The basic research for the therapy of Gram-negative bacterial sepsis by LPS-receptors fusion protein

脂多糖在革兰氏阴性菌感染脓毒症病理生理反应的发展中起关键和主动作用。研究表明，两种不同的受体CD14和toll样受体4 （TLR4）参与细胞对LPS刺激的识别。为探索革兰氏阴性细菌性脓毒症早期治疗的新方法，研究重组CD14-TLR4-Fc融合蛋白对LPS刺激细胞的抑制作用，比较重组CD14-Fc融合蛋白对LPS诱导的U373细胞株和人外周血单核细胞（PBMC）产生白细胞介素-8 （IL-8）的影响。因此，虽然CD14-Fc仅部分抑制LPS诱导的细胞产生IL-8，但CD14-TLR4-Fc在10ng/ml-1000ng/ml LPS下显著抑制细胞产生IL-8。特别是，5 μg/ml的CD14-TLR4-Fc几乎完全阻断了lps诱导的U373和人PBMC中IL-8的产生。CD14-TLR4-Fc之所以能强烈抑制LPS的刺激，可能是由于CD14-TLR4-Fc被这种融合蛋白捕获后，与细胞表面的TLR4区结合，LPS无法与TLR4结合。相反，被CD14-Fc捕获的LPS可能仍然保持刺激细胞表面TLR4的活性，导致LPS诱导的细胞产生IL-8仅部分抑制。本研究结果表明，重组CD14-TLR4-Fc融合蛋白对lps诱导的炎症反应具有较强的阻断作用，为开发抑制革兰氏阴性菌感染脓毒症过度炎症反应的新药提供了潜在的可能性。

LPS plays a pivotaland initiative role in the development of pathophysiological response of sepsis in gram-negative bacterial infection. It has been revealed that two distinct receptors, CD14 and toll-like receptor 4 (TLR4), are involved in the recognition of LPS stimulation by cells. For the purpose of developing a novel therapy in the early phase of gram-negative bacterial sepsis, the inhibitory effect of recombinant CD14-TLR4-Fc fusion protein against cell stimulation by LPS, comparing the effect of recombinant CD14-Fc fusion protein on the LPS-induced interleukin-8 (IL-8) production by U373 cell line and human peripheral blood mononuclear cells (PBMC). As a result, while CD14-Fc only partially suppressed LPS-induced IL-8 production by cells, CD14-TLR4-Fc significantly inhibited IL-8 production by cells in response to 10ng/ml-1000ng/ml LPS. Especially, the administration of 5 μg/ml of CD14-TLR4-Fc almost completely blocked LPS-induced IL-8 production both in U373 and in human PBMC. The reason why CD14-TLR4-Fc strongly inhibited LPS stimulation may be that LPS cannot associated with TLR4 in the cell surface because of the incorporation with the TLR4 region of the CD14-TLR4-Fc after trapping by this fusion protein. In contrast, LPS trapped by CD14-Fc may still remain the activity of stimulating TLR4 in the cell surface, resulting in the only partial suppression of LPS-induced IL-8 production by cells. The results of this study demonstrate the strong blocking effect of recombinant CD14-TLR4-Fc fusion protein against LPS-induced inflammatory response, indicating the potential possibility for the development of the novel drug for the suppression of excessive inflammatory response in sepsis by gram-negative bacterial infection.