Regulation of Tendon Induction and Formation By TGFbeta Signaling

描述（由申请人提供）：功能性肌肉骨骼系统依赖于从肌肉到骨骼的力传递，因此需要肌肉、软骨和肌腱的协调发育。然而，对肌腱形成的细胞和分子方面知之甚少。阐明引导间充质细胞成为腱细胞的信号的进展在阐明塑造肌肉骨骼系统的组织和协调生长的早期事件中特别重要。此外，肌腱诱导活性对于改善肌腱损伤的治疗和从干细胞诱导肌腱形成的生物工程努力都具有相当大的临床重要性。最近的发现，Scleraxis（Scx）的bHLH转录因子是肌腱细胞和肌腱祖细胞的独特标志物，促进肌腱发育的研究和一些后续的研究涉及FGF信号在肌腱祖细胞的诱导。我们实验室的戏剧性结果现在证明了TGF β在肌腱诱导和分化中的作用更加深远。TGF β 2和TGF β 3基因双重突变的胚胎发育出没有肌腱的肢体。此外，在器官培养和组织培养实验中，TGF β信号传导充当Scx的有效诱导剂，显著超过类似实验中FGF信号传导诱导Scx的程度和强度。我们建议在三组不同的实验中探索TGF β在肌腱形成中的作用。第一个目标将解决所有肌腱组织依赖于TGF β信号传导的假设。将在TGF β 2和3突变体和TGF β受体（TGF β（RII））突变体中评估轴向和颅侧肌腱中的表型，其中所有TGF β信号传导被消除。我们将进一步研究这些突变体中肌腱表型的发生，以建立TGF β信号传导对肌腱形成至关重要的阶段。第二个目标是解决TGF β信号传导对于肌腱形成的后期方面也是必不可少的这一假设。TGF β 1的诱导消除将使得能够探索TGF β 1信号传导的后期作用，并确定肌腱是从早期祖细胞的定向池产生的还是通过TGF β 1信号传导持续募集肌腱细胞来支持肌腱生长。最后，将在组织培养和体内探索TGF β信号传导诱导Scx的机制;将比较TGF β和FGF信号传导的活性，并使用特异性拮抗剂和突变组织的组合探索这两种信号传导级联之间协同或相互依赖的可能性。此外，将使用逆转录病毒系统在子宫内应用TGF β，以评估由TGF β诱导的细胞的能力，从而超过Scx的表达并引起异位或扩张肌腱的形成。

DESCRIPTION (provided by applicant): A functional musculoskeletal system depends upon the transmission of force from muscle to bone, thus requiring the coordinated development of muscle, cartilage and tendon. Little however, is known about the cellular and molecular aspects of tendon formation. Progress unraveling the signals that direct mesenchymal cells to become tenocytes is of particular importance in elucidation of the early events that shape the organization and coordinated growth of the musculoskeletal system. Moreover, a tendon inducing activity may be of considerable clinical importance both for improvements in the treatment of tendon injuries and in bioengineering efforts to induce tendon formation from stem cells. The recent finding that Scleraxis (Scx) a bHLH transcription factor is a unique marker of tenocytes and tendon progenitors facilitated studies of tendon development and a number of subsequent studies have implicated FGF signaling in the induction of tendon progenitors. Dramatic results in our laboratory now demonstrate an even more profound role for TGF( signaling in tendon induction and differentiation. Embryos doubly mutant for the TGF(2 and TGF(3 genes develop limbs in which no tendons can be detected. Moreover, TGF( signaling acts as a potent inducer of Scx, in organ culture and in tissue culture experiments, significantly exceeding the extent and intensity of Scx induction by FGF signaling in similar experiments. We propose to explore TGF( role in tendon formation in three separate groups of experiments. The first aim will address the hypothesis that all tendon tissues depend on TGF( signaling. The phenotype in axial and cranial tendons will be evaluated in TGF(2 & 3 mutants and in mutants for a TGF( receptor (TGF(RII), in which all TGF( signaling is eliminated. We will further study the onset of the tendon phenotype in these mutants to establish the stage in which TGF( signaling is essential for tendon formation. The second aim will be to address the hypothesis that TGF( signaling is essential for later aspects of tendon formation as well. Inducible elimination of TGF(RII will enable exploration of later roles for TGF( signaling and establish if tendons are generated from a committed pool of early progenitors or through continuous recruitment of tendon cells by TGF( signaling to support tendon growth. Finally, the mechanism of Scx induction by TGF( signaling will be explored in tissue culture and in vivo; the activities of TGF( and FGF signaling will be compared and the possibilities of synergism or interdependence between these two signaling cascades will be explored using a combination of specific antagonists and mutant tissues. In addition, TGF( will be applied in utero using a retroviral system to evaluate the capacity of the cells induced by TGF( signaling to go beyond expression of Scx and cause the formation of ectopic or expanded tendons.