• Ming Pei, Mark Shoukry, Jingting Li, Scott D Daffner, John C France, and Sanford E Emery.
• Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, WV 26506, USA. mpei@hsc.wvu.edu
• Spine. 2012 Aug 15;37(18):1538-47.

Study DesignTwo 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.ObjectiveModulating the in vitro microenvironment facilitates SDSC-based NP tissue regeneration.Summary Of Background DataAutologous cell therapy is a promising approach for NP regeneration. Current in vitro expansion in monolayer results in cell dedifferentiation.MethodsIn 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.ResultsDECM 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.ConclusionThe in vitro microenvironment can be modulated by low oxygen and tissue-specific cell-based DECM to facilitate NP tissue regeneration.

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