Qi W N, Scully S P
Division of Orthopaedics, Duke University Medical Center, Durham, North Carolina 27710, USA.
Exp Cell Res. 1998 May 25;241(1):142-50. doi: 10.1006/excr.1998.4032.
The in vivo role of the extracellular matrix and the manner in which it interfaces with soluble regulators remains unknown. This study reports the modulation by extracellular type II collagen of TGF-beta 1-stimulated DNA synthesis, proteoglycan synthesis, and mRNA expression for alpha 1(II) procollagen and aggrecan core protein in the adult articular chondrocyte. Bovine chondrocytes were isolated and resuspended in alginate beads which contained increasing amounts of type II collagen from 0 to 1.5% (w/v). Cultures were maintained for 7 days in basal, DMEM, TGF-beta 1 (10 ng/ml), or FBS (10%) supplemented medium. DNA and proteoglycan synthesis were determined by radiotracer incorporation. The relative amounts of mRNA were analyzed by Northern blot analysis. Exogenous collagen increased DNA synthesis in all culture conditions beginning at concentrations of 0.75% (w/v). We observed that extracellular type II collagen augments both TGF-beta 1 stimulated increases of aggrecan gene expression up to 400% and alpha 1(II) procollagen gene expression up to 180% in a dose-dependent fashion. This is distinct from cultures which were either basal or FBS supplemented medium which lacked a dose-dependent change in aggrecan gene expression and demonstrated a decrease in alpha 1(II) procollagen gene expression. Exogenous collagen above 0.75% (w/v) increased proteoglycan synthesis significantly in FBS and TGF-beta 1-stimulated cultures but not in basal cultures. We have demonstrated that the alterations in gene expression that occur in response to TGF-beta 1 are modulated by extracellular type II collagen. This modulation is possible through both transcriptional and posttranscriptional regulatory mechanisms.
细胞外基质在体内的作用以及它与可溶性调节因子相互作用的方式仍然未知。本研究报道了成年关节软骨细胞中细胞外II型胶原对TGF-β1刺激的DNA合成、蛋白聚糖合成以及α1(II)前胶原和聚集蛋白聚糖核心蛋白mRNA表达的调节作用。分离牛软骨细胞并将其重悬于含有0至1.5%（w/v）递增浓度II型胶原的藻酸盐珠中。将培养物在基础培养基、DMEM、TGF-β1（10 ng/ml）或FBS（10%）补充培养基中维持7天。通过放射性示踪剂掺入法测定DNA和蛋白聚糖合成。通过Northern印迹分析来分析mRNA的相对含量。从浓度为0.75%（w/v）开始，外源性胶原在所有培养条件下均增加了DNA合成。我们观察到，细胞外II型胶原以剂量依赖性方式增强了TGF-β1刺激的聚集蛋白聚糖基因表达增加高达400%以及α1(II)前胶原基因表达增加高达180%。这与基础培养基或补充FBS的培养基培养不同，后者聚集蛋白聚糖基因表达缺乏剂量依赖性变化，并且α1(II)前胶原基因表达呈下降趋势。在FBS和TGF-β1刺激的培养物中，高于0.75%（w/v）的外源性胶原显著增加了蛋白聚糖合成，但在基础培养物中未增加。我们已经证明，响应TGF-β1而发生的基因表达改变受到细胞外II型胶原的调节。这种调节可能通过转录和转录后调控机制实现。
