Mechanism of action of ethanol in the down-regulation of Gal(beta)1, 4GlcNAc alpha2,6-sialyltransferase messenger RNA in human liver cell lines.
Gal beta l, 4GlcNAc alpha 2,6-sialyltransferase (2,6-ST) mediates the addition of alpha 2,6-linked sialic acid to glycoproteins in the Golgi compartment. Down-regulation of its gene and consequent impaired activity of 2,6-ST seems to be the major cause for the appearance of asialoconjugates in the blood of long-term alcoholics. Therefore, mechanism(s) involved in the regulation of 2,6-ST gene is important and clinically relevant. Our previous work showed that long-term ethanol feeding in rats caused a marked 59% decrease of 2,6-ST activity as well as 2,6-ST messenger RNA (mRNA) level in liver that were due to the decreased stability of its mRNA. We now mimic these actions of ethanol using ( a ) human liver HepG2 cells stably transfected with ethanol-inducible human cytochrome P4502E1 (CYP2E1 cells), or ( b ) with high alcohol dehydrogenase (HAD cells) but not in wild-type HepG2 cells lacking either of the above 2 enzymes as models. Incubation of these cells for 72 hours with 100 mmol/L ethanol caused decreases (up to 76%, P < .05) of 2,6-ST mRNA levels in CYP2E1 and HAD cells but not in the wild type. However, incubation of wild-type cells with acetaldehyde at concentrations of 50 and 100 micro mol/L showed a dramatic decrease (up to 69%, P < .02) in the 2,6-ST mRNA levels. Furthermore, exposure of CYP2E1 cells to 4-hydroxy-2-nonenal, an endogenous lipid peroxidation product of reactive oxygen species, strongly decreased 2,6-ST mRNA level by 61% ( P < .02). These results demonstrate that 2,6-ST gene is highly sensitive to ethanol action in human liver cells either via its oxidation product, acetaldehyde, or via reactive oxygen species leading to the generation of a more reactive aldehyde such as 4-hydroxy-2-nonenal. Thus, this study assumes major importance and clinical relevance because 2,6-ST gene regulation in a human liver cell model is demonstrated within a few days of ethanol exposure, whereas its in vivo regulation in liver generally takes prolonged period of ethanol exposure.
Garige
Mamatha
M
The Lipid Research Laboratory 151 T, Veterans Affairs Medical Center, NW, Washington, DC 20411, USA.
Gong
Maokai
M
Rao
Manjunath N
MN
Zhang
Yili
Y
Lakshman
M Raj
MR
D000426:Alcohol Dehydrogenase / Q000235:genetics; D002460:Cell Line; D019392:Cytochrome P-450 CYP2E1 / Q000235:genetics; D015536:Down-Regulation; D000431:Ethanol / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D008099:Liver / Q000187:drug effects* / Q000201:enzymology; D012333:RNA, Messenger / Q000032:analysis; D012799:Sialyltransferases / Q000235:genetics*; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
D012333:RNA, Messenger; D000431:Ethanol; D000426:Alcohol Dehydrogenase; D019392:Cytochrome P-450 CYP2E1; D012799:Sialyltransferases; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase