Hepatocyte senescence explains conjugated bilirubinaemia in chronic liver failure
Authors: Aloysious D. Aravinthan, Graeme J.M. Alexander
Journal: Journal of Hepatology (2015)

Conjugated bilirubinaemia in patients with chronic liver disease (CLD) reflects hepatic decompensation and a poor prognosis [[1]Gines P. Quintero E. Arroyo V. Teres J. Bruguera M. Rimola A. et al.Compensated cirrhosis: natural history and prognostic factors.Hepatology. 1987; 7: 122-128Crossref PubMed Scopus (921) Google Scholar]. The pathophysiology that underlies conjugated bilirubinaemia in hepatic decompensation is poorly understood. There is no demonstrable flaw in processing unconjugated bilirubin and a more likely explanation is altered hepatocyte handling of conjugated bilirubin. Hepatocyte senescence is present across diverse aetiologies and as many as 80% of hepatocytes show the senescent phenotype in advanced liver disease [[2]Aravinthan A. Scarpini C. Tachtatzis P. Verma S. Penrhyn-Lowe S. Harvey R. et al.Hepatocyte senescence predicts progression in non-alcohol-related fatty liver disease.J Hepatol. 2013; 58: 549-556Abstract Full Text Full Text PDF PubMed Scopus (194) Google Scholar]. Metabolic activity is altered when a cell becomes senescent and one potential consequence is an alteration of conjugated bilirubin transport in senescent hepatocytes, which accumulate in advanced CLD. Serum bilirubin and hepatocyte telomere length were measured in 70 patients within the spectrum of NAFLD. Mean hepatocyte telomere intensity, a surrogate marker of telomere length, was measured using quantitative fluorescent in-situ hybridization, as described [[3]Verma S. Tachtatzis P. Penrhyn-Lowe S. Scarpini C. Jurk D. Von Zglinicki T. et al.Sustained telomere length in hepatocytes and cholangiocytes with increasing age in normal liver.Hepatology. 2012; 56: 1510-1520Crossref PubMed Scopus (50) Google Scholar]. There was an inverse relationship between serum bilirubin and hepatocyte telomere length (p= 0.04, Fig. 1). Thus, accelerated hepatocyte ageing is associated with jaundice. Liver sections from five of those patients were double-stained using unconjugated mouse monoclonal anti-p21 (Dako; concentration 1:100, heat-induced EDTA-based antigen retrieval, 20 min) and unconjugated mouse monoclonal anti-MRP2 (Merck Millipore; concentration 1:20, heat-induced citrate-based antigen retrieval, 20 min). MRP2 was negative in p21-positive (senescent) hepatocytes and was only detected in p21-negative hepatocytes (Fig. 1). Reliable immunohistochemical staining could not be achieved with available MRP3 antibodies. An in vitro model was used to examine gene expression of MRP2 and MRP3 in senescent hepatocytes by real-time PCR. Cellular senescence was induced in HepG2 cells by incubation with 0.5 mM H2O2 in culture medium for 60 minutes, as described [[4]Aravinthan A. Shannon N. Heaney J. Hoare M. Marshall A. Alexander G.J. The senescent hepatocyte gene signature in chronic liver disease.Exp Gerontol. 2014; 60: 37-45Crossref PubMed Scopus (34) Google Scholar]. Expression of MRP2 was downregulated in senescent HepG2 cells; in contrast, expression of MRP3 was upregulated (Fig. 1). Hepatocytes are polarised cells; MRP2 is restricted to the canalicular (apical) membrane, whereas MRP3 is found only in the sinusoidal (basolateral) membrane [[5]Keppler D. The roles of MRP2, MRP3, OATP1B1, and OATP1B3 in conjugated hyperbilirubinemia.Drug Metab Dispos. 2014; 42: 561-565Crossref PubMed Scopus (153) Google Scholar]. Both MRP2 and MRP3 are unidirectional efflux pumps, which transport conjugated bilirubin into the canalicular space (bile) or the sinusoid (blood), respectively [[5]Keppler D. The roles of MRP2, MRP3, OATP1B1, and OATP1B3 in conjugated hyperbilirubinemia.Drug Metab Dispos. 2014; 42: 561-565Crossref PubMed Scopus (153) Google Scholar]. Reduced MRP2 expression in senescent hepatocytes in vitro and an absence of MRP2 protein in p21-positive (senescent) hepatocytes suggest reduced conjugated bilirubin transport into the biliary canaliculi. Increased MRP3 expression in senescent hepatocytes may be compensatory, increasing transport of conjugated bilirubin into the hepatic sinusoid (Fig. 1). It is, however, not clear why changes in MRP2 and MRP3 accompany hepatocyte senescence. The authors declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. Addenbrooke's Charitable Trust, Cambridge, UK.