Interaction of fibroblasts with cell corpses increases collagen synthesis during lung repair

项目摘要 损伤后的修复是一个基本的生物学过程，对维持肺部健康至关重要。成纤维细胞必须 在损伤后执行关键的促修复功能，包括增殖和合成新的细胞外基质， 尤其是胶原蛋白，它能帮助伤口愈合死细胞（统称为细胞尸体）产生 在肺损伤和炎症过程中，并已被证明可促进从炎症到修复的转变。 先前的研究集中在理解巨噬细胞与凋亡尸体的相互作用。然而，非- 包括成纤维细胞在内的专职吞噬细胞也可以与细胞尸体相互作用， 肺中的相互作用尚未研究。该提案旨在解决有关成纤维细胞的未知问题- 尸体相互作用及其在健康肺修复中的作用。我们已经发现，细胞尸体的识别， 成纤维细胞导致成纤维细胞增加胶原蛋白合成。我们的初步数据支持一种机制 其中磷脂酰丝氨酸受体Axl结合细胞尸体，激活多胺代谢物合成， 增加细胞内的亚精胺水平，从而促进羟腐胺酸化。hypusination发生在一个单一的蛋白质： 核糖体亚基eiF5a。当羟腐胺赖氨酸化时，eiF5a能够稳定多聚脯氨酸重复蛋白的翻译 包括胶原蛋白。Hypusination对细胞增殖也很重要。阻止多胺合成 eiF5a的hypusination响应于尸体识别。阻断Axl或羟腐胺赖氨酸-eIF5a阻止了 对尸体识别的反应。我们还表明，在体内，羟腐胺赖氨酸-eIF5a在小鼠中增加。 肺损伤后胶原沉积和肺泡毛细血管修复减少， 用抑制剂GC 7或通过羟腐胺赖氨酸化酶的靶向缺失来阻断羟腐胺赖氨酸化 脱氧羟腐胺赖氨酸合酶。这使我们假设成纤维细胞与细胞尸体的相互作用 是肺损伤后激活成纤维细胞增殖和胶原蛋白合成的中心线索， 修复.我们将使用三个目标来测试1）通过Axl的尸体识别是否会触发一个受预防性调节的增加 在胶原蛋白中，2）尸体识别驱动多胺合成，增加亚精胺， 羟腐胺赖氨酸-eIF5a并稳定胶原蛋白翻译，和3）完全激活促修复编程成纤维细胞 需要通过Axl感知尸体的能力，通过鸟氨酸脱羧酶合成多胺， eIF5a通过脱氧羟腐胺赖氨酸合成酶来调节肺损伤后细胞外基质的增殖和合成。

PROJECT SUMMARY Repair after injury is a fundamental biologic process that is critical for maintaining lung health. Fibroblasts must perform key pro-repair functions after injury including proliferation and synthesis of new extracellular matrix, particularly collagens, that scaffold wound closure. Dead cells (collectively termed cell corpses) are produced during lung injury and inflammation and have been shown to promote the transition from inflammation to repair. Prior research has focused on understanding macrophage interactions with apoptotic corpses. However, non- professional phagocytes including fibroblasts can also interact with cell corpses and the consequence of these interactions in the lung has not been studied. This proposal seeks to address unknowns regarding fibroblast- corpse interactions and their role in healthy lung repair. We have found that recognition of cell corpses by fibroblasts causes fibroblasts to increase collagen protein synthesis. Our preliminary data support a mechanism where the phosphatidylserine receptor Axl binds cells corpses, activates polyamine metabolite synthesis, increasing intracellular spermidine levels, which fuel hypusination. Hypusination occurs on a single protein: the ribosomal subunit eiF5a. When hypusinated, eiF5a is able to stabilize translation of poly-proline-repeat proteins including collagen. Hypusination is also important for cell proliferation. Blocking polyamine synthesis prevented hypusination of eiF5a in response to corpse recognition. Blocking Axl or hypusine-eiF5a prevented the increase in collagen in response to corpse recognition. We also show that, in vivo, hypusine-eiF5a is increased in fibroblasts after lung injury and that collagen deposition and alveolar-capillary repair are reduced when hypusination is blocked with the inhibitor GC7 or through targeted deletion of the hypusination enzyme deoxyhypusine synthase in fibroblasts. This led us to hypothesize that interaction of fibroblasts with cell corpses is a central cue that activates fibroblast proliferation and collagen synthesis following lung injury, facilitating repair. We will use three aims to test if 1) corpse recognition via Axl triggers a translationally-regulated increase in collagen protein, 2) corpse recognition drives polyamine synthesis that increases spermidine that increases hypusine-eiF5a and stabilizes collagen translation, and 3) to fully activate pro-repair programming fibroblasts require the ability to sense corpses via Axl, synthesize polyamines via ornithine decarboxylase, and hypusinate eiF5a via deoxyhypusine synthase in order to proliferate and synthesize new extracellular matrix after lung injury.