骨髓间充质干细胞与椎间盘髓核细胞在表型表达的相互效应及机制研究

Stem cells offer promise for intervertebral disc (IVD) repair and regeneration because they are easily isolated and expanded, and can differentiate into several mesenchymal tissues. However, the cellular biological mechanisms of interactions between stem cells and degenerative IVD still remain uncertain. The current lack of specific marker(s) for IVD cells limits the ability to meaningfully explore the differentiation of stem cells to an IVD phenotype. Moreover, there have been no studies investigating the proper candidate stem cells specifically for disc regeneration. The purpose of this study is thus to begin to investigate the cellular biological interactions between mesenchymal stem cells and degenerative nucleus pulposus cells and its possible signal pathway mechanism. Homologous human mesenchymal stem cells (MSCs) and degenerative nucleus pulposus cells (NPCs) will be harvested with informed consent from bone marrow, adipose and nucleus pulposus of forty patients undergoing lumbar spine surgery(grouped with age and IVD Pfirrmann's scale) . MSCs will be co-cultured with indirect (Transwell system) and direct cell-cell contact with homologous degenerative NPCs for 14 days. Functinoal cell factors(TGF-β1, IGF-1, BMP-7) and changes of matrix-associated genes and protein (collagen type I,II,III, aggrecan, SOX9 and Versican)of MSCs, ADSCs and degenerative NPCs are to be assessed by quantitative real-time PCR and ELISA in 3rd ,7th and 14th day. We are trying to answer two questions of biological treatment for intervertebral disc degenerative diseases by this comparative study: Does co-culture effectively induce stem cell differentiation toward nucleus pulposus cell phenotype? Does the biological funtion of degenerative nucleus pulposus cells also up-regulated by extrogenic stem cells

应用干细胞修复椎间盘退变已经成为脊柱外科基础研究的重要发展方向。但目前对外源性干细胞与退变髓核发生相互作用的细胞生物学机制缺乏认识，也是其深入研究及临床应用的瓶颈。既缺乏促进干细胞向类髓核细胞定向分化的明确手段，也不清楚体内退变髓核细胞的生物学功能是否受到干细胞的积极影响。本研究拟在实施椎间盘手术同时，采集同源人骨髓与髓核组织，体外分离骨髓基质干细胞与髓核细胞，分别以直接及非直接接触的方式进行共培养，观察共培养前后干细胞向类髓核细胞分化表型及髓核组织功能性细胞因子表达的变化，及其可能作用的细胞信号通路；并探讨供体年龄及受体髓核退变程度对于干细胞修复椎间盘退变的生物学效应的影响。研究基质干细胞与退变髓核细胞相互作用的细胞生物学机制，希望以此回答两个问题：骨髓基质干细胞与退变髓核细胞共培养能否达到"近朱者赤"的定向诱导分化作用？退变髓核细胞功能是否受到干细胞的上调作用以及其可能的调节机制？

应用干细胞修复椎间盘退变已经成为脊柱外科基础研究的重要发展方向。但目前对外源性干细胞与退变髓核发生相互作用的细胞生物学机制缺乏认识，也是其深入研究及临床应用的瓶颈。既缺乏促进干细胞向类髓核细胞定向分化的明确手段，也不清楚体内退变髓核细胞的生物学功能是否受到干细胞的积极影响。本研究拟在实施椎间盘手术同时，采集同源人骨髓与髓核组织，体外分离骨髓基质干细胞与髓核细胞，分别以直接及非直接接触的方式进行共培养，观察共培养前后干细胞向类髓核细胞分化表型及髓核组织功能性细胞因子表达的变化，及其可能作用的细胞信号通路；并探讨供体年龄及受体髓核退变程度对于干细胞修复椎间盘退变的生物学效应的影响。研究基质干细胞与退变髓核细胞相互作用的细胞生物学机制，希望以此回答两个问题：骨髓基质干细胞与退变髓核细胞共培养能否达到“近朱者赤”的定向诱导分化作用？退变髓核细胞功能是否受到干细胞的上调作用以及其可能的调节机制？