古川 聡 東京大学, 医学部・附属病院, 医員
韓 一秀 東京大学, 医学部・附属病院, 医員
深柄 和彦 東京大学, 医学部・附属病院, 医員
井上 知巳 東京大学, 医学部・附属病院, 医員
井上 知己 東京大学, 医学部・附属病院, 医員
FURUKAWA Satoshi Univ.of Tokyo, Faculty of Medicine Clinical Associate
INOUE Tomomi Univ.of Tokyo, Faculty of Medicine Clinical Associate
HAN Ilsoo Univ.of Tokyo, Faculty of Medicine Clinical Associate
|Budget Amount *help
¥13,000,000 (Direct Cost : ¥13,000,000)
Fiscal Year 1998 : ¥2,700,000 (Direct Cost : ¥2,700,000)
Fiscal Year 1997 : ¥10,300,000 (Direct Cost : ¥10,300,000)
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
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