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Role of polymorphonuclear neutrophil (PMN) cell death in inflammation and infection

多形核中性粒细胞 (PMN) 细胞死亡在炎症和感染中的作用

基本信息

批准号：
09470245
负责人：
SAITO Hideaki
金额：
$ 8.32万
依托单位：
University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

We investigated the role of polymorphonuclear neutrophil(PMN)cell death in inflammation and infection, and obtained the following findings :(1) PMNs from healthy volunteers were cocultured with or without live E.coli at different ratios. The PMNs were then analyzed by flow cytometry for cell death, reactive oxygen intermediates (ROI) production, and CD16 expression. Morphologic features also were assessed. PMN apoptosis was confirmed by DNA gel electrophoresis. TNF-alpha, IL-1beta and IL-6 levels in cell free supernatants were also measured. Low doses of E.coli inhibited PMN apoptosis. In contrast, a high dose of E.coli increased PMN necrosis. The mean percentages of PMN apoptosis after coculture with E.coli showed significant inverse correlations with cytokine levels in cell free supernatants. While ROI production increased after coculture with E.coli, CD16 expression decreased after coculture with E.coli.(2) PMNs were cultured with various kinds of cytokines including TNF-alpha, IL-1 … More beta, IL-6, IL-8, GM-CSF and IL-10. Then, PMN cell death was assessed. While low dose of TNF-alpha inibited PMN apoptosis, high dose of TNF-alpha increased PMN apoptosis. IL-1beta, IL-6 and GM-CSF inhibited PMN apoptosis. IL-10 attenuated PMN apoptosis inhibition by IL-6.(3) PMNs were pretreated with GH (0 or 100 ng/mL) for 3 hours, then cultured for 0, 4 or 12 hours and PMNs were analyzed by flow cytometry for cell death, ROI production, CD16 and Fas expression. GH inhibited PMN apoptosis at 12 hours of culture. GH enhanced ROI production by PMNs, and GH decreased Fas expression on PMNs.(4) Peripheral blood samples were collected from patient before surgery, on day 1, 3, and 7 after surgery. Peritoneal samples were obtained aseptically through abdominal drain on day 1, 2 and 3 after surgery. PMNs were analyzed by flow cytometry for cell death. Peripheral PMN apoptosis was inhibited on day 1 and 3, and increased on day 7 after surgery. Peritoneal PMN apoptosis also was inhibited on day 1 after surgery, and significantly increased by days 3.These findings suggest that augmented PMN bactericidal function, via inhibition of PMN cell death, may be beneficial for host defense against bacterial infection and/or sepsis. Less

我们研究了多形核中性粒细胞（PMN）细胞死亡在炎症和感染中的作用，并获得以下发现：（1）将来自健康志愿者的PMN与或不与活大肠杆菌以不同比例共培养。然后通过流式细胞术分析 PMN 的细胞死亡、活性氧中间体 (ROI) 产生和 CD16 表达。还评估了形态学特征。通过DNA凝胶电泳证实PMN凋亡。还测量了无细胞上清液中的 TNF-α、IL-1β 和 IL-6 水平。低剂量的大肠杆菌抑制中性粒细胞凋亡。相反，高剂量的大肠杆菌会增加中性粒细胞坏死。与大肠杆菌共培养后，PMN 凋亡的平均百分比与无细胞上清液中的细胞因子水平呈显着负相关。与大肠杆菌共培养后ROI产量增加，而与大肠杆菌共培养后CD16表达下降。(2)将PMN与TNF-α、IL-1…更多β、IL-6、IL-8、GM-CSF和IL-10等多种细胞因子一起培养。然后，评估中性粒细胞死亡。低剂量的 TNF-α 抑制 PMN 凋亡，而高剂量的 TNF-α 则增加 PMN 凋亡。 IL-1β、IL-6 和 GM-CSF 抑制 PMN 凋亡。 IL-10减弱了IL-6对PMN凋亡的抑制作用。(3)用GH(0或100ng/mL)预处理PMN 3小时，然后培养0、4或12小时，并通过流式细胞术分析PMN的细胞死亡、ROI产生、CD16和Fas表达。培养 12 小时时 GH 抑制 PMN 凋亡。 GH增强PMNs的ROI产生，并且GH降低PMNs的Fas表达。(4)术前、术后第1、3、7天采集患者外周血样本。手术后第1、2和3天通过腹腔引流无菌获取腹膜样本。通过流式细胞术分析 PMN 的细胞死亡情况。外周PMN细胞凋亡在术后第1天和第3天受到抑制，并在术后第7天增加。腹膜 PMN 细胞凋亡在术后第 1 天也受到抑制，并在第 3 天显着增加。这些发现表明，通过抑制 PMN 细胞死亡来增强 PMN 杀菌功能，可能有利于宿主防御细菌感染和/或败血症。较少的