Petersen KF, Dufour S, Hariri A, Nelson-Williams C, Foo JN, Zhand XM, et al. Apolipoprotein C3 gene variants in nonalcoholic fatty liver disease. N Engl J Med 2010;362:1082-1089. (Reprinted with permission.)
Background: Nonalcoholic fatty liver disease is associated with hepatic insulin resistance and type 2 diabetes mellitus. Whether this association has a genetic basis is unknown. Methods: In 95 healthy Asian Indian men, a group known to have a high prevalence of nonalcoholic fatty liver disease, we genotyped two single-nucleotide polymorphisms (SNPs) in the gene encoding apolipoprotein C3 (APOC3) that are known to be associated with hypertriglyceridemia (rs2854116 [T-455C] and rs2854 117 [C-482T]). Plasma apolipoprotein C3 concentrations, insulin sensitivity, and hepatic triglyceride content were measured. We also measured plasma triglyceride concentrations and retinyl fatty acid ester absorption as well as plasma triglyceride clearance after oral and intravenous fat-tolerance tests. Liver triglyceride content and APOC3 genotypes were also assessed in a group of 163 healthy non-Asian Indian men. Results: Carriers of the APOC3 variant alleles (C-482T, T-455C, or both) had a 30% increase in the fasting plasma apolipoprotein C3 concentration, as compared with the wild-type homozygotes. They also had a 60% increase in the fasting plasma triglyceride concentration, an increase by a factor of approximately two in the plasma triglyceride and retinyl fatty acid ester concentrations after an oral fat-tolerance test, and a 46% reduction in plasma triglyceride clearance. The prevalence of nonalcoholic fatty liver disease was 38% among variant-allele carriers and 0% among wild-type homozygotes (P<0.001). The subjects with nonalcoholic fatty liver disease had marked insulin resistance. A validation study involving non-Asian Indian men confirmed the association between APOC3 variant alleles and nonalcoholic fatty liver disease. Conclusions: The polymorphisms C-482T and T-455C in APOC3 are associated with nonalcoholic fatty liver disease and insulin resistance.
Burgeoning waistlines and a more sedentary lifestyle have resulted in nonalcoholic fatty liver disease (NAFLD) becoming the most common cause of chronic liver disease in the Western world. The prevalence of NAFLD has been reported to be as high as 46% in the United States and is associated with obesity, diabetes, metabolic syndrome, and certain ethnicities, including Hispanics and Asian Indians.1 Interestingly, the prevalence of NAFLD is lower among African Americans despite high rates of diabetes, obesity, and metabolic syndrome.2 In addition, many patients with sedentary lifestyles (with or without diabetes) may not develop fatty liver disease. This suggests that other factors are involved in the development of hepatic steatosis and steatohepatitis.
The study by Petersen et al.3 in the March issue of the New England Journal of Medicine provides evidence that genotypic variations and particularly the genes encoding apolipoprotein C3 (APOC3) are important in the development of hepatic steatosis. This gene was selected for study because of its known association with hypertriglyceridemia, and Asian Indians were selected as the study cohort because of their high prevalence of NAFLD.4 Specifically, the authors genotyped two single-nucleotide polymorphisms (SNPs) in the APOC3 gene (rs2854116 and rs2854117) and showed that the presence of one or both of these alleles was associated with a 30% increase in the fasting plasma APOC3 concentration and a 60% increase in the fasting plasma triglyceride concentration. Using magnetic resonance spectroscopy, they further showed that 38% of the Asian Indian study subjects with variant alleles had detectable hepatic steatosis versus 0% of the wild-type Asian Indian study subjects. A control population of mixed ethnicities showed that 9% of the subjects possessing the variant allele had detectable hepatic steatosis versus 0% of those possessing the wild-type genotype.
The implications of these findings are 2-fold. First, APOC3 and plasma triglyceride concentrations appear to be important in the development of hepatic steatosis, even in the absence of obesity. All patients possessing variant alleles for the APOC3 gene had markedly higher average hepatic triglyceride contents (7.5% ± 10.3% versus 1.5% ± 1.3%). In fat-tolerance testing of a subgroup of variant-allele and wild-type individuals, variant-allele subjects were also found to have significantly higher levels of plasma triglyceride. Similarly, plasma triglyceride clearance was decreased in a subgroup of patients with the variant allele versus their wild-type counterparts. In all, this provides compelling evidence that APOC3 can contribute to triglyceride excess both intrahepatically and systemically.
The second and more novel finding of these authors is the genetic association between Asian Indian men and this polymorphism. This population has a high prevalence of NAFLD, and this is the first study to associate a specific genetic variation with hepatic steatosis in healthy men of Asian Indian descent. This finding was substantiated in an independent group of non-Asian men, and the authors noted that no wild-type homozygotes at this allele were found to have hepatic steatosis by magnetic resonance spectroscopy.
It is important to understand that the frequency of variant alleles was no greater in the Asian Indian study population versus the control group composed of multiple ethnicities, and this suggests that this APOC3 variant allele does not explain the high rates of hepatic steatosis among Asian Indians. Other genetic variations, such as that encoding the patatin-like phospholipase domain-containing 3 protein (PNPLA3), have also been independently associated with NAFLD,5 with recent research linking specific polymorphisms to disease severity.6-8 In the study by Petersen etal.,3 PNPLA3 variants were also shown to be associated with NAFLD, and the relative risk appeared additive in those individuals having both PNPLA3 and APOC3 variants. However, only 13.1% of the variance in risk for NAFLD could be explained by the combination of these two different SNPs, and this suggests that additional factors may be involved in the development of NAFLD.
The authors also selected seven subjects with hepatic steatosis and insulin resistance (IR) for enrollment in a 6-month weight-loss program; during this time, they experienced a mean weight loss of approximately 6 kg along with a significant reduction in their hepatic triglyceride content from 14.0% to 3.8% (P = 0.05) and an improvement in the insulin sensitivity index from 1.8 to 3.7 (P < 0.01). This provides further evidence that although genetic variations may predispose someone to a fatty liver, dietary habits and activity levels remain important cofounders in the development of NAFLD and provide the first step in disease prevention, even in normal-weight individuals with NAFLD.
The intriguing results described by Petersen et al.3 provide a foundation for further study. Several questions remain to be answered, however. What factors, genetic or otherwise, allow the development of steatohepatitis and hepatic fibrosis, and what role does IR play in this process? The G allele variant of PNPLA3 has been shown to be associated with the severity of NAFLD but is not associated with IR.8 Conversely, the SNPs in APOC3 have now been linked to NAFLD and IR, but so far, there are no data linking APOC3 variations to the severity of NAFLD. A future study combining tests for multiple SNPs linked to NAFLD with hepatic histology is essential to determine the relationship between this and other genetic variations and NAFLD/nonalcoholic steatohepatitis (NASH). Additionally, it is possible that there are specific genetic SNPs that confer protection against hepatic steatosis, steatohepatitis, or both. It can be postulated that in fact this is the case in patients who are phenotypically predisposed to NAFLD but do not have hepatic steatosis.
The role of IR in hepatic steatosis and NAFLD has yet to be fully understood. It is known that patients with NAFLD have increased IR, both systemically and intrahepatically, but it remains uncertain if this is a cause or effect of hepatic steatosis. Petersen et al.3 suggested that in patients with a normal body mass index, genetic variation in APOC3 leads to hypertriglyceridemia, which in turn causes hepatic steatosis and consequently leads to IR. As evidence, they noted that among the members of their small group who lost weight, hepatic steatosis was reduced with a subsequent improvement in IR. This is by no means resounding proof of cause and effect, and recent data from mouse and human studies suggest that IR is independent of hepatic steatosis.9, 10 A complete picture of the interplay between hepatic steatosis and IR remains to be seen, but the reality is likely much more complex and involves not only triglyceride accumulation in the liver and IR but also the host's defense and repair responses to the potentially hepatotoxic triglyceride precursor molecules (Fig. 1).11 IR not only is a byproduct of hepatic triglyceride excess but also appears to promote hepatic endoplasmic reticulum stress, which may in turn lead to steatohepatitis and even fibrosis.12
In summary, Petersen et al.3 have opened the door to further research aimed at mapping genes associated with NAFLD and, most usefully, advanced disease as evidenced by NASH and fibrosis. This may lead to the ability to predict who is most at risk for progression to cirrhosis or even the development of liver cancer. Many questions, including the complicated relationship of hepatic steatosis and IR and the optimal treatment regimen for NASH patients, still need to be answered along the way.