Targeting host lipid metabolism to limit tissue damage in necrotizing fasciitis

摘要/摘要 坏死性筋膜炎是一种进展迅速的细菌感染，具有严重的 真皮和下面的软组织的坏死。治疗神经营养不良需要全身抗生素和侵袭性治疗 外科清创手术。即使使用这些治疗方法，神经纤维性疾病仍有相当高的发病率和死亡率。因此，一个更好的 了解神经营养不良的病理生理机制并确定新的治疗策略以减轻疾病 进步是必需的。最近的研究表明，促炎信号可以增加或减少 细胞对胆固醇依赖的细胞溶血素(CDCs)的抵抗力，CDCs是使细胞通透的关键微生物毒素 并破坏组织。对吞噬细胞的CDC“抵抗或敏感状态”的诱导被发现是 依赖于细胞胆固醇动态平衡的快速重新编程。此外，扰乱了 巨噬细胞重新编程其脂代谢状态破坏炎症诱导的保护性状态 信号。因此，宿主细胞中的炎症-脂质代谢回路是致病的决定因素。 CDC的潜力，这是坏死性皮肤感染的一个主要毒力因素。在此应用程序中，我们结合了 先进的方法(例如，质谱仪、单细胞测序和成像)与遗传和 脂代谢的药理学模型，以了解组织脂代谢是否是决定 疾病预防控制中心和A群链球菌(GAS)感染的致病潜力。具体目标1将决定 CH25H-LXR代谢轴介导CDC细胞保护作用的分子机制 毒性。具体地说，我们将深入研究我们的发现，LXR信号通路的激活 保护吞噬细胞免受CDC介导的膜完整性的破坏。结合脂质组学，转录组学， 成像和功能分析与功能增减模型，我们将从分子上剖析脂质 LXR介导的保护CDC介导的细胞毒性所需的代谢途径。特定目标2将 重点促进我们对皮肤中细胞类型的理解，这些细胞类型是LXR诱导的必要和充分的 防止疾病预防控制中心组织损伤。我们将结合鼠标模型应用先进的分析技术 以确定细胞类型和脂代谢途径参与诱导 皮肤对CDC的抵抗状态。特定目标3确定哪些宿主脂代谢途径是关键 用于抵抗局部或类似核因子的实验性气体皮肤感染模型。我们的数据显示，监管失调 胆固醇代谢增强CDC介导的组织损伤，但激活LXR途径诱导 保护状态。在这个目标中，我们扩展了这些令人兴奋的观察结果，并从机械上测试了是否调节血脂 宿主组织的动态平衡改变实验性核因子模型的发病机制，并可能作为辅助因素 治疗。我们期望这些研究将在分子水平上确定感染组织中的脂类代谢 影响CDC致孔毒素引起的组织损伤，并可能提供概念验证证据 靶向脂质稳态是减轻坏死性感染发病机制的有效方法。

ABSTRACT/SUMMARY Necrotizing Fasciitis (NF) or “flesh-eating disease” is a rapidly progressing bacterial infection with severe necrosis of the dermis and underlying soft tissues. Treatment of NF requires systemic antibiotics and aggressive surgical debridement. Even with these treatments, NF has considerable morbidity and mortality. Thus, a better understanding of the pathophysiology of NF and identification of new treatment strategies to attenuate disease progression is required. Recent work has revealed that pro-inflammatory signals can increase or decrease cellular resistance to the cholesterol-dependent cytolysins (CDCs), key microbial toxins that permeabilize cells and destroy tissues. The induction of a CDC “resistant or sensitive state” for phagocytes was found to be dependent on the rapid reprogramming of cellular cholesterol homeostasis. Moreover, disrupting the ability of macrophages to reprogram their lipid metabolic state disrupts the induction of protective states by inflammatory signals. Thus, an inflammatory-lipid metabolic circuit in host cells serves as a determinant of the pathogenic potential of CDCs, a major virulence factor in necrotizing skin infections. In this application, we combine advanced methodologies (e.g., mass spectrometry, single-cell sequencing, and imaging) with genetic and pharmacologic models of lipid metabolism to understand if tissue lipid metabolism is a host factor that determines the pathogenic potential of CDCs and group A strep (GAS) infections. Specific Aim 1 will determine the molecular mechanism underlying how the CH25H-LXR metabolic axis mediates the protection of cells from CDC toxicity. Specifically, we will pursue our discovery that activation of the LXR signaling pathway profoundly protects phagocytes from CDC-mediated loss of membrane integrity. Combining lipidomics, transcriptomics, imaging, and functional assays with gain- and loss-of function models, we will molecularly dissect the lipid metabolic pathways necessary for LXR-mediated protection from CDC-mediated cytotoxicity. Specific aim 2 will focus on advancing our understanding of the cell types in the skin necessary and sufficient for LXR-induced protection from CDC tissue damage. We will apply advanced analytical techniques combined with mouse models of altered lipid metabolism to determine the cell types and lipid metabolic pathways involved in inducing a resistant state to CDCs in the skin. Specific Aim 3 determines which host lipid metabolism pathways are critical for resistance to localized or NF-like experimental GAS skin infection models. Our data shows that dysregulation of cholesterol metabolism potentiates CDC-mediated tissue damage but activating the LXR pathway induces a protective state. In this aim, we extend these exciting observations and mechanistically test if modulating lipid homeostasis in host tissues alters the pathogenesis of experimental NF models and may serve as an adjunct treatment. We expect that these studies will define at the molecular level how lipid metabolism in infected tissues influences tissue damage caused the CDC pore-forming toxins and could provide proof-of-concept evidence that targeting lipid homeostasis is a productive approach to attenuating the pathogenesis of necrotizing infections.