Targeting host lipid metabolism to limit tissue damage in necrotizing fasciitis

摘要/总结 坏死性筋膜炎（NF）或“食肉疾病”是一种快速进展的细菌感染， 真皮和下层软组织坏死。NF的治疗需要全身性抗生素和积极的治疗。 外科清创术。即使使用这些治疗，NF也具有相当大的发病率和死亡率。一个更好的 了解NF的病理生理学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 细胞对胆固醇依赖性溶细胞素（CDC）的抵抗力，CDC是使细胞透化的关键微生物毒素 破坏组织发现诱导吞噬细胞的CDC“抗性或敏感状态”是 依赖于细胞胆固醇稳态的快速重编程。此外，破坏能力， 巨噬细胞重新编程其脂质代谢状态，破坏了炎症介质对保护性状态的诱导， 信号.因此，宿主细胞中的炎症-脂质代谢回路充当致病性脂质代谢的决定因素。 CDCs是坏死性皮肤感染的主要毒力因子。在本申请中，我们将联合收割机 先进的方法（例如，质谱、单细胞测序和成像）， 脂质代谢的药理学模型，以了解组织脂质代谢是否是决定 CDC和A组链球菌（GAS）感染的致病潜力。具体目标1将决定 CH 25 H-LXR代谢轴如何介导细胞免于CDC的分子机制 毒性具体地说，我们将继续研究我们的发现，即LXR信号通路的激活深刻地影响了 保护吞噬细胞免于CDC介导的膜完整性丧失。结合脂质组学，转录组学， 成像和功能测定与增益和损失的功能模型，我们将分子解剖脂质 LXR介导的保护免受CDC介导的细胞毒性所必需的代谢途径。具体目标2将 重点是推进我们对皮肤中细胞类型的理解，这些细胞类型对于LXR诱导的 防止CDC组织损伤。我们将应用先进的分析技术结合小鼠模型 脂质代谢的改变，以确定细胞类型和脂质代谢途径参与诱导 皮肤对CDC的抵抗状态。特定目标3确定哪些宿主脂质代谢途径是关键的 对局部或NF样实验性GAS皮肤感染模型的抗性。我们的数据显示失调 胆固醇代谢的降低会增强CDC介导的组织损伤，但激活LXR通路会诱导 保护国家。在这个目标中，我们扩展了这些令人兴奋的观察，并从机制上测试，如果调节脂质 宿主组织的内稳态改变了实验性NF模型的发病机制， 治疗我们希望这些研究能在分子水平上确定受感染组织的脂质代谢是如何 影响CDC致孔毒素引起的组织损伤，并可以提供概念验证证据， 靶向脂质体内平衡是减弱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.