Panyutich A, Shi J, Boutz P L, Zhao C, Ganz T
Department of Medicine, School of Medicine, University of California at Los Angeles, 90095, USA.
Infect Immun. 1997 Mar;65(3):978-85. doi: 10.1128/IAI.65.3.978-985.1997.
Antimicrobial peptides of several structural classes have been found in phagocytes and epithelial cells of many animals. The broadly microbicidal protegrins (PG1, -2, and -3) were originally isolated as 16 to 18-amino-acid peptides from pig neutrophil lysates, but the corresponding cDNA sequences encoded much larger precursors that belonged to the cathelicidin family of antimicrobial peptides. We explored the storage, secretion, and microbicidal activation of protegrins in porcine neutrophils and in a model system consisting of recombinant proprotegrin 3 (pPG3) and various serine proteases and their inhibitors. Protegrins were stored in neutrophils as inactive proforms that were cleaved by neutrophil elastase to mature protegrins during the preparation of granule lysate and during phorbol myristate acetate-stimulated granule secretion from intact neutrophils. Recombinant pPG3 was efficiently cleaved by trace amounts of human neutrophil elastase or equivalent amounts of elastase activity from porcine neutrophils, but pPG3 was relatively resistant to porcine pancreatic elastase or human neutrophil cathepsin G. The recombinant pPG3 and neutrophil proprotegrins lacked microbicidal activity, but the mature protegrins generated in the elastase-mediated cleavage reaction were as active against Listeria monocytogenes as the chemically synthesized protegrin. The secretion and elastase-mediated activation of proprotegrins accounted for much of the stable microbicidal activity of porcine neutrophil secretions against L. monocytogenes. Secreted proprotegrins and trace amounts of elastase constitute a binary microbicidal system that is likely to contribute to the antimicrobial activity of porcine inflammatory fluids.
在许多动物的吞噬细胞和上皮细胞中发现了几种结构类型的抗菌肽。最初从猪中性粒细胞裂解物中分离出具有广泛杀菌作用的防御素（PG1、-2和-3），它们是16至18个氨基酸的肽，但相应的cDNA序列编码的前体要大得多，属于抗菌肽的cathelicidin家族。我们研究了猪中性粒细胞以及由重组前防御素3（pPG3）与各种丝氨酸蛋白酶及其抑制剂组成的模型系统中防御素的储存、分泌和杀菌激活情况。防御素在中性粒细胞中以无活性的前体形式储存，在制备颗粒裂解物过程中以及佛波酯刺激完整中性粒细胞的颗粒分泌过程中，被中性粒细胞弹性蛋白酶切割成成熟的防御素。重组pPG3能被痕量的人中性粒细胞弹性蛋白酶或等量的猪中性粒细胞弹性蛋白酶活性有效切割，但pPG3对猪胰弹性蛋白酶或人中性粒细胞组织蛋白酶G相对耐药。重组pPG3和中性粒细胞前防御素缺乏杀菌活性，但弹性蛋白酶介导的切割反应产生的成熟防御素对单核细胞增生李斯特菌的活性与化学合成的防御素一样。前防御素的分泌和弹性蛋白酶介导的激活占猪中性粒细胞分泌物对单核细胞增生李斯特菌稳定杀菌活性的很大一部分。分泌的前防御素和痕量的弹性蛋白酶构成了一个二元杀菌系统，可能有助于猪炎症液的抗菌活性。
