cyclin-d1 and Liver-Diseases--Alcoholic

Alcoholic liver injury is associated with hepatic stellate cell (HSC) activation. A-kinase anchoring protein 12 (AKAP12) scaffolds protein kinase C and cyclin-D1, which is regulated by its phosphorylation, and spatiotemporally controls cell proliferation, invasiveness, and chemotaxis. HSC activation induces AKAP12 expression, but the role of AKAP12's scaffolding activity in liver function is unknown. Because AKAP12 phosphorylation is enhanced in ethanol-treated HSCs, we examined AKAP12's scaffolding functions in alcohol-mediated HSC activation and liver injury. AKAP12 expression, interaction, and phosphorylation were assayed in in vitro and in vivo ethanol models and human subjects by real-time PCR, coimmunoprecipitation, immunoblotting, and phosphorylated proteomics/Phos-tag. Ethanol induced AKAP12 phosphorylation in the liver and in primary HSCs, but not in hepatocytes. AKAP12's scaffolding activity for protein kinase C/cyclin-D1 decreased in ethanol-treated HSCs but not hepatocytes. AKAP12 negatively regulated HSC activation, which was reversed by ethanol-mediated AKAP12 phosphorylation. AKAP12 interacted with heat shock protein 47 (HSP47), which chaperones collagen and induces its secretion. Ethanol inhibited AKAP12-HSP47 and induced HSP47-collagen interaction. Ethanol-induced phosphorylated AKAP12 was unable to bind to HSP47 compared with its unphosphorylated counterpart, thereby proving that ethanol-mediated phosphorylation of AKAP12 inhibited the HSP47-AKAP12 scaffold. Silencing AKAP12 facilitated the chaperoning of collagen by HSP47. Hence, AKAP12 scaffolds HSP47 and regulates collagen-HSP47 interaction. Ethanol quenches AKAP12's scaffolding activity through phosphorylation and facilitates HSC activation.

Topics: A Kinase Anchor Proteins; Animals; Cell Cycle Proteins; Collagen; Cyclin D1; Ethanol; Hepatic Stellate Cells; Hepatocytes; Humans; Liver; Liver Diseases, Alcoholic; Mice; Phosphorylation; Protein Kinase C

Both human and rodent females are more susceptible to developing alcoholic liver disease following chronic ethanol (EtOH) ingestion. However, little is known about the relative effects of acute EtOH exposure on hepatotoxicity in female versus male mice. The nuclear receptor pregnane X receptor (PXR; NR1I2) is a broad-specificity sensor with species-specific responses to toxic agents. To examine the effects of the human PXR on acute EtOH toxicity, the responses of male and female PXR-humanized (hPXR) transgenic mice administered oral binge EtOH (4.5 g/kg) were analyzed. Basal differences were observed between hPXR males and females in which females expressed higher levels of two principal enzymes responsible for EtOH metabolism, alcohol dehydrogenase 1 and aldehyde dehydrogenase 2, and two key mediators of hepatocyte replication and repair, cyclin D1 and proliferating cell nuclear antigen. EtOH ingestion upregulated hepatic estrogen receptor α, cyclin D1, and CYP2E1 in both genders, but differentially altered lipid and EtOH metabolism. Consistent with higher basal levels of EtOH-metabolizing enzymes, blood EtOH was more rapidly cleared in hPXR females. These factors combined to provide greater protection against EtOH-induced liver injury in female hPXR mice, as revealed by markers for liver damage, lipid peroxidation, and endoplasmic reticulum stress. These results indicate that female hPXR mice are less susceptible to acute binge EtOH-induced hepatotoxicity than their male counterparts, due at least in part to the relative suppression of cellular stress and enhanced expression of enzymes involved in both EtOH metabolism and hepatocyte proliferation and repair in hPXR females.

Topics: Alcohol Dehydrogenase; Alcohol Drinking; Aldehyde Dehydrogenase; Aldehyde Dehydrogenase, Mitochondrial; Animals; Cyclin D1; Cytochrome P-450 CYP2E1; Endoplasmic Reticulum Stress; Estrogen Receptor alpha; Ethanol; Female; Hepatocytes; Humans; Lipid Peroxidation; Liver; Liver Diseases, Alcoholic; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Pregnane X Receptor; Receptors, Steroid; Sex Characteristics