两种景东湍蛙抗菌肽新家族的抗菌功能与作用机制研究

The drug resistance of the traditional antibiotics is becoming a more and more serious problem. Antimicrobial peptides (AMPs) become the hot topic of the antibiotic research due to their characteristics of high efficiency and safety. Amphibian skin secretes a large number of AMPs to prevent microbial infections and is one of the most important sources of AMPs. It has been proved that there are abundant AMPs in skin secretions of Amolops jingdongensis. Thirty-one antimicrobial peptides belonging to nine families were identified from the skin of this frog. Among them, Jingdongin-1 and Jingdongin- 2 are two novel antimicrobial families with unique structural motifs. We were interested in the two new antimicrobial families and preliminarily studied their hemolytic activity and antimicrobial activities to some standard strains. Based on the previous study, we will modify the structures of the two new antimicrobial peptides and compare their antimicrobial activities to some drug resistant strains isolated from clinic in the hope of obtaining safer and more effective antimicrobial precursors. At the same time, we will study their interacting with the cytomembrane, cytoderm and intracellular contents of microbes to clarify the mechanisms behind their antimicrobial activities. This study will be contribute to screen safe and efficient peptide drugs resisting to drug-resistance bacterias.

目前传统抗生素的耐药问题日益严重。抗菌肽因具高效和相对安全的特点成为抗生素研究新热点。两栖动物皮肤为抵御微生物的感染而分泌大量的抗菌肽，是抗菌肽重要来源之一。前期研究证实，景东湍蛙皮肤抗菌肽种类非常丰富。在其皮肤中克隆得到了31条编码不同抗菌肽的序列，属于9个不同家族。其中Jingdongin-1和Jingdongin-2有着独特的结构序列，是两个新鉴定的家族，并对这两个新抗菌肽的溶血活性和对部分标准菌株的抗菌活性进行了初步研究。本研究拟在前期工作的基础上，对这两个抗菌肽进行分子改造，并深入比较它们对标准菌株和临床耐药菌株的抗菌活性，以期获得对临床耐药菌株更高效、更安全的新型抗菌肽前体分子。另外，研究它们与微生物的细胞膜、细胞壁和细胞内物质的相互作用，阐明它们杀灭微生物的作用机制。本研究将有助于筛选到安全高效抗临床耐药菌的多肽药物。

抗菌肽除了具有直接的杀菌功能，还具有调控宿主抗菌免疫应答的功能，在临床细菌感染的治疗中具有良好的应用前景。两栖动物皮肤为抵御微生物的感染而分泌大量的抗菌肽，是发掘抗菌多肽的重要途径之一。随着抗生素的滥用，鲍曼不动杆菌的临床耐药性日益严重。我们的研究发现，景东湍蛙皮肤来源的新型抗菌肽Jingdongin-2，可有效抵抗临床耐药性鲍曼不动杆菌在小鼠腹腔局部的感染，腹腔注射Jingdongin-2显著降低了小鼠的鲍曼不动杆菌的载量，显著抑制了鲍曼不动杆菌感染诱导的小鼠肺部炎症损伤。抗菌机制研究发现，该抗菌肽不能直接作用于鲍曼不动杆菌，但可以激活小鼠的抗菌免疫应答。进一步分析发现，腹腔注射Jingdongin-2，可显著趋化吞噬细胞（巨噬细胞、中性粒细胞）到小鼠腹腔局部。Jingdongin-2可直接诱导小鼠骨髓来源的巨噬细胞（BMDM）产生趋化因子，可激活巨噬细胞的丝裂原活化蛋白激酶（mitogen-activated protein kinase，MAPK）和核因子κB（nuclear factor kappa-B，NF-κB）。与传统的抗生素和抗菌多肽直接作用于耐药菌相比，Jingdongin-2直接靶向宿主的免疫细胞，而非病原菌，能够避免对病原菌的选择压力。本课题将为临床鲍曼不动杆菌感染的多肽免疫疗法提供新策略。