Protein kinase C-zeta mediates apoptosis of mouse Kupffer cells via ERK-1/2: a novel mechanism.
BACKGROUND
We have demonstrated that activated Kupffer cells undergo accelerated apoptosis via Toll-like receptor (TLR)-4 and protein kinase C (PKC)-ζ-dependent nuclear factor (NF)-κB activation. Because PKC-ζ plays a pivotal role in cell signaling, we sought to determine the signaling pathway of PKC-ζ in Kupffer cell apoptosis.


METHODS
Mouse Kupffer cell line (MKCL3-2) were transfected with PKC-ζ small interfering RNA (siRNA) and then treated with elastase alone or elastase along with the extracellular signal-regulated kinase (ERK) inhibitor U0126. Cell extracts were assayed for PKC-ζ (protein and activity), TLR-4, NF-κB nuclear translocation, phosphorylated ERK-1/2, activated caspase-3, and DNA fragmentation. All n ≥3; data are expressed as mean values ± standard deviations; means were compared using the t test; P < .05 was considered significant.


RESULTS
Elastase upregulated TLR-4, PKC-ζ, NF-κB, ERK-1/2, caspase-3, and DNA fragmentation (all P < .01 versus control). Transfection with PKC-ζ siRNA attenuated the elastase-induced upregulation of PKC-ζ activity, NF-κB, ERK-1/2, caspase-3, and DNA fragmentation (all P < .01 versus control). The interaction of PKC-ζ with ERK-1/2 was increased by elastase and was attenuated by PKC-ζ siRNA as confirmed by co-immunoprecipitation and immunofluorescent staining.


CONCLUSIONS
Activation of Kupffer cells upregulates PKC-ζ activity, increases apoptosis, and induces nuclear translocation of NF-κB via ERK-1/2-dependent pathways. Inhibiting the activity of PKC-ζ significantly attenuates Kupffer cell apoptosis, NF-κB, and ERK-1/2 activation. The interaction of PKC-ζ and ERK-1/2 warrants further investigation.
Peng
Yanhua
Y
Department of Surgery, James A. Haley Veterans Affairs Medical Center, University of South Florida Health Sciences Center, Tampa, FL, USA.
Sigua
Celia A
CA
Murr
Michel M
MM
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D002070:Butadienes / Q000494:pharmacology*; D053938:DNA Fragmentation; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D047468:Immunoprecipitation; D007728:Kupffer Cells / Q000166:cytology* / Q000187:drug effects; D051379:Mice; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D023421:Models, Animal; D016328:NF-kappa B / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology*; D010196:Pancreatic Elastase / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D011493:Protein Kinase C; D034741:RNA, Small Interfering / Q000378:metabolism; D011897:Random Allocation; D012016:Reference Values; D015398:Signal Transduction; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D015854:Up-Regulation / Q000235:genetics; D000099221:Protein Kinase C zeta
D002070:Butadienes; D048052:Mitogen-Activated Protein Kinase 3; D016328:NF-kappa B; D009570:Nitriles; D010196:Pancreatic Elastase; D011493:Protein Kinase C; D034741:RNA, Small Interfering; D051197:Toll-Like Receptor 4; D000099221:Protein Kinase C zeta; C113580:U 0126