Repression of Cytochrome P450 Activity in Human Hepatocytes in Vitro by a Novel Hepatotrophic Factor, Augmenter of Liver Regeneration

Abstract

Pathological disorders of the liver were shown to be associated with an impairment of hepatic drug metabolism mediated in part by growth factors. Augmenter of liver regeneration (ALR) is a novel liver-specific hepatotrophic growth factor, whereas its action on cytochrome P450 (P450) metabolism is completely unknown. Application of ALR to primary human hepatocytes in vitro reduced P450 isoenzyme activities (1A2 and 2A6) in a dose-dependent manner. Time-course analysis revealed that the maximal inhibitory effect was reached after 24 to 72 h of exposure with 50 nM ALR. The reduction of basal activities upon ALR treatment was 35% for CYP1A2, 56% for CYP2A6, 18% for CYP2B6, and 45% for CYP2E1. Additionally, after induction of P450 with specific inducers, ALR revealed an inhibitory effect on the isoenzyme activities (CYP1A2, 41%; CYP2B6, 35%). Investigations of protein and mRNA expression of basal and induced CYP1A2 and CYP3A4 after ALR treatment by Western blotting and real-time reverse transcriptase-polymerase chain reaction, respectively, suggest a regulation on the transcriptional level. Furthermore, ALR treatment increased nuclear factor kB activity and reduced constitutive androstane receptor but not pregnane X receptor or aryl hydrocarbon receptor expression. In contrast, ALR revealed no effects on phase II reactions (glutathione/oxidized glutathione, UDP-glucuronyltransferase conjugation). Our results indicate that ALR, as a member of hepatotrophic factors, down-regulates basal and induced P450 in human liver and therefore cross-links growth signals to regulation of hepatic metabolism. These findings further imply a possible role of ALR in drug interactions during impaired hepatic function, whereas liver regeneration is triggered.

Liver Regeneration Associated Protein (ALR) Exhibits Antimetastatic Potential in Hepatocellular Carcinoma

“…Recently, we reported that ALR induced c-myc expression and activated polyamine metabolism with both being critically important in cellular growth (17). Furthermore this isoform modulated hepatic detoxification by cross-linking growth signals to the regulation of hepatic metabolism (18). The proliferation augmenting effect of ALR was attributed to its ability to activate EGF receptor phosphorylation and the subsequent stimulation of the MAPK cascade (29).…”

“…The short form of ALR (15 kDa) is found not only at extracellular sites but also at nuclear and cytosolic localizations participating in intracellular redox-dependent signaling pathways (15,16). ALR is a hepatotrophic factor stimulating proliferation of hepatocytes (17,18) and augmenting liver regeneration (15,16), thereby exerting beneficial effects in models of hepatic failure (19) and liver fibrosis (20).…”

Hepatitis B Virus Induces Expression of Antioxidant Response Element-regulated Genes by Activation of Nrf2

“…Primary human hepatocytes were isolated by a modified two-step EGTA/collagenase perfusion procedure as described previously (19,20) and infected with HepAD38-derived supernatant as described previously (21). Primary mouse hepatocytes were isolated from Nrf2 Ϫ/Ϫ (22) and WT C57BL/6 mice using a two-step collagenase perfusion and cultivated as described previously (23).…”

Insulin Decreases Inflammatory Signal Transcription Factor Expression in Primary Human Liver Cells after LPS Challenge

“…Experimental procedures were performed according to the guidelines of the charitable state-controlled foundation, Human Tissue and Cell Research (HTCR), with informed patient's consent (12) and approved by the local ethics committee of the University of Regensburg. Human hepatocytes were isolated using a modified two-step EGTA/ collagenase perfusion procedure and maintained in culture as described (13,14). Viability of isolated hepatocytes was determined by trypan blue exclusion, and cells with viability >85% were used for cell culture.…”