NF-kappa B in Murine Sepsis

尽管使用有效的抗生素与心肺支持相结合，但在过去三十年中，脓毒症和脓毒性休克的死亡率仍然很高（29%）。此外，脓毒症和脓毒性休克的发生率似乎正在增加。需要具有广泛临床适用性的新治疗方法来降低该综合征的高死亡率。 在严重感染并发脓毒症和脓毒性休克时，炎症介质的过度释放直接导致器官损伤和死亡。核因子κ B（NF-κ B）是一种核转录调节蛋白，其对脓毒症期间编码与炎症反应相关的蛋白质的几种不同基因的激活起中心作用。在正常条件下，NF-κ B在其胞质抑制剂（I-（B）蛋白的控制下保持在胞质中的无活性状态。然而，包括LPS（革兰氏阴性细菌的毒性部分）和细胞因子（例如TNF α和白细胞介素-6）的不同种类的刺激引起I-KB蛋白的磷酸化、泛素化和随后的降解，进而导致NF-κ B的活化。然后，NF-κ B的DNA结合亚基迁移到细胞核中并激活编码炎症和免疫应答中的蛋白质的靶基因的表达，所述靶基因例如趋化因子、细胞因子、诱导型一氧化氮合酶（iNOS）和粘附分子。这些基因产物中的许多与脓毒症和脓毒性休克期间发生的血流动力学不稳定和器官损伤的发病机制密切相关。因此，设计用于抑制NF-κ B的药物可能具有广泛的抗炎作用，这在脓毒症期间可能是有益的。然而，许多与炎症反应相关并在NF-κ B控制下的宿主介质也有助于先天免疫和细菌感染的清除。因此，脓毒症期间NF-κ B的抑制也可能使潜在感染恶化。 本方案已经测试了在LPS攻击的小鼠脓毒症模型中调节NF-κ B的作用。最初的研究调查了银胶菊的效果，银胶菊是一种来源于菊科植物的倍半萜内酯。据报道，在分别用腹膜内或静脉内LPS攻击的小鼠或大鼠中，当在静脉内LPS刺激后长达3小时施用孤雌菊时，孤雌菊抑制NF-κ B并改善存活。然而，在迄今为止已经完成的研究中，在我们的小鼠模型中，用LPS攻击，孤雌生殖器恶化了存活率。此外，孤雌菊对NF-κ B的影响是时间依赖性的，早期降低这些水平，但随后增加。观察到血浆细胞因子水平的类似变化。 我们现在已经扩展了这些研究，研究丙酮酸乙酯，第二种药物，据报道，它可以抑制NF-κ B和炎症反应，并改善脓毒症模型（包括采用LPS攻击的模型）的存活率。然而，与我们对孤雌菊的发现一致，丙酮酸乙酯在我们最近的研究中也是有害的，并且对炎性细胞因子和趋化因子释放具有类似的时间依赖性作用。本研究正在进行组织NF-κ B和应激激酶测量。 然而，总体而言，这些研究表明，NF-κ B抑制在脓毒症动物模型中的作用可能是可变的。了解抑制这种中央介质在脓毒症期间的炎症和宿主反应的影响，必须明确这是在患者中探索之前。

Despite the use of effective antibiotics in combination with cardiopulmonary support, the mortality rate from sepsis and septic shock for the last three decades has remained high (29%). Furthermore, the incidence of sepsis and septic shock appear to be increasing. New therapeutic approaches with wide clinical applicability are needed to lower the high mortality rate of this syndrome. Excessive release of inflammatory mediators contributes directly to the pathogenesis of organ injury and death occurring during severe infection complicated by sepsis and septic shock. Nuclear factor kappa B (NF-KB) is a nuclear transcription regulatory protein central to the activation of several different genes encoding proteins associated with the inflammatory response during sepsis. Under normal conditions, NF-KB remains sequestered in an inactive state in the cytoplasm under the control of its cytoplasmic inhibitor (I-(B) proteins. However, differing kinds of stimuli including LPS (the toxic moiety of gram-negative bacteria) and cytokines (e.g. TNF alpha and interleukin-6) cause the phosphorylation, ubiquitinylation, and the subsequent degradation of I-KB proteins in turn resulting in the activation of NF-KB. Then the DNA-binding subunits of NF-KB migrate into the nucleus and activate expression of target genes that code for proteins in the inflammatory and immune responses, such as chemokines, cytokines, inducible nitric oxide synthase (iNOS), and adhesion molecules. Many of these gene products have been closely associated with the pathogenesis of the hemodynamic instability and organ injury occurring during sepsis and septic shock. Therefore, agents designed to inhibit NF-KB may have broad antiinflammmatory effects that could be beneficial during sepsis. However, many of the host mediators associated with the inflammatory response and under the control of NF-KB also contribute to innate immunity and the clearance of bacterial infection. Suppression of NF-KB during sepsis could therefore also worsen underlying infection. The present protocol has tested the effects of of modulating NF-KB in an LPS challenged mouse sepsis model. Initial studies investigated the effects of parthenolide, a sesquiterpene lactone derived from Asteraceae plants. Parthenolide had been reported to inhibit NF-KB and improve survival when administered up to 3 hours following intravenous LPS stimulation in mice or rats challenged with intraperitoneal or intravenous LPS respectively. However in investigations that have thus far been completed, parthenolide worsened survival with LPS challenge in our mouse model. Furthermore, the effects of parthenolide on NF-KB were time dependent, decreasing these levels early but increasing them later. Similar changes were noted with plasma cytokine levels. We have now extended these studies investigating ethyl pyruvate, a second agent which has been reported to inhibit NF-KB and the inflammatory response and to improve survival in sepsis models including ones employing LPS challenge. Consistent with our findings with parthenolide however, ethyl pyruvate has also been harmful in our most recent studies, and has had similar time dependent effects on inflammatory cytokine and chemokine release. Tissue NF-KB and stress kinase measurements are ongoing in this study. Overall however, these studies suggest that the effects of NF-KB inhibition in animal models of sepsis may be variable. Understanding the effects of inhibiting such a central mediator in the inflammatory and host responses during sepsis must be well defined before this is explored in patients.