Chondrocyte DNA Double-Strand Breaks in the Pathogenesis of Osteoarthritis

软骨细胞 DNA 双链断裂在骨关节炎发病机制中的作用

• 批准号: 10624247
• 负责人: Jennifer Harrell Jonason
• 金额: $ 33.88万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-07 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10624247
• 关键词: Acceleration; Adaptor Signaling Protein; Adult; Affect; Age; Age Months; Age of Onset; Aging; Arthritis; Automobile Driving; Biological Assay; Bleomycin; Bromodeoxyuridine; Cartilage; Cell Aging; Cell Culture Techniques; Cell Cycle; Cell Survival; Cells; Chondrocytes; Chronic; Clinic; Clinical; Coculture Techniques; DNA Damage; DNA Double Strand Break; Data; Degenerative polyarthritis; Development; Disease; Disease Progression; Endogenous Factors; Exhibits; Extracellular Matrix; Gene Expression; Gene Expression Profiling; Generations; Genotoxic Stress; Growth Factor; Harvest; Histology; Homing; Human; Immune; In Vitro; Inflammation; Inflammatory; Inflammatory Response; Innate Immune Response; Interferons; Investigation; Joints; Knee joint; Knock-out; Mass Spectrum Analysis; Matrix Metalloproteinases; Mediating; Methods; Modeling; Molecular; Mus; NF-kappa B; Nuclear; Pathogenesis; Pathway interactions; Phenotype; Population; Proliferating; Protein Inhibition; Proteins; Public Health; Reporter; Risk Factors; Role; STING agonists; Signal Pathway; Signal Transduction; Source; Stimulator of Interferon Genes; Tamoxifen; Testing; Therapeutic; Tissues; Toxic effect; Western Blotting; Wild Type Mouse; age related; aged; articular cartilage; cartilage degradation; cartilage regeneration; chemokine; cytokine; design; disability; endonuclease; experience; experimental study; in vivo; insight; joint destruction; loss of function; microCT; migration; novel; paracrine; postnatal; prevent; programs; response; senescence; small molecule inhibitor; therapeutic development; transcriptome sequencing

Osteoarthritis (OA) is the most common form of arthritis and a leading cause of disability in adults. While aging is an important risk factor for the development of OA, the molecular mechanisms responsible for OA in the context of aging remain unclear. Here, we propose to test the hypothesis that DNA DSBs in articular chondrocytes promote cell cycle re-entry and induce both IKKBeta/NF-kappaB and IRF signaling downstream of STING to mediate a pro-inflammatory secretory phenotype that leads to cartilage degeneration and inflammation in surrounding joint tissues. We show that IKKBeta/NF-kappaB signaling increases with age in the articular chondrocytes of wild type mice and that, by 24 months, these mice develop a knee joint phenotype exhibiting early hallmarks of OA. We also show that sustained IKKBeta activation in the chondrocytes of young mice greatly accelerates the onset of this age-related OA phenotype likely through paracrine actions on surrounding cells via a proinflammatory secretory program consisting of numerous cytokines, chemokines, growth factors, and MMPs that can alter proliferation and viability of neighboring cells, remodel the cartilage ECM, and initiate an innate immune response. DNA double-strand breaks (DSBs) are known to induce IKKBeta/NF-kappaB signaling and we provide evidence here of DNA DSBs in aged murine articular chondrocytes. We also provide gene expression data showing that IRF and Interferon signaling are among the most significantly affected pathways in aged articular chondrocytes. DNA DSBs can induce both NF-kappaB and IRF signaling through Stimulator of interferon genes, or STING. Thus, we hypothesize that STING may be upstream of both NF-kappaB and IRF to produce a proinflammatory secretory phenotype in chondrocytes with DSBs. In the first Aim of this proposal, we will test the effects of chondrocyte-specific DNA DSBs on chondrocyte function and OA development using AcanCreERT2/+; R26I-PpoI mice where DSBs are produced specifically in chondrocytes upon administration of tamoxifen. We will determine how chondrocytes respond to this genotoxic stress and also whether persistent DSBs are capable of promoting OA. In the second Aim, we will test whether STING is specifically involved in the activation of IKKBeta/NF-kappaB and IRF signaling following induction of DSBs and in the onset of age-related OA using in vitro cell culture methods as well as an in vivo STING knockout model in combination with NF-kappaB and IRF endogenous reporters. Results from these studies will provide valuable insight to the effects of DNA DSBs on chondrocyte fate and also on the identity of upstream activators of NF-kappaB in aged chondrocytes, potentially leading to the identification of a novel mechanism in which to inhibit IKKBeta/NF-kappaB signaling and inflammation in OA chondrocytes.

骨关节炎（OA）是成年人最常见的关节炎形式，也是残疾的主要原因。尽管衰老是OA发展的重要危险因素，但在衰老的背景下，负责OA的分子机制尚不清楚。在这里，我们建议测试以下假设：关节软骨细胞中的DNA DSB促进细胞周期重新进入，并诱导sting下游的Ikkkbeta/nf-kappab和IRF信号传导，以介导促炎的分泌表型，从而导致软骨变性和炎症在周围的关节组织中。我们表明，ikkbeta/nf-kappab信号在野生型小鼠的关节软骨细胞中随着年龄的增长而增加，并且在24个月后，这些小鼠发展出膝关节表型，表现出OA的早期标志。 We also show that sustained IKKBeta activation in the chondrocytes of young mice greatly accelerates the onset of this age-related OA phenotype likely through paracrine actions on surrounding cells via a proinflammatory secretory program consisting of numerous cytokines, chemokines, growth factors, and MMPs that can alter proliferation and viability of neighboring cells, remodel the cartilage ECM, and initiate an innate免疫反应。已知DNA双链断裂（DSB）诱导Ikkbeta/nf-kappab信号传导，我们在这里提供了老年鼠关节软骨软骨细胞DNA DSB的证据。我们还提供了基因表达数据，表明IRF和干扰素信号传导是老年关节软骨细胞中最显着影响的途径之一。 DNA DSB可以通过干扰素基因的刺激器或刺激诱导NF-kappab和IRF信号传导。因此，我们假设Sting可能是NF-KAPPAB和IRF的上游，以在带有DSB的软骨细胞中产生促炎性分泌表型。在该提案的第一个目的中，我们将使用ACANCREERT2/+测试软骨细胞特异性DNA DSB对软骨细胞功能和OA发育的影响。 r26-ppoi小鼠，在他莫昔芬给药后，在软骨细胞中专门产生了DSB。我们将确定软骨细胞如何应对这种遗传毒性应激，以及持续的DSB是否能够促进OA。在第二个目标中，我们将测试sting是否特别参与了DSB诱导后的IKKBETA/NF-KAPPAB和IRF信号传导，并在使用体外细胞培养方法以及与NF-Kappab和IRF内源性的组合中使用体外细胞培养方法以及与年龄相关的OA发作。 Results from these studies will provide valuable insight to the effects of DNA DSBs on chondrocyte fate and also on the identity of upstream activators of NF-kappaB in aged chondrocytes, potentially leading to the identification of a novel mechanism in which to inhibit IKKBeta/NF-kappaB signaling and inflammation in OA chondrocytes.