Regulation of T Cell TNF Production

说明书(申请人提供)：雌激素(E2)具有多样性和复杂性 对T细胞功能的调节作用可能与 绝经后骨质疏松症的发病机制。然而，一个因果关系 T细胞变化与绝经后残留骨量的关系 待定。我们发现卵巢切除(OVX)不能导致骨丢失 在T细胞缺陷裸鼠体内。OVX诱导骨丢失的能力是 通过与野生型T细胞的T细胞重组而不是通过 用来自肿瘤坏死因子-/-小鼠的T细胞构建。此外，OVX还可增加T细胞肿瘤坏死因子 通过促进T细胞增殖和减少T细胞凋亡而产生。 这些事件导致体内产生肿瘤坏死因子的T细胞数量增加。 骨髓。这些发现表明，T细胞和T细胞产生肿瘤坏死因子 是体内E2缺乏所致骨耗损效应的重要介体。 然而，E2调节的产生肿瘤坏死因子的T细胞的特定表型 仍有待定性。同样令人费解的是雌二醇组 下调骨髓中产生肿瘤坏死因子的T细胞的数量。 因此，我们的第一个特定目标将是识别特定的T细胞 产生增加的群体(CD4+CD8+和/或γ-增量T细胞) 大量的肿瘤坏死因子，并导致去卵巢小鼠的骨丢失。这一点与 候选T细胞群将通过确定OVX是否诱导 缺乏CD4+、CD8+和/或伽马-德尔塔淋巴细胞的小鼠的骨丢失。 在特定的目标2中，我们将确定E2是否阻止T细胞依赖性的骨丢失 通过直接靶向T细胞或通过对骨髓附件的作用 细胞。这将通过分析OVX和E2的影响来实现 替代治疗对骨密度、T细胞增殖、T细胞凋亡和T细胞的影响 从雌二醇提取的T细胞重组裸鼠体内肿瘤坏死因子的产生 缺乏受体α和β的供者。我们的实验证据表明 E2通过抑制IL-7抑制T细胞肿瘤坏死因子的产生 调节T细胞增殖，诱导骨质流失。因此，我们将 通过测定IL-7在体内的中和作用来评估IL-7的贡献 IL-7抑制T细胞肿瘤坏死因子的产生和骨丢失 被OVX公司。我们的数据还表明，E2通过以下方式抑制T细胞肿瘤坏死因子的产生 抑制干扰素-γ诱导的巨噬细胞抗原提呈细胞(A-PC)活性； 通过下调CIITA基因的表达。因此，我们将得出结论 目的2用干扰素-γ中和法和干扰素-GR1-/-小鼠测定 CITA沉默对T细胞肿瘤坏死因子产生和骨丢失的影响在……里面 具体目标3我们将研究E2抑制CIITA基因的机制 表情。这将通过分析E2对 CIIT A的转录和翻译以及通过确定顺式调节 赋予CIITA启动子E2响应性的因素。这个 这个项目的意义是很高的，因为它可能会揭示发病机制。 发现绝经后骨质疏松症的新靶点，并确定骨中E2的新靶点。

DESCRIPTION (provided by applicant): Estrogen (E2) has multiple and complex regulatory effects on T cell function which may be relevant for the pathogenesis of postmenopausal osteoporosis. However, a cause-effect relationship between T cell alterations and postmenopausal bone loss remains to be established. We found that ovariectomy (ovx) failed to induce bone loss in T cell deficient nude mice. The capacity of ovx to induce bone loss was restored by T cell reconstitution with Wild type T cells but not by constitution with T cells from TNF-/- mice. Moreover, ovx increased T cell TNF production by enhancing T cell proliferation and decreasing T cell apoptosis. These events led to an increase in the number of TNF producing T cells in the bone marrow. These findings demonstrated that T cells and T cell produced TNF are essential mediators of the bone-wasting effects of E2 deficiency in vivo. However, the specific phenotype of the E2 regulated TNF producing T cell remains to be characterized. Equally enigmatic are the mechanisms by which E2 down regulates the number of TNF producing T cells in the bone marrow. Therefore, our first Specific Aim will be to identify the specific T cell population (CD4+ CD8+ and/or gamma-delta T cells) that produces increased amounts of TNF and cause bone loss in ovx mice. The relevance of the candidate T cell population will be demonstrate by determining if ovx induces bone loss in mice lacking either CD4+, CD8+ and/or gamma-delta lymphocytes. In Specific Aim 2 we will determine if E2 blocks T cell dependent bone loss via a direct targeting of T cells or via effects on bone marrow accessory cells. This will be accomplished by analyzing the effects of ovx and E2 replacement on bone density, T cell proliferation, T cell apoptosis and T cell TNF production in nude mice reconstituted with T cell harvested from E2 receptor alpha and beta deficient donors. Our experimental evidence suggests that E2 blocks T cell TNF production by repressing IL-7, a cytokine which regulates T cell proliferation and induces bone loss. Therefore, we will evaluate the contribution of IL-7 by determining if in vivo neutralization of IL- 7 prevents the increase in T cell TNF production and the bone loss induced by ovx. Our data also suggest that E2 represses T cell TNF production by blunting IFN-gamma induced macrophage antigen presenting cell (A PC) activity, through down regulation of CIITA gene expression. Therefore, we will conclude Aim 2 by using IFN-gamma neutralization and IFN-gR1 -/- mice to determine the effects of CIITA silencing on T cell TNF production and bone loss. In specific Aim 3 we will investigate the mechanism which E2 represses CIITA gene expression. This will be accomplished by analyzing the effects of E2 on the transcription and the translation of CIIT A and by determining the Cisregulating factors that confer E2 responsiveness to the CIITA promoter. The significance of this project is high, as it may shed light on the pathogenesis of postmenopausal osteoporosis and identify novel target of E2 in bone.