2007 Fiscal Year Final Research Report Summary
The effect of intermedilysin on human bile duct cells
| Project/Area Number |
18592003
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Morphological basic dentistry
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| Research Institution | The University of Tokushima |
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Principal Investigator |
HIROTA Katsuhiko The University of Tokushima, Institute of Health Bioscience Graduate School, associate professor (60199130)
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| Co-Investigator(Kenkyū-buntansha) |
MIYAKE Yoichiro The University of Tokushima, Institute of Health Bioscience Graduate School, professor (80136093)
NEMOTO Ken The University of Tokushima, Institute of Health Bioscience Graduate School, assistant professor (10218274)
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| Project Period (FY) |
2006 – 2007
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| Keywords | S.intermedius / intermedilysin / bile duct cells / non-apoptotic cell death / Ca^<2+> oscillation / BiP / calcineurin / NFAT |
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| Research Abstract |
Streptococcus intermedius is a commensal member of the human oral, gastrointestinal and urinary floras, but may also be associated with deep-seated purulent infections, particularly in the liver abscess.
It was recently reported that the function of bacterial pore-forming toxins is biphasic. Not only do they act cytolytically on nucleated target cells when present in higher concentrations, but they can also affect signal-transduction pathways in cells when the toxin is present in low, sublytic concentrations. Intermedilysin (ILY), a human-specific pore-forming cytolysin, has been suggested as a potentially important virulence factor of S.intermedius.
Whereas Ca^<2+> is a key regulator of cell survival, the breakdown of Ca^<2+> homeostasis due to its sustained elevation was able to trigger cell death. The goals here were to examine whether a low, sublytic concentration of ILY affects Ca^<2+> homeostasis in human bile duct cells by induction of [Ca^<2+>]I oscillation and [Ca^<2+>]I related
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other cell responses (the activation of calcineurin/NFAT pathway, PI3/AKT/MTOR pathway, and endoplasmic reticulum stress).
Fron the calcium imaging, five rain ILY treatment generated an increase in[Ca^<2+>]i level from the average base line of 50 nM up to the average peak of 400 nM marked by two repeated oscillations every minute. Increased [Ca^<2+>]i was initially detected in the nuclei of ILY-treated cells subsequently followed with markedly increase in ER area. Immunocytochemistry and Westernbloting analysis, ILY treatment caused activation of calcineurin/NFAT1 signalling pathway and increased expression of AKT(p-S473),BiP, and PTEN. Interestingly the expression of BiP and AKT(p-S473)decreased in an hour post ILY treatment. Calcineurin inhibitor cyclosporine A prevented ILY-induced calcineurin/NFAT1 activation, cell death, and calcium oscillations.
ILY in concentration of 10-40 ng/ml capable of inducing calcium oscillations, activation of calcineurin/NFAT1 signalling pathway ; inactivation of PI3K/AKT/MTOR pathway, and caused ER stress disorder resulted non-apoptotic cell death in HuCCT1 cells. Less
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