Toll样受体4与严重多菌种脓毒症心功能不全相关性的研究

Sepsis is a systemic inflammatory response syndrome that occurs during infection and its induced mortality increased annually. Most antisepsis clinical trials have yielded frustratingly negative results. To date, the pathophysiology of sepsis is considered to be the over activation of amounts of cytokine production and immune maladjustment. Toll-like receptor 4 （TLR4）is an important transmembrane receptor expressed by heart. Various studies in different models have suggested that TLR4-mediated signaling pathway is critical for the production of inflammatory factors and related to the development of cardiac dysfunction in certain conditions. Septic cardiomyopathy is a main feature of severe sepsis and contributes to its high mortality, so the early diagnosis plays an important role in preventing the deterioration of the disease. To the best of our knowledge, there has been no report of the role of TLR4 signaling during polymicrobial sepsis induced cardiac dysfunction. Velocity vector imaging（VVI）is a new developed technique which has been widely used to assess the ventricular contractility. However little is studied about its use in detection of sepsis induced cardiac dysfunction. The present study is to assess 1）whether VVI can sensitively detect septic cardiac dysfunction in mouse model and patients; 2) the role of TLR4 signaling in cardiac dysfunction during severe polymicrobial sepsis，using the combination of new ultrasound technique（VVI）, pathological, biochemical, molecular biological and immunological methods; 3）the possibility of TLR4-targeted therapy for severe polymicrobial sepsis.

脓毒症的发病率及死亡率高，诸多治疗方法疗效欠佳。脓毒症的基本病理机制是在感染情况下发生的全身性过度炎性反应。Toll样受体4（TLR4）是心肌表达的重要跨膜受体，其介导的信号转导通路是脓毒症中炎性因子产生的重要途径，与脓毒症性心功能异常的发生发展密切相关。心功能不全是严重脓毒症的主要特征之一，早期诊断脓毒症所致心功能不全对控制疾病、防止病情恶化有重要意义。目前，关于TLR4在严重多菌性脓毒症性心功能不全中的作用尚未见报道。本研究利用超声新技术（速度矢量成像技术）结合病理学、生物化学、分子生物学、免疫学来评价TLR4在脓毒症性心功能异常中的作用，试图在整体、器官、细胞及分子四个不同层面对脓毒症心脏功能进行动物实验与临床研究，为临床早期发现脓毒症心功能不全，并利用抗TLR4抗体治疗提供科学依据。

脓毒症是感染情况下发生的全身炎症反应综合征。Toll样受体是一类表达于细胞膜上能够识别病原体的受体家族，作为天然免疫的重要组成部分。心功能不全是严重多菌种脓毒症的主要特征之一。严重多菌种脓毒症所致的心功能障碍的评估及其潜在的信号机制尚不完全明确，因而在严重多菌种脓毒症急性心肌损伤中研究TLR4的作用及其干预机制对指导临床治疗严重多菌种脓毒症具有重要意义。VVI是一种研究心肌结构力学、定量评估心脏整体舒缩功能和局部心肌运动的新方法。国内外尚未见应用VVI技术评价严重多菌性脓毒症心功能不全与TLR4相关性的报导。. 本研究通过建立小鼠（C57BL/6）严重多菌种脓毒症模型，应用常规超声技术、超声速度矢量成像技术（VVI）、离体心脏灌流技术等从整体、器官、细胞及分子四个不同层面，探讨TLR4在脓毒症所致心功能不全所起的作用。观察抗TLR4抗体对脓毒症心功能障碍及死亡率的作用。. 本研究结果显示，WT小鼠CLP术后小鼠24h小时存活率约32%，而Sham小鼠与TLR4-KO小鼠在CLP术后24h内无死亡（p<0.01）。. 通过VVI对小鼠心功能的测定结果显示，术后6h，WT-CLP小鼠的左室后壁的应变率下降了20%,而WT-sham组仅下降了6%（p<0.01）,术后12h和24h，TLR4-KO-CLP的应变率也明显高于WT-CLP小鼠（p<0.05）。而传统超声结果显示CLP术后小鼠12h、24h心功能有轻度下降，左室舒张末期内径略有扩张（p>0.05）。应用离体心脏灌流法对小鼠心功能进行再评估，CLP术后TLR4-KO组的LVDP和dP/dt max均高于WT组(p<0.05)。. 应用ELISA法测定24h血清炎症因子水平， WT-CLP组中TNF-α、IL-6、IL-10均明显高于WT-sham组， TLR4-KO-CLP组较WT-CLP组各炎症因子均明显降低 (p<0.01)。中性粒细胞内MPO的分组测定发现TLR4-KO-CLP组心肌组织内MPO的活性明显高于WT-CLP组（p<0.01)。心肌的组织病理切片结果显示，TLR4-KO-WT小鼠心肌组织损伤表现较WT-CLP组轻，且PMNs的数量也较少。. 综上所述，我们认为 TLR4信号在严重脓毒症所致的心功能障碍和高致死率起到重要作用。VVI技术能够早期、敏感、准确地评价严重脓毒症心功能障碍。