmacology; D012038:Regeneration / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D013552:Swine; D013583:Synovial Membrane / Q000166:cytology* / Q000378:metabolism; D013997:Time Factors; D023822:Tissue Engineering / Q000379:methods*
chemical_list: D006025:Glycosaminoglycans; C013786:A73025; D003094:Collagen; D010100:Oxygen