Role of cytokines in ethanol-induced cytotoxicity in vitro in Hep G2 cells.
OBJECTIVE
As shown previously by us, ethanol (EtOH) causes time- and concentration-dependent reduction in cytoviability. Tauroursodeoxycholic acid (TUDCA) and ursodeoxycholic acid (UDCA) were shown to reduce cytotoxicity. Long-term EtOH exposure leads to immunoregulatory and detoxification impairment. This study aimed to determine the relationship between cytokine (interleukin [IL]-1 alpha and IL-6 and tumor necrosis factor [TNF]-alpha) production and expression, glutathione (GSH) status, and EtOH-induced cytotoxicity on Hep G2 cells.


METHODS
Cells were incubated with 80 mmol/L EtOH or alpha-minimal essential medium (control) in the presence or absence of 50 mumol/L TUDCA or UDCA. Cytokine release was quantitated by enzyme-linked immunosorbent assay. Cytokine expression was measured by reverse-transcription polymerase chain reaction. GSH content was determined in both the cytosolic and mitochondrial fractions.


RESULTS
After 24 hours of EtOH exposure, the release of IL-1 alpha doubled, that of IL-6 increased 10 times, and that of TNF-alpha increased 3.5 times. Cytokine expression was up-regulated compared with control for IL-1 alpha (42%), IL-6 (26%), and TNF-alpha (52%). Addition of 50 mumol/L TUDCA or UDCA reduced cytokine release and expression. TNF-alpha increased cytotoxicity by 18%. Anti-TNF-alpha antibody almost abolished it. EtOH depleted mGSH levels by 55% (P < 0.001). TUDCA replenished them by 88%.


CONCLUSIONS
EtOH up-regulated expression of cytokines in Hep G2 cells is down-regulated by bile acids. Increased amounts of TNF-alpha and depletion in both cytosolic and mitochondrial GSH contribute to EtOH cytotoxicity. Bile acids prevent toxicity.
Neuman
M G
MG
Division of Clinical Pharmacology, Sunnybrook Health Science Center, Ontario, Canada.
Shear
N H
NH
Bellentani
S
S
Tiribelli
C
C
D017209:Apoptosis / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D016207:Cytokines / Q000502:physiology*; D000431:Ethanol / Q000378:metabolism / Q000633:toxicity*; D005978:Glutathione / Q000032:analysis; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology
D016207:Cytokines; D014409:Tumor Necrosis Factor-alpha; D000431:Ethanol; D005978:Glutathione