HIF-1alpha, a Survival and Differentiation Factor for Cartilage

摘要 氧(O2)不仅是各种酶反应中不可或缺的代谢底物，包括 线粒体呼吸，也是控制！“#$转录因子稳定性和活性的调节信号 低氧诱导因子-1(HIF-1)，是细胞适应低氧压(低氧)的关键介质。这个 胎儿生长板是一种独特的间充质组织，因为它是无血管的，尽管它需要血管生成转换。 用骨头代替的命令。多年来，我们已经证明，与其无血管一致，胎儿 生长板内有缺氧区。我们已经提供了遗传证据，证明HIF-1是一种生存因素 体内低氧软骨细胞。我们已经证明，肢芽的间质凝聚也是低氧的， 在体内，肢芽间充质中缺乏HIF-1会延迟间充质细胞向软骨细胞的分化。 在这项资助中，我们建议确定调节HIF-1作为生存的作用的分子机制。 体内软骨中的分化因子。沿着这些思路，我们已经报道了在软骨细胞 HIF-1生长板的周边和肢芽的HIF-1间充质凝集 比对照组低氧多了。此外，我们还提供了基因证据，证明极端缺氧 缺氧诱导因子-1零细胞的减少不是氧对生长板可利用性降低的结果。因此，我们 假设这一定是氧气消耗量增加的结果。我们的假设符合井的假设- 已证实HIF-1能够在体外损害线粒体呼吸。基于这些发现，我们现在建议 HIF-1的一个关键功能是减少细胞的氧气消耗，因为有限的 氧气的可用性，以防止它们变成几乎缺氧的状态，这种状态与细胞不相容 生存和分化。具体地说，我们假设HIF-1对低氧生存是必不可少的。 并通过负向诱导低氧间充质细胞向软骨细胞的分化 调节线粒体的呼吸，从而调节线粒体的氧气消耗。我们将通过以下方式验证我们的假设 抑制HIF-1缺失软骨细胞和HIF-1缺失间充质细胞的线粒体呼吸 体内和体外的肢芽细胞(特异靶II)。此外，我们将确定HIF-1是否降低O2 软骨细胞在体外的消耗(特定目标III)。我们的发现可能导致范式的转变，如果我们 确定与在氧气充足的组织背景下所报道的不同， 线粒体呼吸障碍是生存和早期生存不可或缺的条件 缺氧软骨细胞的分化阶段。 $

ABSTRACT Oxygen (O2) is not only an indispensable metabolic substrate in various enzymatic reactions including mitochondrial respiration, but also a regulatory signal that controls stability and activity of !"#$transcription factor Hypoxia Inducible Factor-1¿ (HIF-1¿), a key mediator of the cellular adaptation to low O2 tension (hypoxia). The fetal growth plate is a unique mesenchymal tissue because it is avascular, albeit it requires the angiogenic switch in order to be replaced by bone. Over the years, we have demonstrated that, consistent with its avascularity, the fetal growth plate has an inner hypoxic region. We have provided genetic evidence that HIF-1¿ is a survival factor for hypoxic chondrocytes in vivo. We have shown that mesenchymal condensations of the limb bud are also hypoxic, and lack of HIF-1¿ in limb bud mesenchyme delays differentiation of mesenchymal cells into chondrocytes in vivo. In this grant, we propose to identify the molecular mechanisms that mediate the role of HIF-1¿ as a survival and differentiation factor in cartilage in vivo. Along these lines, we have reported that viable chondrocytes at the periphery of HIF-1¿ null growth plates and HIF-1¿ null mesenchymal condensations of the limb bud are considerably more hypoxic than controls. Moreover, we have provided genetic evidence that the extreme hypoxia of HIF-1¿ null cells is not the consequence of reduced availability of O2 to the growth plate. Therefore, we hypothesized it had to be the consequence of increased O2 consumption. Our hypothesis is in line with the well- documented ability of HIF-1¿ to impair mitochondrial respiration in vitro. Based on these findings, we now propose that a key function of HIF-1¿ is to reduce O2 consumption in cells that are already hypoxic because of limited availability of O2, in order to prevent them from becoming virtually anoxic, a status that is not compatible with cell survival and differentiation. Specifically, we hypothesize that HIF-1¿ is essential for survival of hypoxic chondrocytes and for timely differentiation of hypoxic mesenchymal cells into chondrocytes by negatively regulating mitochondrial respiration, and thus mitochondrial O2 consumption. We will test our hypothesis by inhibiting mitochondrial respiration in HIF-1¿ null chondrocytes (Specific Aim I) and in HIF-1¿ null mesenchymal cells of the limb bud (Specific Aim II) in vivo and in vitro. Moreover, we will establish whether HIF-1¿ lowers O2 consumption in chondrocytes in vitro (Specific Aim III). Our findings may lead to a paradigm shift if we determine that, differently from what has been reported in the context of well-oxygenated tissues, impairment of mitochondrial respiration is an indispensable requirement for survival and for early differentiation stages of hypoxic chondrocytes. $