2006 Fiscal Year Final Research Report Summary
Mechanisms of Activation and Induction of Cell Death of Oral Polymorphonuclear neutrophils
| Project/Area Number |
17592000
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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 |
Conservative dentistry
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| Research Institution | Meikai University |
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Principal Investigator |
TAKAHASHI Keiso Meikai University, School of Dentistry, Associate Professor, 歯学部, 助教授 (70243475)
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| Project Period (FY) |
2005 – 2006
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| Keywords | oral polymorphnuclear leucocytes / reactive oxygen species / signal transduction / anti-oxidant therapy / caspase / glutathione |
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| Research Abstract |
Oral PMNs (OPMNs) were collected from periodontally healthy volunteers by the mouth washing. Unstimulated OPMNs produced approximately 2-fold the amount of active oxygens and had approximately half the intracellular concentration of GSH compared to peripheral PMNs (PPMNs). The intracellular GSH level negatively correlated with that of active oxygen produced by stimulation with PMA or opsonized zymosan A. H-7, did not inhibit the active oxygen production of OPMNs, whereas W-7, inhibited it partially. Most of the OPMNs rapidly underwent cell death within 1 hour. Flow cytometry showed that more than 60% of OPMNs became apoptotic or necrotic within 2 hours after isolation. The death of these cells was not effectively inhibited by caspase inhibitors or p38 mitogen-activated protein kinase inhibitor. The activation of caspase-3, -8 and -9 in OPMNs were significantly lower than that in PPMNs (p<0.05). Electron microscopic observation revealed the vacuolization of the cytoplasm of OPMNs after 2 hours in culture. The mitochondrial membrane potential of OPMNs was reduced. Western blot analysis showed that the expression of Bad, Bax and cytochrome c were significantly elevated (p<0.05), whereas the expression of Bc1-2 and phospho-Bad (Ser136) was nil or significantly reduced, in OPMNs compared with their levels in PPMNs (p<0.05). Both NAC and GSH, but neither superoxide dismutase nor catalase, effectively delayed the cell death. NAC inhibited the reduction in mitochondrial membrane potential of the OPMNs. These results suggest that oxidative stress, accompanied by the decrease of intracellular GSH level, plays a crucial role in the activation and induction of cell death of OPMNs, and that OPMNs undergo cell death via a caspase-independent pathway involving proapoptotic molecules around the mitochondrial membrane. It may thus be possible to regulate the activation and cell death of OPMNs by SH-containing antioxidants.
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