Role of aTGF-beta Regulated Gene in human and mouse osteoblasts and skeleton

描述（由申请人提供）：TGF β在骨骼生长、成骨细胞（OB）分化以及牙周炎（成人牙齿脱落的主要原因）的发生和进展中起关键作用。尽管其在骨生物学和牙周组织重建中的作用，但TGF β在骨组织中作用的分子机制尚未完全了解。在本实验室研究雌激素（E2）和TGF β对OB的作用过程中，我们发现并表征了新的TGF β诱导早期基因-1（TIEG）作为Kruppel转录因子家族（KLF-10）的成员。OBs中的TIEG表达显示由E2、TGF β和BMP诱导。在过去的资助期间，我们发现TIEG蛋白通过转录抑制Smad 7和激活Smad 2激活TGF β/Smad通路，并调节其他重要的OB标记基因的表达。为了更好地理解TIEG在骨中的功能，我们已经产生了TIEG-null（TIEG-/-）小鼠，并且发现雌性而不是雄性具有较小和较弱的骨，相对于野生型同窝仔被分类为骨质减少表型。在过去的一年中，我们报道了颅骨来源的OB具有显著降低的矿化骨和支持破骨细胞生成的能力。这些OB的进一步表征显示Runx 2、osterix、碱性磷酸酶和其他重要OB标记基因的表达水平降低。最近，我们已经证明，TIEG是能够直接调节Runx 2的转录，Runx 2似乎是，至少部分，负责TIEG-/- OB矿化中观察到的缺陷。我们的初步研究表明，E2诱导TIEG的表达在雌激素受体（ER）亚型特异性的方式。最后，E2还诱导Runx 2在野生型OB中的表达，但在TIEG-/-OB中不诱导Runx 2的表达，这表明TIEG在介导E2在骨中的作用中具有重要作用。基于这些数据，我们假设TIEG的E2调节和Runx 2的后续TIEG调节至少部分地导致观察到的OB缺陷，并且可能涉及在TIEG-/-小鼠中观察到的性别特异性骨质减少表型。为了验证这一假设，我们计划确定：1）TIEG对Runx 2表达的调节对观察到的TIEG-/-OB缺陷的作用，2）E2对TIEG表达的调节对TIEG-/- OB表型的贡献，3）TIEG在介导E2激活OB中的Runx 2表达中的作用，（4）TIEG-/-小鼠性腺切除对骨骼表型的影响。这些研究的完成将有助于确定TIEG在OB功能以及骨骼发育和维护中的生物学作用。此外，这些研究应该为E2和TIEG调节Runx 2表达的机制及其对骨疾病（包括牙周炎和骨质疏松症）的贡献提供新的见解。

DESCRIPTION (provided by applicant): TGFb, plays a pivotal role in skeletal growth, osteoblast (OB) differentiation, as well as the onset and progression of periodontitis, a main cause for tooth loss in adults. In spite of its role in bone biology and periodontal tissue remodeling, the molecular mechanisms of action of TGFb in bone tissue are not fully understood. During this laboratory's investigations of estrogen (E2) and TGFb actions on OBs, we discovered and characterized the novel TGFb Iducible Early Gene-1 (TIEG) as a member of the Kruppel family of transcription factors (KLF-10). TIEG expression in OBs was shown to be induced by E2, TGFb, and BMPs. During the past funding period, we revealed that TIEG protein activates the TGFb/Smad pathway via transcriptional repression of Smad 7 and activation of Smad 2 and regulates the expression of other important OB marker genes. In order to better understand the function of TIEG in bone, we have generated TIEG-null (TIEG-/-) mice and found that the females, but not males, have smaller and weaker bones, classified as an osteopenic pheontype relative to wild-type littermates. In this past year, we reported that calvarial-derived OBs have a markedly reduced capacity to mineralize bone and to support osteoclastogenesis. Further characterization of these OBs revealed decreased expression levels of Runx2, osterix, alkaline phosphatase, and other important OB marker genes. Recently, we have demonstrated that TIEG is capable of directly regulating the transcription of Runx2 and that Runx2 appears to be, at least in part, responsible for the observed defects in TIEG-/- OB mineralization. Our preliminary studies have shown that E2 induces the expression of TIEG in an estrogen receptor (ER) isoform specific manner. Finally, E2 also induces the expression of Runx2 in wild-type OBs, but not in TIEG-/- OBs, suggesting an important role for TIEG in mediating E2 action in bone. Based on these data, it is our hypothesis that the E2 regulation of TIEG, and the subsequent TIEG regulation of Runx2, is at least in part responsible for the observed defects in OBs and could be involved in the gender specific osteopenic phenotype observed in TIEG-/- mice. In order to test this hypothesis, we plan to determine: 1) the role that TIEG regulation of Runx2 expression has on the observed defects in TIEG-/- OBs, 2) the contribution of E2 regulation of TIEG expression to the TIEG-/- OB phenotype, 3) the role of TIEG in mediating E2 activation of Runx2 expression in OB, and 4) the effects of gonadectomy of male and female TIEG-/- mice on the skeletal phenotype. The completion of these studies should help determine the biological role of TIEG in OB functions as well as skeletal development and maintenance. In addition, these studies should provide new insights into the mechanisms of E2 and TIEG regulation of Runx2 expression and their contribution to bone disease, including periodontitis and osteoporosis.