Cell-specific response to mechanical stress and regulatory mechanism of differentiation : ligament/tendon cells versus osteoblasts

本实验以本实验室建立的韧带细胞系PDL-L2为材料，证明：（1）韧带细胞在成骨细胞分化的早期阶段具有成骨细胞的特征。(2)尽管表达Runx 2（成骨细胞分化的关键转录因子），韧带细胞在体外不会产生矿化结节（3）这是由于同源框蛋白Msx 2抑制Runx 2活性。(4)Msx 2的过表达抑制成骨细胞的基质矿化（5）相反，Msx 2的敲低导致韧带细胞的基质矿化（6）机械应力加速并增强成骨细胞的基质矿化，但对韧带细胞没有影响。(7)这可能是由于这两种细胞类型之间整合素的差异。此外，我们鉴定了PIASxβ，首先鉴定为E3连接酶，作为矿化的关键调节因子。观察结果包括（8）对于基质矿化，在成骨细胞分化的早期阶段需要PIASxβ的瞬时增加，这也是机械应力诱导的矿化所需的。(9)PIASxβ的这种作用是SUMOYLATION依赖性的，并且通过激活Osterix，Osterix是位于Runx 2下游的成骨细胞分化的另一个关键转录因子。因此，PIASxβ是Runx 2和osterix在成骨细胞分化和矿化中的信号分子。(10)Osterix启动子具有自身调节作用，可能在成骨细胞控制基质矿化中发挥重要作用。

Using PDL-L2, a ligament cell line established in our laboratory, we have demonstrated the followings:(1) Ligament cells have characteristics of osteoblastic cells at the early stage of osteoblast differentiation.(2) Ligament cells do not produce mineralized nodule in vitro in spite of expressing Runx2, a key transcription factor for osteoblast differentiation(3) This is due to suppression of Runx2 activity by homeobox protein Msx2.(4) Overexpression of Msx2 suppresses matrix mineralization by osteoblasts(5) Conversely, knockdown of Msx2 causes matrix mineralization by ligament cells(6) Mechanical stress accelerates and enhances matrix mineralization by osteoblasts, but without effect on ligament cells.(7) This is likely due to difference in integrins between these two cell types.Furthermore, we identified PIASxβ, first identified as an E3 ligase, as a key regulator for mineralization. The observations includes(8) For matrix mineralization, a transient increase of PIASxβ is required at an early stage of osteoblast differentiation and this is also required for mechanical stress-induced mineralization.(9) This effect of PIASxβ is sumoylation-dependent and through activation of osterix, another key transcription factor for osteoblast differentiation located at the immediate downstream of Runx2. Thus PIASxβ is a signaling molecule between Runx2 and osterix in osteoblast differentiation and mineralization.(10) Osterix promoter is auto-regulated by itself and this may play an important role in controlling matrix mineralization by osteoblasts.