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Oxidative stress-induced cell death of human oral neutrophils
Eisuke F Sato1, Masahiro Higashino, Kazuo Ikeda
1Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, Osaka 545-8585, Japan. sato@med.osaka-cu.ac.jp
Abstract:
Polymorphonuclear leukocytes (PMN) play crucial roles in protecting hosts against invading microbes and in the pathogenesis of inflammatory tissue injury. Although PMN migrate into mucosal layers of digestive and respiratory tracts, only limited information is available of their fate and function in situ. We previously reported that, unlike circulating PMN (CPMN), PMN in the oral cavity spontaneously generate superoxide radical and nitric oxide (NO) in the absence of any stimuli. When cultured for 12 h under physiological conditions, oral PMN (OPMN) showed morphological changes that are characteristic of those of apoptosis. Upon agarose gel electrophoresis, nuclear DNA samples isolated from OPMN revealed ladder-like profiles characteristic of nucleosomal fragmentation. l-cysteine, reduced glutathione (GSH), and herbimycin A, a protein tyrosine kinase inhibitor, suppressed the activation of caspase-3 and apoptosis of OPMN. Neither thiourea, superoxide dismutase (SOD), nor catalase inhibited the activation of caspase-3 and apoptosis. Moreover, N-acetyl-Asp-Glu-Val-Asp-aldehyde (Ac-DEVD-CHO), inhibitor for caspase-3, inhibited the fragmentation of DNA. These results suggested that oxidative stress and/or tyrosine-kinase-dependent pathway(s) activated caspase-3 in OPMN, thereby inducing their apoptosis.
Insights
Oral polymorphonuclear leukocytes (PMN) undergo apoptosis via caspase-3 activation, suggesting oxidative stress and tyrosine kinase pathways are involved. These findings offer insights into PMN fate in the oral cavity.
Area of Science:
- Immunology
- Cell Biology
- Oral Biology
Background:
- Polymorphonuclear leukocytes (PMN) are vital for host defense and inflammatory processes.
- Limited knowledge exists regarding the fate and function of PMN within mucosal tissues.
- Oral PMN (OPMN) differ from circulating PMN (CPMN), spontaneously generating superoxide and nitric oxide (NO).
Purpose of the Study:
- To investigate the mechanisms underlying apoptosis in OPMN.
- To identify signaling pathways involved in OPMN cell death.
- To elucidate the role of oxidative stress and specific enzymes in OPMN apoptosis.
Main Methods:
- Culture of OPMN under physiological conditions.
- Morphological assessment of apoptosis.
- Agarose gel electrophoresis for DNA fragmentation analysis.
- Inhibition studies using l-cysteine, reduced glutathione (GSH), herbimycin A, thiourea, superoxide dismutase (SOD), catalase, and Ac-DEVD-CHO.
Main Results:
- OPMN exhibited morphological and DNA fragmentation characteristic of apoptosis after 12-hour culture.
- l-cysteine, GSH, and herbimycin A inhibited caspase-3 activation and OPMN apoptosis.
- Thiourea, SOD, and catalase did not affect caspase-3 activation or apoptosis.
- Ac-DEVD-CHO, a caspase-3 inhibitor, blocked DNA fragmentation.
Conclusions:
- Oxidative stress and/or tyrosine-kinase-dependent pathways activate caspase-3 in OPMN.
- Caspase-3 activation leads to nucleosomal DNA fragmentation and apoptosis in OPMN.
- These findings provide insights into the programmed cell death of PMN in the oral environment.