FoxO transcription factors in joint aging and osteoarthritis pathogenesis

摘要 骨性关节炎是最常见的关节疾病。尽管衰老是最重要的 骨性关节炎的危险因素，导致衰老相关软骨退变的机制仍有待确定。 我们报告了自噬和氧化防御等细胞动态平衡机制 老化和骨关节炎影响的软骨受损。在我们最近的研究中，我们研究了FoxO转录 调节自噬蛋白表达和自噬激活的因子。特别是FOXO3a是 被称为细胞衰老的分子守门人。我们观察到FoxO mRNA和蛋白的减少 在人和小鼠衰老和骨关节炎影响的软骨中的表达。 这些发现支持这样的假设，即与衰老相关的FoxO表达减少损害了保护性 细胞内稳态机制，损害软骨细胞的生存和生物合成能力，并导致 加速关节老化和启动骨性关节炎的发病机制。这一假设将在以下目标中得到检验。 目的1：FoxO在软骨细胞中的表达和功能调节。我们将检查细胞外 FoxO抑制或激活的调节因子和机制，并确定在FoxO抑制或激活中负责的顺式元件 FOXO推动者。利用FoxO基因敲除和过表达模型在正常人和骨性关节炎患者中的应用 从FoxO缺陷小鼠的软骨细胞和软骨移植中，我们将研究FoxO在调节 软骨细胞的功能。 目的2：软骨中的FoxO信号网络。FOXO靶基因是特定于组织和环境的。我们会 利用RNA-seq、ChIP-seq和FoxO-seq分别在小鼠软骨和人软骨细胞中建立FoxO靶基因 组蛋白标记的芯片序列。对这些数据集的综合分析将识别路径和信号 由FoxO监管的机制。 目的3：FoxO在软骨内稳态、衰老和实验性骨性关节炎中的作用。我们的初步研究表明 FoxO1的条件性软骨特异性缺失导致出生后软骨异常生长和 6个月后出现自发性骨性关节炎样降解。我们将产生出生后的三重和单基因敲除小鼠 使用Acan-Creer，并测试其老化相关变化和实验性骨关节炎的严重程度。 目的4：FoxO过度表达和激活的保护作用 我们将产生在软骨中过度表达活性形式的FoxO1或FOX03的小鼠，以平衡OA 相关抑制并确定与内稳机制和骨性关节炎严重程度相关的结果 老龄化和外科手术模型。 该项目将确定与衰老相关的FoxO表达减少是骨性关节炎的早期和关键事件 发病机制。这将为旨在调节FoxO的治疗方法提供基础 表达和/或活动，以防止年龄相关和损伤诱导的骨性关节炎的发生和发展。 好了！

ABSTRACT Osteoarthritis (OA) is the most prevalent joint disease. Although aging represents one of the most important risk factors for OA, mechanisms leading to the aging-related cartilage degeneration remain to be determined. We reported that cellular homeostasis mechanisms such as autophagy and oxidant defenses are compromised in aging and OA-affected cartilage. In our recent studies we investigated FoxO transcription factors, which regulate expression of autophagy proteins and autophagy activation. FoxO3a in particular is known as molecular gatekeeper of cellular aging. We observed a reduction in FoxO mRNA and protein expression in aging and OA-affected cartilage in humans and mice. These findings support the hypothesis that ‘Aging-related reduction of FoxO expression impairs protective cellular homeostasis mechanisms, compromises chondrocyte survival and biosynthetic capacity and leads to accelerated joint aging and initiation of OA pathogenesis’. This hypothesis will be tested in the following aims. Aim 1: Regulation of FoxO expression and function in chondrocytes. We will examine the extracellular regulators and mechanisms of FoxO suppression or activation and identify the responsible cis-elements in the FoxO promoters. Using in vitro models of FoxO knock down and overexpression in normal and OA human chondrocytes and cartilage explants from FoxO deficient mice we will examine the role of FoxO in regulating chondrocyte functions. Aim 2: The FoxO signaling network in cartilage. FoxO target genes are tissue and context specific. We will establish FoxO target genes in mouse cartilage and human chondrocytes by using RNA-seq, ChIP-seq and ChIP-seq for histone marks. Integrative analysis of these datasets will identify pathways and signaling mechanisms that are regulated by FoxO. Aim 3: Role of FoxO in cartilage homeostasis, aging and experimental OA. Our preliminary studies show that conditional cartilage specific deletion of FoxO1 leads to abnormal cartilage growth postnatally and spontaneous OA-like degradation by 6 months. We will generate postnatal triple and single knock out mice using Acan-CreER and test them for aging related changes and severity of experimental OA. Aim 4: Protective effects of FoxO overexpression and activation We will generate mice that overexpress active forms of FoxO1 or FoxO3 in cartilage to balance the OA associated suppression and determine outcomes with respect to homeostasis mechanisms and OA severity in the aging and surgical models. This project will establish that aging-related reduction in FoxO expression is an early and critical event in OA pathogenesis. This will provide the foundation for therapeutic approaches aimed at modulating FoxO expression and/or activity to prevent age-related and injury-induced onset and progression of OA. !