Phlpp protein phosphatases in cartilage development and disease

摘要 腕骨是一种软组织，参与骨延长、骨折修复和无痛关节运动。损坏 骨骼软骨可阻碍骨发育并引起疼痛的关节疾病如骨关节炎（OA）， 这是美国和世界各地残疾和住院的主要原因。该项目将 探索酶Phlpp 1和Phlpp 2如何调节骨骼的形成、维持和再生 软骨Phlpp 1和Phlpp 2是细胞内蛋白磷酸酶，通过以下途径控制软骨细胞凋亡： 后修饰蛋白质底物（例如，AKT、PKCα）。来自OA患者的关节软骨细胞 表达异常高水平的Phlpp 1和Phlpp 2。Phlpp 1的整体缺失增加软骨细胞 增殖和基质合成，并减少损伤诱导的OA，同时刺激软骨的表达 生长因子和受体（Fgf 18、Tgfβ和Pthr 1）。Phlpp 1或Phlpp 2的缺失也诱导了 小蛋白聚糖（Prg 4，Dcn，Spp 1），改善软骨组织完整性和关节润滑。中 创伤后OA的临床前动物模型，单次关节内注射小分子Phlpp 抑制剂暂时减少软骨损失和与关节损伤相关的疼痛，而没有不良事件 五个星期这些Phlpp抑制剂还促进软骨细胞增殖和基质基因的产生。 因此，Phlpp抑制是OA的一种新的治疗策略，但是关于Phlpp 1和Phlpp 2如何 调节信号通路和促进软骨细胞粘附是必要的，以最大限度地提高治疗效果。 Phlpp抑制剂的潜力。该项目的总体目标是阐明新的分子和细胞 Phlpp 1和Phlpp 2在软骨发育和创伤后OA中的作用机制 发病机制，并确定在受伤的关节炎关节Phlpp抑制剂的特异性。具体目标是 目的：1）确定Phlpp 1和Phlpp 2在胶原2阳性细胞增殖和成熟中的作用， 2）确定Phlpp 1和Phlpp 2在成人聚集蛋白聚糖表达中的分子功能， 3）测试Phlpp抑制剂对Phlpp 1和Phlpp 2的特异性，并改善其对关节损伤后的细胞的影响。 在创伤后OA模型中的有效性;以及4）阐明GIRK通道对 Phlpp 1和Phlpp 2与合成代谢信号事件的整合。

ABSTRACT Cartilage is a soft tissue involved in bone lengthening, fracture repair and painless joint motion. Damage to skeletal cartilages can hinder bone development and cause painful joint diseases like osteoarthritis (OA), which is a leading cause of disability and hospital visits in the USA and around the world. This project will explore how the enzymes Phlpp1 and Phlpp2 regulate the formation, maintenance and regeneration of skeletal cartilage. Phlpp1 and Phlpp2 are intracellular protein phosphatases that control chondrocyte anabolism by post-translationally modifying protein substrates (e.g., AKT, PKCα). Articular chondrocytes from OA patients express abnormally high levels of Phlpp1 and Phlpp2. Global deletion of Phlpp1 increases chondrocyte proliferation and matrix synthesis, and reduces injury-induced OA while stimulating the expression of cartilage growth factors and receptors (Fgf18, Tgfβ and Pthr1). Phlpp1 or Phlpp2 depletion also induces the expression of small proteoglycans (Prg4, Dcn, Spp1) that improve cartilage tissue integrity and joint lubrication. In a preclinical animal model of post-traumatic OA, a single intra-articular injection of a small molecule Phlpp inhibitor temporarily reduced cartilage loss and pain associated with a meniscal injury without adverse events for five weeks. These Phlpp inhibitors also promoted chondrocyte proliferation and production of matrix genes. Thus Phlpp inhibition is a new therapeutic strategy for OA, but additional knowledge of how Phlpp1 and Phlpp2 regulate signaling pathways and promote chondrocyte anabolism is necessary to maximize the therapeutic potential of Phlpp inhibitors. The overall goals of this project are to elucidate novel molecular and cellular mechanisms of Phlpp1 and Phlpp2 action during endochondral development and post-traumatic OA pathogenesis, and to define the specificity of Phlpp inhibitors in injured articulating joints. The specific aims are to: 1) define the roles of Phlpp1 and Phlpp2 in proliferation and maturation of collagen 2-positive cells during skeletal development; 2) determine the molecular functions of Phlpp1 and Phlpp2 in adult aggrecan-expressing cells following a joint injury; 3) test the specificity of Phlpp inhibitors to Phlpp1 and Phlpp2 and improve their effectiveness in post-traumatic OA models; and 4) elucidate the contribution of GIRK channels to the integration of Phlpp1 and Phlpp2 with anabolic signaling events.