Mechanisms of Novel Herbal Therapies for Sepsis.

 描述（由申请人提供）：尽管最近在抗生素治疗和重症监护方面取得了进展，但脓毒症仍然是重症监护病房中最常见的死亡原因，仅在美国每年就有> 225，000名受害者。脓毒症的发病机制仍然不清楚，但部分归因于晚期促炎介质（例如，HMGB 1、CIRP和NO）。我们对HMGB 1作为致死性全身炎症（LSI）的晚期介质的开创性发现（Science，285：248-51，1999）促使了对传统草药药理学调节HMGB 1分泌的复杂分子机制的研究。传统上，甘草被用于治疗消化性溃疡、肝损伤和肝炎，但其保护机制仍不清楚。我们的初步数据表明，甘珀酸（CBX），一种主要的甘草成分，甘草酸（甘草酸，GZA）的衍生物，剂量依赖性地消除LPS诱导的PKR（双链RNA激活蛋白激酶R）磷酸化和HMGB 1分泌，并拯救小鼠从致命的败血症（盲肠结扎和穿刺，CLP诱导），即使在延迟的方式。同时，已经出现令人信服的证据来支持以下的关键作用：i）细胞质HMGB 1易位中的STAT 1信号传导（PNAS，111：3068-73，2014）;和ii）炎性小体激活和HMGB 1分泌中的PKR（Nature，488：670-4，2012）。鉴于Cx43和Panx 1半通道在ATP流出中的可能作用，以及超纯LPS诱导的HMGB 1分泌对ATP的需求，重要的是要检验一种新的假设，即用粗LPS或其他关键的HMGB 1分泌刺激物延长刺激（如SAA和CIRP）可能导致Panx 1和/或Cx43半通道蛋白的上调，其促进ATP依赖性PKR和STAT 1磷酸化，从而导致HMGB 1分泌和致死性全身炎症（LSI）。因此，我们建议测试：1）新的假设，即粗LPS和其他重要的炎症刺激物（例如，SAA和CIRP）上调Panx 1和Cx43半通道表达，并触发PKR磷酸化和HMGB 1分泌（Aim 1）：2）Panx 1和Cx43半通道在致死性全身性炎症（LSI）发病机制中的新作用（Aim 2）;和3）Panx 1或Cx43半通道抑制剂的新机制（如模拟肽拮抗剂、中和抗体和草药抑制剂如GZA和CBX）影响LSI的结果（目的3）。总的来说，这些拟议的研究将提高我们对草药有效抑制外源性PAMPs或内源性促炎介质诱导的HMGB 1分泌的机制的理解，并为LSI的新型半通道靶向治疗的发展提供线索。

 DESCRIPTION (provided by applicant): Despite recent advances in antibiotic therapy and intensive care, sepsis remains the most common cause of death in the intensive care units, claiming > 225,000 victims annually in the U.S. alone. The pathogenesis of sepsis remains obscure, but is partly attributable to dys-regulated inflammatory responses sustained by late pro-inflammatory mediators (e.g., HMGB1, CIRP, and NO). Our seminal discovery of HMGB1 as a late mediator of lethal systemic inflammation (LSI) (Science, 285: 248-51, 1999) has prompted an investigation of the intricate molecular mechanisms underlying the pharmacological modulation of HMGB1 secretion by traditional herbal medicine. Gancao (Radix glycyrrhizae, or licorice) has been traditionally used in the treatment of peptic ulcer, hepatic injury, and hepatitis, but its protective mechanisms remain elusive. Our preliminary data indicated that carbenoxolone (CBX), a derivative of the major Gancao component, glycyrrhizin (glycyrrhizic acid, GZA), dose-dependently abrogated LPS-induced PKR (double-stranded RNA-activated protein kinase R) phosphorylation and HMGB1 secretion, and rescued mice from lethal sepsis (induced by cecal ligation and puncture, CLP) even if given in a delayed fashion. Meanwhile, compelling evidence has emerged to support the critical roles of: i) STAT1 signaling in cytoplasmic HMGB1 translocation (PNAS, 111: 3068-73, 2014); and ii) PKR in inflammasome activation and HMGB1 secretion (Nature, 488: 670-4, 2012). In light of the possible role of Cx43 and Panx1 hemichannels in ATP efflux, as well as the requirement of ATP for ultrapure LPS-induced HMGB1 secretion, it is important to test a novel hypothesis that prolonged stimulation with crude LPS or other key HMGB1 secretion stimuli (such as SAA and CIRP) may lead to upregulation of Panx1 and/or Cx43 hemichannel proteins, which facilitate ATP-dependent PKR and STAT1 phosphorylation, thereby contributing to HMGB1 secretion and lethal systemic inflammation (LSI). Accordingly, we propose to test: 1) the novel hypothesis that crude LPS and other important inflammatory stimuli (e.g., SAA and CIRP) up-regulate Panx1 and Cx43 hemichannel expression, and trigger PKR phosphorylation and HMGB1 secretion (Aim 1); 2) the novel roles of Panx1 and Cx43 hemichannels in the pathogenesis of lethal systemic inflammation (LSI) (Aim 2); and 3) the novel mechanisms by which Panx1 or Cx43 hemichannel inhibitors (such as mimetic peptides antagonists, neutralizing antibodies, and herbal inhibitors such as GZA and CBX) affect the outcomes of LSI (Aim 3). Collectively, these proposed studies will improve our understanding of the mechanisms by which herbal medicine effectively inhibits exogenous PAMPs or endogenous proinflammatory mediators-induced HMGB1 secretion, and shed light on the development of novel hemichannel-targeting therapies for LSI.