中文摘要
钩端螺旋体病是全球流行人兽共患病。病理学显示钩体病患者病变特征为脂多糖（LPS）中毒性炎症及损伤。然而，体外培养钩体LPS毒性较低，难以解释上述病理变化。我们研究显示，感染人或小鼠单核-巨噬细胞的问号钩体LPS活性显著增强，其中合成量升高约2倍，脂质A中月桂烯酸链被豆蔻烯酸链替代。低pH是感染时钩体LPS合成增加及脂质A结构改变的环境信号。LA2222和LA2541产物均为二元信号系统，可感受低pH并调控靶基因表达，其中LA2222上调合成限速酶lpxC表达；LA2541上调脂肪酸转移酶lpxD2和htrB表达。lpxC或htrB敲除后，上述LPS合成增加以及脂质A变构现象消失。变构后的脂质A可被TLR2 和TLR4识别，并通过NF-κB和JNK信号传导通路上调促炎细胞因子TNF-α,、IL-1β和IL-8的合成与释放，未变构脂质A仅可被TLR2识别。综上所述，问号钩体感染时通过增加LPS合成、脂质A变构而使LPS活性显著升高，LA2222和LA2541感受低pH并通过上调LPS合成限速酶、脂肪酸转移酶表达水平导致感染时LPS合成增加及脂质A变构，脂质A变构物诱导炎症反应能力显著增强。
英文摘要
Infection of L. interrogans causes leptospirosis, a world-spread zoonotic infectious disease. The disease is pathologically characterized as lipopolysaccharide (L-LPS)-induced systemic inflammation and tissue injury. However, the contradiction between much lower toxicity of L-LPS from the spirochete in medium and pathologic changes in leptospirosis remains unanswered. LPS has been considered as stability in structure and quantity in whole life-process of Gram-negative bacteria, and fatty acid types in lipid A of LPS have been confirmed to decide LPS toxicity. However, we found that L-LPS from L. interrogans strain Lai during infection of mouse or human macrophage was about two-fold increased compared that from the spirochete in medium. Interestingly, the lauroleic acid in lipid A of L-LPS (L-lipid A) was changed into myristoleic acid after infection. The infection-induced pH decrease (6.0), rather than temperature or osmotic pressure, triggered the signaling for the numeral and structural changes of L-LPS. Leptospiral LA2222 and LA2541 gene-encoding histidine kinase-response regulator hybrid proteins were confirmed to sense the pH change. In the lower pH condition, the LA2222 gene up-regulated the expression of lpxC gene, whose product is a rate-limiting enzyme in L-LPS synthesis, while the LA2541 gene up-regulated the expression of lpxD2 and htrB genes, whose products are fatty acid ferases in lipid A formation. Deletion of the lpxC or htrB gene caused the absence of L-LPS synthesis increase and fatty acid change as described above. The modified L-lipid A could be recognized by both TLR2 and TLR4 that induced higher levels of TNF-α, IL-1β and IL-8 though NF-κB and JNK signaling pathways compared to the unmodified L-lipid A that recognized by TLR2 alone. Thus, our study revealed that the enhancement of L-LPS toxicity by increase of L-LPS synthesis and structural modification of L-lipid A through pH-dependent signaling in L. interrogans during infection of host cells.