Functional Significance of Ferritin Light Chain in Sepsis-associated Kidney Injury

脓毒症占重症监护病房急性肾损伤(AKI)的近50%，并对 死亡率。然而，脓毒症相关性AKI(SA-AKI)的治疗仍然难以捉摸，因为 损伤的病理生理学还不是很清楚。证据强调全身细胞因子的致病作用 风暴和近端肾小管损伤(氧化应激和线粒体功能障碍)在SA-AKI。细胞因子 髓系细胞(尤其是巨噬细胞)以NFkB和HIF-1α信号为主 脓毒症的发病机制。在这种情况下，用铁蛋白轻链(FTL)处理巨噬细胞会减少 脂多糖诱导NFkB和HIF-1α及随后的促炎基因的激活 表达(IL-6、肿瘤坏死因子α)。此外，对野生型小鼠给予FTL可减轻盲肠结扎和穿孔 (CLP)诱导的过度炎症反应。给药小鼠肾细胞群的单细胞RNA测序 FTL(或生理盐水)，然后内毒素血症鉴定为胱抑素A(Csta)最显著 FTL给药小鼠髓系细胞中基因上调。CSTA是一种细胞内抑制物， 组织蛋白酶。虽然csta在脓毒症中的作用尚不清楚，但组织蛋白酶通过 核因子κB的活化。因此，我们认为FTL通过csA-1途径来预防高炎性反应。 组织蛋白酶和核因子κB/缺氧诱导因子-1α信号通路在脓毒症中的介导抑制与肾脏有关，而中电 导致肾脏损伤标志物(NGAL和Kim-1)的显著表达，FTL给药阻止了这种 过氧化物酶体增殖物激活受体-γ共激活因子1-α的诱导和保存表达 (pGC-1α)，线粒体生物发生和代谢的调节因子。近端小管(PT)是 脓毒症和FTL时最大的损伤表现在这段肾小管上。因此，我们建议确定 脓毒症时巨噬细胞和PT来源的FTL在保护性反应中的不同作用。我们的发现是 临床上相关，因为血清铁蛋白水平(主要是FTL)在炎症期间经常升高，但 它在炎症中的作用尚不清楚。我们认为FTL的增加是一种适应性的生理反应 控制炎症，促进生存。在目标1中，使用两种脓毒症模型(CLP和LPS)联合 删除或过度表达FTL的新工具，我们将确定FTL是否诱导csta表达并抑制 巨噬细胞缺氧诱导因子-1α信号转导，从而防止线粒体功能障碍、糖酵解和随后的 减轻细胞因子的过度产生。在目标2中，我们将确定巨噬细胞的不同功能角色 与PT特异性FTL表达相比，减轻肾功能损失、肾脏炎症、氧化应激和 线粒体功能障碍。采用体外和体内模型的综合方法，我们将确定 FTL在脓毒症中的作用。如果我们的假设被证实，结果将证明开发一种新的 治疗SA-AKI可以减轻脓毒症引起的发病率、死亡率和 可观的医疗保健支出。

Sepsis accounts for nearly 50% of acute kidney injury (AKI) in the intensive care unit and significantly impacts mortality. However, therapy for sepsis-associated AKI (SA-AKI) has remained elusive because the pathophysiology of injury is not well understood. Evidence emphasizes the pathogenic role of systemic cytokine storm and proximal tubular damage (oxidative stress and mitochondrial dysfunction) in SA-AKI. The cytokine response, mediated by NFkB and HIF-1α signaling in myeloid cells (particularly, macrophages) is a dominant pathogenic mechanism in sepsis. In this context, treatment of macrophages with ferritin light chain (FtL) reduces lipopolysaccharide (LPS)-induced activation of NFkB and HIF-1α and subsequent pro-inflammatory gene expression (IL-6, TNFα). Additionally, administration of FtL to wildtype mice mitigates cecal ligation and puncture (CLP) induced hyperinflammation. Single cell RNA sequencing on renal cell populations from mice administered FtL (or saline) and then subjected to LPS endotoxemia identified cystatin A (CSTA) as the most significantly upregulated gene in the myeloid populations of FtL administered mice. CSTA is an intracellular inhibitor of cathepsins. While the role of CSTA in sepsis is unknown, cathepsins augment the inflammatory response via activation of NFκB. Therefore, we propose that FtL prevents the hyperinflammatory response via CSTA- mediated inhibition of cathepsin and NFκB/HIF-1α signaling during sepsis. Pertinent to the kidney, while CLP led to marked expression of kidney injury markers (NGAL and KIM-1), FtL administration prevented such induction and preserved expression of peroxisome proliferator-activated receptor gamma coactivator 1- alpha (PGC-1α), a regulator of mitochondrial biogenesis and metabolism. Proximal tubules (PT) are the sites of maximal injury during sepsis and FtL is expressed in this tubular segment. Thus, we propose to determine the disparate roles of macrophage- and PT-derived FtL in the protective response during sepsis. Our findings are clinically relevant because serum ferritin levels (predominantly FtL) are often elevated during inflammation but its role in inflammation is unknown. We propose that an increase in FtL is an adaptive physiological response to control inflammation and promote survival. In Aim 1, using two models of sepsis (CLP and LPS) combined with novel tools to delete or overexpress FtL, we will determine whether FtL induces CSTA expression and inhibits macrophage HIF-1α signaling, thereby preventing mitochondrial dysfunction, glycolysis and subsequently mitigating overproduction of cytokines. In Aim 2, we will determine the distinct functional roles of macrophage vs. PT-specific FtL expression in mitigating loss of kidney function, renal inflammation, oxidative stress and mitochondrial dysfunction. Using an integrative approach of in vitro and in vivo models, we will determine the function of FtL during sepsis. If our hypothesis is validated, the results will justify the development of a new treatment for SA-AKI that could alleviate the significant burden of sepsis induced morbidity, mortality and substantial health care expenditures.