Toll-like Receptors and Host Response to Vibrio vulnificus, An Emerging Pathogen

描述(申请人提供)：创伤弧菌(VV)是一种新出现的革兰氏阴性细菌病原体，在世界各地温暖的海洋环境中发现。慢性肝病，特别是酒精性肝病，是严重VV感染的主要危险因素。即使使用抗生素治疗，严重VV感染的死亡率也可以超过50%。大多数死亡是由感染性休克引起的，这种休克是由促炎症细胞因子如TNF1的失调引起的，推测是由于VV激动剂与Toll样受体(TLRs)的相互作用。TLRs是一个模式识别受体家族，在有害的炎症反应中发挥重要作用。最近的一项研究表明，VV脂蛋白通过TLR2信号通路诱导人单核细胞产生TNF1。我们用VVATCC 27562的临床(血液)分离株获得的初步数据表明：(1)TLR4信号在VV细胞刺激的小鼠血液和脾细胞产生TnF1中起关键作用；(2)TLR4信号在VV感染的小鼠模型中是有害的；(3)除TLR4外，TLR4信号是根除VV感染所必需的；(4)TLR4介导的TnF1反应在VV感染的结局中起着关键作用。与VV引起的脓毒症相关的高发病率和死亡率强调了新的治疗策略的必要性。我们推测，用血管紧张素受体拮抗剂(S)选择性阻断TLR介导的有害炎症反应，结合抗生素治疗，将改善VV诱导的脓毒症患者的临床结局。这项研究的长期目标是更好地了解TLR介导的宿主对VV的反应，以便开发新的辅助疗法来阻断有害的TLR信号。具体目的是：(1)比较野生型(WT)、TLR2基因敲除(KO)、TLR2/TLR4KO和TLR2/TLR4KO与VV临床分离株ATCC-27562、MO6-24、YJ016和C7184体外刺激的野生型(WT)、TLR2基因敲除(KO)、TLR2/TLR4KO和VV临床分离株ATCC 27562、MO6-24、YJ016和C7184对VV感染小鼠存活和清除感染能力的影响；(3)采用小鼠脓毒症模型，观察选择性阻断TLR信号通路联合抗生素治疗对VV感染WT对照组小鼠和酒精性肝病WT小鼠存活和清除感染能力的影响。这项研究将产生新的信息，对于设计新的治疗策略至关重要，以治疗由VV和潜在的其他病原体引起的严重感染，这些病原体通过TLR介导的炎症反应损害宿主组织。 公共卫生相关性：创伤弧菌(VV)严重感染患者的发病率和死亡率很高，尤其是那些酒精性肝损伤患者。这项研究将提供新的信息，将显著加强我们对TLRs在VV先天反应中作用的有限了解，并(B)通过选择性阻断介导有害炎症反应的TLR信号，指导新辅助疗法的开发，以改善VV诱导的脓毒症患者的临床结果。这项研究还将提供与设计新的治疗策略相关的基本信息，以治疗由其他病原微生物引起的脓毒症，这些病原微生物会引起有害的TLR介导的炎症反应。

DESCRIPTION (provided by applicant): Vibrio vulnificus (Vv), an emerging Gram-negative bacterial pathogen, is found in warm marine environments worldwide. Chronic liver disease, particularly alcohol-induced liver disease, is a major risk factor for severe Vv infection. Even with antibiotic treatment, the mortality rate for severe Vv infection can exceed 50%. Most fatalities are caused by septic shock that results from dysregulation of proinflammatory cytokines such as TNF1, presumably due to interaction of Vv agonists with Toll-like receptors (TLRs). TLRs are a family of pattern recognition receptors that play a major role in the harmful inflammatory response. A recent study showed that Vv lipoprotein elicits TNF1 production by human monocytes via TLR2 signaling. Our preliminary data obtained with Vv ATCC 27562, a clinical (blood) isolate, demonstrates that: (1) TLR4 signaling plays a key role in TNF1 production by mouse blood and splenocytes stimulated with Vv cells; (2) TLR4 signaling is deleterious in a mouse model of Vv infection; (3) signaling by TLR(s), exclusive of TLR4, is needed to eradicate Vv infection; and (4) the TLR-mediated TNF1 response plays a critical role in determining the outcome of Vv infection. The high morbidity and mortality associated with Vv-induced sepsis underscore the need for new therapeutic strategies. We hypothesize that selective blockade of the TLR-mediated harmful inflammatory response with TLR antagonist(s), in combination with antibiotic therapy, will improve the clinical outcome of patients with Vv-induced sepsis. The long-term goal of this study is to gain a better understanding of the TLR-mediated host response to Vv in order to develop new adjunctive therapies that block harmful TLR signaling. Specific aims are to: (1) Compare, using ELISA, the TNF1 response of wild-type (WT), TLR2 knockout (KO), TLR4 KO and TLR2/TLR4 KO mouse whole blood and resident liver macrophages stimulated ex vivo with the genotypically distinct Vv clinical strains ATCC 27562, MO6-24, YJ016, and C7184; (2) Determine, using the mouse sepsis model, the effect of TLR2 KO and TLR2/TLR4 KO on the ability of Vv-infected mice to survive and clear infection; and (3) Investigate, using the mouse sepsis model, the effect of selective blockade of TLR signaling combined with antibiotic therapy on the ability of Vv-infected WT control mice and WT mice with alcohol-induced liver disease to survive and clear infection. This study will yield new information that is essential for designing novel therapeutic strategies for serious infections caused by Vv and potentially other pathogens that damage host tissues via the TLR-mediated inflammatory response. PUBLIC HEALTH RELEVANCE: Morbidity and mortality rates are high for patients with severe V. vulnificus (Vv) infection, particularly those with alcohol-induced liver damage. This study will provide new information that will (a) significantly enhance our limited understanding of the role of TLRs in the innate response to Vv and (b) guide development of new adjunctive therapies to improve the clinical outcome of patients with Vv-induced sepsis by selectively blocking TLR signaling that mediates the harmful inflammatory response. This study will also provide basic information that is relevant to the design of new therapeutic strategies for treatment of sepsis caused by other pathogenic microbes that evoke a harmful TLR-mediated inflammatory response.