Metabolic Regulation of Articular Cartilage and Joint Homeostasis

关节软骨的代谢调节和关节稳态

基本信息

批准号：
10656369
负责人：
Regis J O'Keefe
金额：
$ 60.68万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-08 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10656369
关键词：
Ablation Acceleration Acetyl Coenzyme A Address Affect Aging Anabolism Attenuated Carbon Carrier Proteins Cartilage Catabolism Chondrocytes Citrates Clinical Treatment Cytoplasm Data Degenerative polyarthritis Development Disease Enzymes Equilibrium Gene Deletion Gene Expression Genes Genetic Glucose Glutamine Glycolysis Glycosaminoglycans Goals Hexosamines High Pressure Liquid Chromatography Homeostasis Hospital Costs Hyaluronic Acid IGF1 gene In Vitro Injury Joints Knowledge Label MAP3K7 gene Mass Spectrum Analysis Mediating Medical Medicare Medicine Metabolic Mitochondria Mus Pathway interactions Patients Persons Pharmaceutical Preparations Production Proteoglycan Radiolabeled Receptor Gene Regulation Replacement Arthroplasty Replacement Therapy Role Signal Transduction Source Symptoms Therapeutic United States Up-Regulation Work articular cartilage blood glucose regulation fructose-6-phosphate gain of function genetic approach glucose metabolism glucose uptake in vitro Model in vivo innovation insight joint injury loss of function metabolomics novel overexpression prevent receptor receptor expression response therapeutic target transcriptome sequencing uptake

项目摘要

ABSTRACT TGF is an essential regulator of articular chondrocyte/cartilage homeostasis. However, reduced/absent TGF receptor (Tgfbr2) expression with aging, joint injury, and in osteoarthritis (OA) prevents the use of TGF1 as a clinical treatment for OA. Therefore, the goal of this proposal is to identify the key genes, pathways, and potential therapeutic targets that are regulated by TGF1. Our preliminary data shows that TGF1 regulates chondrocyte homeostasis and anabolic biosynthesis through stimulation of glucose uptake, glycolysis and anabolic Hexosamine Biosynthetic Pathway (HBP). Specifically, we show that TGF, via TAK1 signaling, induces the HBP through upregulation of 3 key genes/targets: i) Glut1, the major enzyme involved in glucose uptake; ii) Gfpt2 (glutamine-fructose-6-phosphate amidotransferase-2, the rate limiting enzyme of HBP), and iii) Slc25a1, the key mitochondrial citrate transport protein that provides a source of cytoplasmic Acetyl CoA necessary for production of UDP-GlcNAc. UDP-GlcNAc is the terminal metabolite in the HBP pathway and is required for matrix synthesis of hyaluronic acid and glycosaminoglycans (GAGs). Our mass spectrometry (MS) data establish that TGF1 enhances the production of UDP-GlcNAc and increases the proportion of carbons in UDP-GlcNAc derived from radiolabeled glucose. Moreover, our RNA-seq data and additional in vitro data identify Igf1 as a critical downstream target of TGF1 since the induction of glucose metabolism, glycolytic gene expressions, glucose uptake, HBP, and proteoglycan production is abolished in in TGF1 treated articular chondrocytes with Igf1r gene deletion. In contrast, Igf1 over-expression mimics the effect of TGF1 on glucose metabolism as well as cartilage anabolism and homeostasis. Collectively, these novel findings indicate the existence of a TGF/IGF1 signaling axis in chondrocytes, and that modulation of this axis may be a promising therapeutic strategy to treat OA. Two Specific Aims are proposed. Specific Aim 1 will define the upregulation of Hexosamine Biosynthesis Pathway (HBP) as a key mechanism involved in TGF-mediated homeostasis of articular cartilage. Complementary in vitro and in vivo genetic approaches targeting Tgfbr2, Tak1, Glut1, Gfpt2 and Slc25a1 as well as HPLC-MS will be used to establish regulation of the HBP as an essential anabolic pathway necessary for articular chondrocyte homeostasis. Specific Aim 2 will utilize Igf1r loss-of-function and Igf1 gain-of-function models in vitro and in vivo to establish Igf1 signaling as a downstream effector of TGF regulation of glucose metabolism and articular cartilage homeostasis. In summary, the proposed studies will define TGF/IGF1 as a novel pathway axis in regulation of glucose metabolism, HBP, and articular chondrocytes homeostasis in the context of OA. This work will enhance our understanding of mechanisms regulating OA and provide novel targets for innovative therapeutic approaches.

抽象的 TGF是关节软骨细胞/软骨稳态的基本调节剂。但是，减少/缺失衰老，关节损伤和骨关节炎（OA）的TGF受体（TGFBR2）表达可防止使用TGF1 作为OA的临床治疗。因此，该提案的目的是确定关键基因，途径和由TGF1调节的潜在治疗靶标。我们的初步数据表明，TGF1调节软骨细胞稳态和合成代谢生物合成通过刺激葡萄糖摄取，糖酵解和合成代谢己胺的生物合成途径（HBP）。具体而言，我们证明TGF通过TAK1信号传导通过3键的上调诱导HBP 基因/靶标：i）Glut1，涉及葡萄糖摄取的主要酶； ii）GFPT2（6-磷酸谷氨酰胺 - 果糖 - 果糖 Amidotransferase-2，HBP的速率限制酶和III）SLC25A1，键柠檬酸盐的键运输蛋白质提供了生产UDP-GLCNAC所需的细胞质乙酰基COA的来源。是HBP途径中的末端代谢物，是透明质酸和基质合成所必需的糖胺聚糖（插科打）。我们的质谱（MS）数据建立TGF1增强了生产 UDP-GLCNAC的含量并增加了从放射性标记的葡萄糖衍生的UDP-GLCNAC中的碳比例。此外，我们的RNA-seq数据和其他体外数据将IGF1识别为TGF1的关键下游目标由于诱导葡萄糖代谢，糖酵解基因表达，葡萄糖摄取，HBP和蛋白聚糖在用IGF1R基因缺失的TGF1处理的关节软骨细胞中废除生产。相反，IGF1 过表达模仿TGF1对葡萄糖代谢以及软骨合作和软骨的影响稳态。总的来说，这些新颖的发现表明存在TGF/IGF1信号轴软骨细胞以及该轴的调节可能是治疗OA的有前途的治疗策略。提出了两个具体目标。特定目标1将定义己糖胺生物合成的上调途径（HBP）是关节软骨TGF介导的稳态涉及的关键机制。靶向TGFBR2，TAK1，GLUT1，GFPT2和SLC25A1的互补体外和体内遗传方法因为HPLC-MS将用于建立对HBP的调节，作为必需的合成代谢途径关节软骨细胞稳态。特定目标2将利用IGF1R功能丧失和IGF1功能获得体外和体内模型以建立IGF1信号作为TGF调节葡萄糖的下游效应子代谢和关节软骨稳态。总而言之，拟议的研究将将TGF/IGF1定义为葡萄糖代谢，HBP和关节软骨细胞稳态调节的新型途径轴 OA的背景。这项工作将增强我们对调节OA机制的理解并提供新颖的目标用于创新的治疗方法。