the gene expression and release of the multifuctional cytokine, tumor necrosis factor alpha(TNFα) from human peripheral blood lymphocytes(HPBL) were investigated.
Normal HPBL were culture with or without viable C. albicans9VCA) for up to 72hours. After various incubation times, cell culture supernatants were collected and tested for TNF
activity by using L929 cytotoxicity. Total mRNA was isolated from cells at various times after stimulation and assessed for TNFαmRNA by polymerase chain reaction-assisted mRNA amplification (RT-PCR). The mRNA encodint TNFαwas detected at
various
times
(up to eight hours) and the intensity of the band for TNFαwas increased greatly after four hour hours of in vitro stimulation with C. albicans. In the case of mRNA encoding LI-6, the band for IL-6 was detected up to 48hours. Cell supernatants
from
HPBL
stimulated with C. albicans (KCA) or lipopolysaccharide(LPS), the activity of the TNFαat various times showed higher and continuous patterans compared with VCA and mRNA encoding TNFαwere detected up to 24hrs(KCA) and 72hrs(LPS). After 0.5, 3, 5
hours
reaction with actinomycin D(5 μg/ml) or cycloheximide(25μg/ml), the TNFαactivities and mRNA encoding TNFαwere not detected at all time points, but band for IL-6 were detected at 0.5 hour reaction in actinomycin D and all time points in
cycloheximide.
These results suggest that new transcription and translation occur in HPBL responding to C. albicans with the release of TNFαwithin the first few hours of stimulation.
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