APOC3 polymorphisms and non-alcoholic fatty liver disease: Resolving some doubts and raising others
Authors: Manuel Romero-Gómez
Journal: Journal of Hepatology (2011)

The APOC3 T-455C and C-482T promoter region polymorphisms are not associated with the severity of liver damage independently of PNPLA3 I148M genotype in patients with nonalcoholic fatty liverJournal of HepatologyVol. 55Issue 6PreviewThe T-455C and C-482T APOC3 promoter region polymorphisms (SNPs) have recently been reported to predispose to dyslipidemia, insulin resistance, and nonalcoholic fatty liver disease (NAFLD) in Indian subjects, but the association with liver damage has not been evaluated so far. The aim was to assess the association between APOC3 SNPs and liver damage in Caucasian patients. Full-Text PDF [genes]so taken one by one they are like dust, they are nothingJosé Agustín Goytisolo Non-alcoholic fatty liver disease (NAFLD) is the histological expression of different assaults on the liver, including lipid, glucose abnormalities, and obesity. The injury is indistinguishable from that caused by alcohol. Susceptibility and natural history of NAFLD seem to be modulated by gene-environment interactions. However, our current knowledge of the genetic traits involved in NAFLD is limited. In the current issue of the Journal of Hepatology, Valenti et al. [[1]Valenti L. Nobili V. Al-Serri A. Rametta R. Leathart J.B.S. Zappa M.A. et al.The APOC3 T-455C and C-482T promoter region polymorphisms are not associated with the severity of liver damage independently of PNPLA3 I148M genotype in patients with non-alcoholic fatty liver.J Hepatol. 2011; 55: 199-204Abstract Full Text Full Text PDF Scopus (65) Google Scholar], report a well-powered study on a large cohort of European patients with biopsy-proven NAFLD that showed lack of association between apolipoprotein C3 (APOC3) promoter SNPs (T-455C and C-482T) and the severity of liver damage, even after controlling for PNPLA3 mutation. Apolipoprotein C3 (ApoC3) is a 9 kDa protein inhibitor of lipoprotein lipase, which modulates the binding of chylomicron remnants and very-low-density lipoproteins (VLDL) to low-density-lipoproteins (LDL) receptors. Lipoprotein lipase hydrolyses triacylglycerols in chylomicrons, VLDL, LDL, and diacylglycerols. Thus, ApoC3 decreases the uptake of triglyceride-rich particles by liver cells for subsequent degradation. APOC3, located in the chromosome 11, is a good candidate gene because, in the original study [[2]Petersen K.F. Dufour S. Hariri A. Nelson-Williams C. Foo J.N. Zhang X.M. et al.Apolipoprotein C3 gene variants in nonalcoholic fatty liver disease.N Engl J Med. 2010; 362: 1082-1089Crossref PubMed Scopus (325) Google Scholar], both SNPs were associated with fat deposition in Indian men despite not having risk factors for hepatic steatosis, and was validated in a similar cohort of non-Asian men. Moreover, APOC3 promoter region polymorphisms C-482T and T-455C were found to be associated with lipid metabolism and insulin signalling and previously with metabolic syndrome (MetS) and hypertriglyceridaemia [[3]Miller M. Rhyne J. Chen H. Beach V. Ericson R. Luthra K. et al.APOC3 promoter polymorphisms C-482T and T-455C are associated with the metabolic syndrome.Arch Med Res. 2007; 38: 444-451Crossref PubMed Scopus (75) Google Scholar]. Further confirmation came from a meta-analysis indicating that the functional −455 T>C promoter polymorphism in APOC3 was associated with an approximately 2-fold increased risk in MetS [[4]Pollex R.L. Ban M.R. Young T.K. Bjerregaard P. Anand S.S. Yusuf S. et al.Association between the −455T>C promoter polymorphism of the APOC3 gene and the metabolic syndrome in a multi-ethnic sample.BMC Med Genet. 2007; 8: 80Crossref PubMed Scopus (41) Google Scholar] (Table 1). Differences in genetic analyses and methods to define phenotypes could explain, at least in part, differences in results between both studies: (a) Petersen et al. [[2]Petersen K.F. Dufour S. Hariri A. Nelson-Williams C. Foo J.N. Zhang X.M. et al.Apolipoprotein C3 gene variants in nonalcoholic fatty liver disease.N Engl J Med. 2010; 362: 1082-1089Crossref PubMed Scopus (325) Google Scholar] classified patients according to content of liver fat using proton magnetic resonance spectroscopy while Valenti et al. [[1]Valenti L. Nobili V. Al-Serri A. Rametta R. Leathart J.B.S. Zappa M.A. et al.The APOC3 T-455C and C-482T promoter region polymorphisms are not associated with the severity of liver damage independently of PNPLA3 I148M genotype in patients with non-alcoholic fatty liver.J Hepatol. 2011; 55: 199-204Abstract Full Text Full Text PDF Scopus (65) Google Scholar] used histological analysis of liver biopsies. Fat deposition does not necessarily mean liver disease. Indeed, triglyceride deposition per se is not directly injurious to hepatocytes, as demonstrated in patients with abetalipoproteinaemia who have high levels of triglycerides in the liver but with little or no injury. High high-fat diets in transgenic mice that overexpress ApoC3 had greater accumulation of liver diacylglycerol and this was associated with hepatic insulin resistance [[5]Samuel V.T. Petersen K.F. Shulman G.I. Lipid-induced insulin resistance: unravelling the mechanism.Lancet. 2010; 375: 2267-2277Abstract Full Text Full Text PDF PubMed Scopus (806) Google Scholar]. Dissociation of liver fat content from steatohepatitis should be borne in mind when using proton MRS as diagnostic tool for NAFLD. This limitation is the most commonly observed when defining phenotypes across the majority of published studies; (b) Valenti et al. included adult patients with altered liver function test values, morbidly obese patients, and children with diabetes or obesity; whereas Petersen et al. included a well-selected cohort of patients without "apparent" risk factors of NAFLD; (c) Ethnic origin did influence NAFLD prevalence and, thus, the diagnostic accuracy of genetic markers depends on the race studied. Recent studies confirmed lack of association between APOC3 promoter polymorphisms and fat deposition in patients with and without type 2 diabetes mellitus [6Kozlitina J. Boerwinkle E. Cohen J.C. Hobbs H.H. Dissociation between APOC3 variants, hepatic triglyceride content and insulin resistance.Hepatology. 2011; 53: 467-474Crossref PubMed Scopus (97) Google Scholar, 7Petit JM, Guiu B, Masson D, Cercueil JP, Hillon P, Verges B. Lack of association between an apoC3 genetic variant and liver fat content in patients with type 2 diabetes. Hepatology, in press.Google Scholar]. Of note is that the study by Valenti et al. addressed the heritability of NAFLD in 71 Italian trios and reported a marginal away association. Heritability of fatty liver has been reported as between 39% and 58%, a range commonly seen in frequent traits [[8]Schwimmer J.B. Celedon M.A. Lavine J.E. Salem R. Campbell N. Schork N.J. et al.Heritability of nonalcoholic fatty liver disease.Gastroenterology. 2009; 136: 1585-1592Abstract Full Text Full Text PDF PubMed Scopus (324) Google Scholar], but these promoter polymorphisms appear not to be implicated.Table 1Influence of two main SNPs in the promoter region of APOC3 in different phenotypes associated with NAFLD. Percentage of fat accumulation in the liver, steatosis, non-alcoholic steatohepatitis, fibrosis progression, and metabolic syndrome. Odds Ratios (effect), and the corresponding 95% CI limits (lower and upper) and significance (p value) were calculated by comparing fatty liver, NASH, fibrosis >F1, and metabolic syndrome as dichotomized variables segregated according to APOC3 variants. Validation studies confirmed an association between APOC3 polymorphisms and metabolic syndrome but not steatosis, steatohepatitis or fibrosis progression. Open table in a new tab Several types of genetic studies on NAFLD have been reported including candidate genes, genome wide association studies (GWAS), and bioinformatics-based analyses. Initial studies of association between candidate genes and disease traits increase the possibility of a subsequent positive replication of the findings when odds ratios are >3, the cohort includes >150 individuals per phenotype trait and meta-analysis would confer further validation [[9]Ioannidis J.P. Ntzani E.E. Trikalinos T.A. Contopoulos-Ioannidis D.G. Replication validity of genetic association studies.Nat Genet. 2001; 29: 306-309Crossref PubMed Scopus (1614) Google Scholar]. A long list of candidate genes in NAFLD reaching negative association has been reported [10Kantartzis K. Schick F. Häring H.U. Stefan N. Environmental and genetic determinants of fatty liver in humans.Dig Dis. 2010; 28: 169-178Crossref PubMed Scopus (32) Google Scholar, 11Romero-Gómez M. Saenz-Goñi M.E. The genetics of non-alcoholic steatohepatitis and insulin resistance.in: Genetics in liver disease. vol. 2. Permanyer Publications, 2007: 1-14Google Scholar]. Moreover, the candidate gene approach does not take into account the interaction between genetic signals. New genotype research technologies have provided incontrovertible evidence that genomic variation in patatin-like phospholipase domain-containing 3 (PNPLA3) or adiponutrin gene rs738409 C>G polymorphism located in chromosome 22 is associated with fat accumulation in the liver [[12]Romeo S. Kozlitina J. Xing C. Pertsemlidis A. Cox D. Pennacchio L.A. et al.Genetic variation in PNPLA3 confers susceptibility to nonalcoholic fatty liver disease.Nat Genet. 2008; 40: 1461-1465Crossref PubMed Scopus (2121) Google Scholar]. A recent meta-analysis confirmed the association of genotype CC not only with increased risk of steatosis, but also with fibrosis progression i.e. patients with the CC genotype showed 3.2-fold higher risk of developing fibrosis [[13]Sookoian S, Pirola JC. Meta-analysis of the influence of I148M variant of patatin-like phospholipase domain containing 3 gene (PNPLA3) on the susceptibility and histological severity of nonalcoholic fatty liver disease. Hepatology, in press.Google Scholar]. However, no GWAS has been conducted in biopsy-proven NAFLD patients yet. In addition to information obtained using genome-wide scans and the candidate gene approach, bioinformatic analyses in search of new genetic markers for NAFLD have yielded negative results [[14]Banasik K. Justesen J.M. Hornbak M. Krarup N.T. Gjesing A.P. Sandholt C.H. et al.Bioinformatics-driven identification and examination of candidate genes for non-alcoholic fatty liver disease.PLoS One. 2011; 6: e16542Crossref PubMed Scopus (23) Google Scholar]. Using several databases, 247 genes were selected and, after removing 26 due to symbol overlap, 37 genes showed a priority score >2 based on the data source and OMIM phenotypes. Five genes remained within the centre of the interactome and showed 5 or more connections with other nodes in the network. These 5 genes (peroxisomal bifunctional enzyme, mitochondrial enoyl-CoA hydratase, mitochondrial long-chain-specific acyl-CoA dehydrogenase, mitochondrial trifunctional enzyme subunit alpha, and mitochondrial trifunctional enzyme subunit beta), which are completely different from the list of candidate genes or genetic signals extracted from genome-wide association studies, were tested in a cohort of 10,196 patients with different metabolic disorders. NAFLD was assessed using surrogate quantitative traits, such as waist circumference, serum triglyceride, fasting insulin, and plasma glucose, but not liver biopsy. No statistically significant associations remained after Bonferroni corrections. To better evaluate the contribution of our genes to NAFLD we need to conduct genome-wide association studies in large biopsy-proven patient populations. Histological end-points are preferred because these are better associated with overall prognoses in these patients. Histological phenotypes should include simple steatosis, non-alcoholic steatohepatitis, and fibrosis stage. Any potential genetic signal would need to clearly segregate patients according to the presence or absence of these phenotypes. Indeed, inflammation is a major predictor of fibrosis progression [[15]Argo C.K. Northup P.G. Al-Osaimi A.M. Caldwell S.H. Systematic review of risk factors for fibrosis progression in non-alcoholic steatohepatitis.J Hepatol. 2009; 51: 371-379Abstract Full Text Full Text PDF PubMed Scopus (407) Google Scholar] and advanced fibrosis independently predicts liver-related mortality [[16]Younossi Z. Stepanova M. Rafiq N. Makhlouf H. Younoszai Z. Agrawal R. et al.Pathologic criteria for Non-alcoholic steatohepatitis (NASH): Inter-protocol agreement and ability to predict liver-related mortality.Hepatology. 2011; 53: 1874-1882Crossref PubMed Scopus (440) Google Scholar]. However, this task would seem to be further complicated since PNPLA3 I148M did not distinguish simple steatosis from non-alcoholic steatohepatitis, despite a strong association with susceptibility and fibrosis progression [[17]Speliotes E.K. Butler J.L. Palmer C.D. Voight B.F. et al.GIANT Consortium, MIGen Consortium, PNPLA3 variants specifically confer increased risk for histologic nonalcoholic fatty liver disease but not metabolic disease.Hepatology. 2010; 52: 904-912Crossref PubMed Scopus (286) Google Scholar]. Also, this approach could provide information about new therapeutic targets for this major cause of chronic liver disease which, to date, remains without effective treatment. There is still a considerable gap between genetic discoveries and application of the findings to innovative clinical practice. Fortunately, this "laboratory bench to bedside" gap is slowly closing. The author declared he does not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. The research leading to these results has received funding from the European Union Seventh Framework Programme (FP7/2007-2013) under Grant agreement Health-F2-2009-241762, for the project FLIP.