| Co-Investigator(Kenkyū-buntansha) |
IIBOSHI Yasuhiko Hyogo college of medicine, Faculty of Medicine, Research Associate, 医学部, 助手 (10340994)
FUJIMOTO Jirou Hyogo college of medicine, Faculty of Medicine, Professor, 医学部, 教授 (90199373)
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| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2004: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2003: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Research Abstract |
Mucosal model of Caco-2 cells, human colon cancer cells, was formed on wells. The 4 groups were made and incubated for 48 hours and 72 hours. One is that mucosal model was incubated without cytokine, another is that mucosal model was incubated with IL-1, the other with IL-6, the forth with TNF-α. The content of RNA was extracted from mucosa of each group. The expression of intracellular phospholipase A2 mRNA of each group was measured with Northern Blot, compared to the relative expression of β-actin. The expression of intracellular phospholipase A2 mRNA of IL-6 group was significantly increased than the others at 48 hours. At 72 hours, the expression of intracellular phospholipase A2 mRNA of TNF-α group was significantly increased than the others. Mucosal models were made on transwells with/without cytokine, to measure the transepithelial electrical resistance (TEER) at 6, 12, 24, 72 hours after the addition of cytokine.
The TEER of IL-6 group was significantly higher compared to contr
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ol group at 6,12,24 hours. At 72 hours, the TEER of IL-1 and IL-6 group was significantly decreased than the control group. The proinflammatory cytokine IL-6 increased the expression of sPLA2 (a hydrolyzer of phosphatidylcholine) of both intracellular and media.
Mucosal models were incubated for 72 h with IL-6 or media alone (control). Both media and cell lysate were analyzed for PL composition using thin-layer chromatography. The PL composition in the media did not show any differences between the two groups. Total intracellular PL contents were also unchanged ; however, IL-6 led to significant changes in PL composition including an increase in phosphatidylethanolamine(PE) and sphingomyelin(SM) and a decrease in phosphatidylcholine(PC) and lysophosphatidylcho-line(LPC) (p<0.05). Both PE and SM are known as inflammatory signaling factors involved in human IBD.
Our study suggests that cytokine may change the mucosal permeability, however this change may not be the effect of lysophospholipids made from phospholipids by phospholipase A2. Less
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