The p105/50 NF-kappaB pathway is essential for Mallory body formation.
To determine if nuclear factor-kappaB (NF-kB) plays a role in Mallory body (MB) formation, quantitative real-time RT-PCR assay was used to measure liver NF-kappaB1/p105 mRNA levels in 4 different groups of mice. Group 1: mice given IP saline for 15 weeks; group 2: mice fed diethyl 1,4-dihydro-2,4,6,-trimethyl-3,5-pyridinedicarboxylate (DDC) for 10 weeks when MBs were formed; group3: mice fed DDC 10 weeks, then withdrawn 5 weeks when MBs disappeared; group 4: mice fed DDC 10 weeks, withdrawn 4 weeks, then fed DDC+chlormethiazole (CMZ) for 1 week when MBs again formed. The mRNA for p105 NF-kappaB expression was significantly increased in the livers of mice treated with DDC (group 2) and DDC+CMZ (group 4) compared with the control livers (group 1) as well as the drug-withdrawal livers (group 3). Primary cultures of hepatocytes from drug-primed mice (the group 4 mice were withdrawn for another 4 weeks when the MBs had disappeared) were studied. The hepatocytes from drug-primed mice were MB free when isolated and used for primary culture. MBs began to form spontaneously within their cytoplasm after 2-3 days of culture. The NF-kappaB inhibitor (NF-kappaBi), a cell-permeable quinazoline compound that acts as a potent inhibitor of NF-kappaB transcriptional activation, was added to the medium 3 h after planting the cultures of liver cells. No MBs formed in the cells treated with 10 microM, 1 microM, and 0.1 microM NF-kappaBi for 6 days. MBs still formed in the cells treated with 10 nM NF-kappaBi for 6 days. Both DDC-primed and normal control liver cells began to enlarge and elongate after a few hours of culture. In contrast, the cells treated with NF-kappaBi stayed polyhedral in shape just as they appeared prior to culturing. The level of NF-kappaB1/p105 mRNA significantly increased in DDC-primed hepatocytes after 24 h of culture and in normal control hepatocytes after 48 h of culture. In DDC-primed hepatocytes, NF-kappaBi 0.1 muM treatment for 6 days significantly decreased mRNA expression of Src, p105/NF-kappaB1, ERK1, MEKK1, and JNK1/2. In normal control liver cells, NF-kappaBi treatment decreased mRNA expression of Src and JNK1 and stimulated the mRNA expression of p105/NF-kappaB1 and Junk2. NF-kappaBi treatment significantly decreased the total ERK1/2 protein and further decreased the phosphorylated (activated) form of ERK1/2 in the cultured hepatocytes. The results indicate that the p105 NF-kappaB pathway which putatively regulates ERK at both the transcriptional and post-translational levels regulates MB formation by way of changes in gene expression.
Nan
Li
L
Department of Pathology, Harbor-UCLA Medical Center, 1000 W. Carson Street, Torrance, CA 90502, USA.
Wu
Yong
Y
Bardag-Gorce
Fawzia
F
Li
Jun
J
French
Barbaba A
BA
Wilson
La Toyia
LT
French
Samuel W
SW
D000818:Animals; D034961:Antigens, Nuclear / Q000187:drug effects / Q000378:metabolism; D015153:Blotting, Western; D002478:Cells, Cultured; D056486:Chemical and Drug Induced Liver Injury; D002719:Chlormethiazole / Q000494:pharmacology; D002868:Chromosomal Proteins, Non-Histone / Q000187:drug effects / Q000378:metabolism; D004095:Dihydropyridines / Q000633:toxicity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002479:Inclusion Bodies / Q000378:metabolism* / Q000473:pathology; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D008107:Liver Diseases / Q000378:metabolism / Q000473:pathology*; D048670:MAP Kinase Kinase 4; D048728:MAP Kinase Kinase Kinase 1 / Q000378:metabolism; D008297:Male; D051379:Mice; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D008954:Models, Biological; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism*; D052002:NF-kappa B p50 Subunit; D011498:Protein Precursors / Q000187:drug effects / Q000378:metabolism*; D011799:Quinazolines / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology
D034961:Antigens, Nuclear; D002868:Chromosomal Proteins, Non-Histone; D004095:Dihydropyridines; D004791:Enzyme Inhibitors; D016328:NF-kappa B; D052002:NF-kappa B p50 Subunit; D011498:Protein Precursors; D011799:Quinazolines; D012333:RNA, Messenger; D002719:Chlormethiazole; C061879:diethyl 1,4-dihydro-2,4,6-trimethyl-3,5-pyridinedicarboxylate; D048031:JNK Mitogen-Activated Protein Kinases; D048052:Mitogen-Activated Protein Kinase 3; D048728:MAP Kinase Kinase Kinase 1; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases