Chertov O, Michiel D F, Xu L, Wang J M, Tani K, Murphy W J, Longo D L, Taub D D, Oppenheim J J
Laboratory of Molecular Immunoregulation, Biological Response Modifiers Program, Division of Cancer Treatment, NCI, National Institutes of Health, Frederick Cancer Research and Development Center, Frederick, Maryland 21702, USA.
J Biol Chem. 1996 Feb 9;271(6):2935-40. doi: 10.1074/jbc.271.6.2935.
Reports that interleukin-8 (IL-8) induces the infiltration of neutrophils followed by T-cells into injection sites led us to postulate that by stimulation of neutrophil degranulation IL-8 may cause the release of factors with chemoattractant activity for T-lymphocytes. Extracts of human neutrophil granules were chromatographed to isolate and purify T-lymphocyte chemoattractant factors. Two major peaks of T-cell chemotactic activity were purified by C18 reversed phase high pressure liquid chromatography (HPLC). The first peak was resolved further by C4 reversed phase HPLC and yielded an active fraction shown by NH2-terminal amino acid sequence analysis to contain defensins HNP-1, HNP-2, and HNP-3. Purified defensins HNP-1 and HNP-2 (kindly provided by Dr. R. I. Lehrer, UCLA) were also potent chemoattractants for human T-cells, while HNP-3 was inactive. The second peak of T-cell chemoattractant activity was also further purified to homogeneity by C4 reversed phase HPLC and identified by NH2-terminal sequence analysis as CAP37/azurocidin, a protein with sequence homology to serine proteases. 0.1 100 ng of defensins and 1.0 100 ng/ml CAP37 were able to stimulate in vitro T-cell chemotaxis. Neutrophil activating factors, i.e. IL-8, phorbol 12-myristate 13-acetate/ionomycin, and formylmethionylleucylphenylalanine each induced the release of CAP37 and defensins from neutrophil granules. Subcutaneous administration of defensins or CAP37/azurocidin into BALB/c mice resulted in a moderate neutrophil and mononuclear cell infiltrate by 4 h, which was greater by 24 h at the site of injection. Additionally, subcutaneous injection of defensins into chimeric huPBL-SCID mice resulted in significant infiltration by human CD3+ cells within 4 h. These results identify the antimicrobial proteins, CAP37/azurocidin and defensins HNP-1 and HNP-2, as potent neutrophil-derived chemoattractants for T-cells. These proteins represent primordial antimicrobial peptides which may have evolved into acute inflammatory cell-derived signals that mobilize immunocompetent T-cells and other inflammatory cells.
有报道称，白细胞介素-8（IL-8）可诱导中性粒细胞随后是T细胞浸润到注射部位，这使我们推测，通过刺激中性粒细胞脱颗粒，IL-8可能会导致释放出对T淋巴细胞具有趋化活性的因子。对人中性粒细胞颗粒提取物进行色谱分析，以分离和纯化T淋巴细胞趋化因子。通过C18反相高压液相色谱（HPLC）纯化出两个主要的T细胞趋化活性峰。第一个峰通过C4反相HPLC进一步分离，得到一个活性组分，通过氨基末端氨基酸序列分析表明其含有防御素HNP-1、HNP-2和HNP-3。纯化的防御素HNP-1和HNP-2（由加州大学洛杉矶分校的R.I. Lehrer博士惠赠）对人T细胞也是有效的趋化剂，而HNP-3无活性。T细胞趋化活性的第二个峰也通过C4反相HPLC进一步纯化至均一，并通过氨基末端序列分析鉴定为CAP37/天青杀素，一种与丝氨酸蛋白酶具有序列同源性的蛋白质。0.1至100 ng的防御素和1.0至100 ng/ml的CAP37能够刺激体外T细胞趋化。中性粒细胞激活因子，即IL-8、佛波醇12-肉豆蔻酸酯13-乙酸酯/离子霉素和甲酰甲硫氨酰亮氨酰苯丙氨酸，均可诱导中性粒细胞颗粒释放CAP37和防御素。将防御素或CAP37/天青杀素皮下注射到BALB/c小鼠体内，4小时时会导致中度的中性粒细胞和单核细胞浸润，注射部位在24小时时浸润更明显。此外，将防御素皮下注射到嵌合huPBL-SCID小鼠体内，4小时内会导致人CD3+细胞显著浸润。这些结果表明，抗菌蛋白CAP37/天青杀素以及防御素HNP-1和HNP-2是有效的中性粒细胞衍生的T细胞趋化剂。这些蛋白质代表了原始的抗菌肽，它们可能已经进化为动员免疫活性T细胞和其他炎症细胞的急性炎症细胞衍生信号。
