Mechanism of ITAM Signal Regulation in Osteoclasts

描述（由申请人提供）：通过成骨细胞的骨沉积和破骨细胞（OC）的骨吸收来保持骨完整性。骨重塑发生在整个生命过程中，形成和吸收的不平衡导致人类疾病，包括骨质疏松症和牙周炎，该机构的使命。我们正在研究itam -适配器信号在破骨细胞中的负调控。具体来说，我们正在研究磷酸酶可能调节itam -适配器DAP12信号传导的可能性。我们发现了活化的DAP12与含有sh2的肌醇-5′-磷酸酶1 （SHIP1）之间的一种新的关联。我们的中心假设是SHIP1在体外和体内OC发育和功能过程中调节DAP12信号。特异性目的1：确定SHIP1抑制DAP12信号传导的机制。这些研究旨在确定SHIP1抑制DAP12信号所需的功能域、近端激酶的作用以及SHIP1负调控的特定下游信号通路。我们将确定SHIP1在MCSF、RANKL和整合素刺激下游的DAP12负调控中的作用。我们将研究DAP12相关受体在介导SHIP1- DAP12关联中的具体作用。特异性目的2：确定SHIP1抑制DAP12信号传导如何影响OC的体外吸收、肌动蛋白环形成和存活功能。我们将在DAP12刺激期间确定SHIP1和DAP12的细胞定位。特异性目的3：确定SHIP1在体内对DAP12信号传导的影响。我们将在抗trem2抗体直接激活DAP12或用trem2融合蛋白阻断DAP12后，确定SHIP1是否在体内调节DAP12信号。特异性目的4：确定SHIP1和TREM2/DAP12在牙龈卟啉单胞菌（P. gingivalis）诱导的牙槽骨丢失中的作用。此外，我们将在体内研究DAP12和SHIP1在慢性低剂量LPS治疗或急性牙龈卟啉卟啉LPS刺激诱导的炎性小梁和牙槽骨丢失中的作用。这些研究不仅将阐明SHIP1如何调节OC中的DAP12信号，还将潜在地深入了解DAP12如何在巨噬细胞和OC中作为激活和抑制信号发挥作用。这种认识可能会导致新的治疗干预牙周病和骨质疏松症。项目描述：拟议研究的目的是确定抑制破骨细胞（溶解骨的细胞）活性所需的关键调控途径。这种认识可能会导致新的治疗干预慢性牙周病和骨质疏松症，疾病相关的过度破骨细胞活动。

DESCRIPTION (provided by applicant): Bone integrity is maintained via the careful balance of bone deposition by osteoblasts and bone resorption by osteoclasts (OC). Bone remodeling occurs throughout life and imbalances in formation and resorption lead to human diseases including osteoporosis and periodontitis, a mission of the agency. We are studying the negative regulation of ITAM-adapter signals in osteoclasts. Specifically we are investigating the possibility that phosphatases may regulate ITAM-adapter, DAP12, signaling. We have found a novel association of activated DAP12 with SH2-containing inositol-5'-phosphatase 1 (SHIP1). Our central hypothesis is that SHIP1 regulates DAP12 signaling during OC development and function in vitro and in vivo. Specific Aim 1: Determine the mechanism by which SHIP1 inhibits DAP12 signaling. These studies aim to determine the functional domains of SHIP1 that are required for inhibition of DAP12 signaling, the role of proximal kinases, and the specific downstream signaling pathways negatively regulated by SHIP1. We will determine the role of SHIP1 in negatively regulating DAP12 downstream of MCSF, RANKL, and integrin stimulation. We will investigate the specific role of DAP12-associated receptors in mediating SHIP1- DAP12 association. Specific Aim 2: Determine how SHIP1 inhibition of DAP12 signaling affects the OC functions of resorption, actin ring formation and survival in vitro. We will determine the cellular localization of SHIP1 and DAP12 during DAP12 stimulation. Specific Aim 3: Determine the affect of SHIP1 on DAP12 signaling in vivo. We will determine whether SHIP1 regulates DAP12 signaling in vivo after direct activation of DAP12 with anti-TREM2 antibodies or blockade of DAP12 with TREM2-fusion protein. Specific Aim 4: Determine the role of SHIP1 and TREM2/DAP12 in response to Porphyromonas gingivalis (P. gingivalis) induced alveolar bone loss. Additionally we will investigate the roles of DAP12 and SHIP1 in inflammatory trabecular and alveolar bone loss induced by chronic low dose LPS treatment or acute stimulation with P. gingivalis LPS in vivo. These studies will not only elucidate how SHIP1 regulates DAP12 signaling in OC but will potentially give insights into how DAP12 can function as both an activating and inhibitory signal in macrophages and OC. This understanding might lead to novel therapeutic interventions in periodontal disease and osteoporosis. Project Narrative: The aim of proposed studies is to define a key regulatory pathway needed to inhibit activity of osteoclasts, cells that dissolve bone. This understanding might lead to novel therapeutic interventions in chronic periodontal disease and osteoporosis, diseases associated with excessive osteoclast activity.