In this study, we explored for the first time epigenetic aging within multiple datasets and tissue types of individuals with ALC and controls. We observed positive age acceleration in one blood dataset and one liver tissue dataset, but not in brain tissue. These results are particularly interesting, given the known negative effect of prolonged excessive alcohol consumption on these parts of the body1.

The positive age acceleration trend observed in each of the cirrhotic liver samples was consistent with our hypothesis. However, we expected to see a greater magnitude of positive age acceleration, as both datasets consisted of samples from individuals with alcoholic cirrhosis, a severe end-stage disease caused by particularly heavy alcohol consumption. Analyses conducted in such tissue must be interpreted with caution, as hepatocytes found in cirrhotic liver tissue differ significantly from healthy hepatocytes and DNA methylation might vary5. In fact, previous research shows that the process of alcoholic cirrhosis may directly influence DNA methylation. In addition, cell composition differences in cirrhotic liver versus normal liver might account for changes in DNA methylation age, as not only the hepatocytes, but also cells linked to fibrosis and inflammation can be abnormal in alcohol cirrhosis25. One way to address this limitation would be the use of Andres Housman’s reference-free method epigenetic clock, which accounts for cell type-specific DNA methylation effects and different cell proportions26. It remains unknown to what degree progressive alcoholic liver damage correlates with increased epigenetic aging, as longitudinal liver biopsy samples from individuals across the life span are not available to our knowledge. Although both liver datasets show age acceleration in alcohol cirrhosis, future larger studies are needed that in particular take into account various demographic covariates as well as cell-specific analyses to confirm this finding.

ALC is strongly associated with degeneration of brain tissue as well as premature cognitive decline3,4,27,28. Thus, we expected to find positive age acceleration in PFC tissue, rather than the negative age acceleration that we observed. However, there may be an explanation for these findings; of our sample of 23 participants, 14 (61%) were diagnosed with DSM-IV alcohol abuse and only nine (39%) were diagnosed with ALC. Alcohol abuse is a milder phenotype than ALC and is thus associated with significantly lower levels of consumption. In fact, it is possible that this sample is more representative of a population that consumes alcohol at moderate levels, which some research suggests may be associated with healthy aging, reduced risk of coronary heart disease, and reduced risk of cognitive decline and Alzheimer’s disease29,30,31,32,33,34. Future larger studies should confirm the effects of alcohol on epigenetic age in PFC tissue and might also explore additional brain regions, as results in our PFC sample may not represent those in other regions.

Studying epigenetic aging in the blood is important, as such results may be combined with those from other tissues to eventually permit the use of DNA methylation signatures in blood as a useful biomarker of aging and to assess age acceleraton across the life span. The difference in results between the NIAAA sample and Grady Trauma project sample may be explained by the multi-year difference in age both within and between the two samples13. In addition, the NIAAA sample was characterized by a current ALC diagnosis, as well as current desire for inpatient treatment, while the Grady Trauma Project was characterized by a diagnosis of ALC anytime in one’s lifetime. Thus, the NIAAA sample may represent a more severe phenotype than the Grady Trauma project sample. In fact, previous research indicates that differences in genome-wide DNA methylation in individuals with ALC and controls may be diminished following periods of abstinence35. In addition, false reporting is common when patients are asked to self-report alcohol use, and may significantly influence the make-up of the case and control groups. This highlights the importance of developing useful biomarkers to measure alcohol use36. Finally, the Grady Trauma project was primarily a study of childhood trauma, which may explain the small difference between cases and controls. We did not control for this possible covariate but it might have an effect on our results as a previous epigenetic clock analysis using this data found that cumulative lifetime stress was associated with accelerated aging37.

Further analyses would benefit from increased statistical power. Many of our datasets had sample sizes ranging from 25 to 50 individuals per group, which may partially explain or influence these findings. In addition, clinical heterogeneity remains a concern for studies of individuals with substance use disorders and the comparison of multiple datasets can be problematic. For example, the average chronological age in the NIAAA blood sample was approximately 15 years younger than that of the postmortem PFC sample and two liver cirrhosis samples. In addition, ALC has been shown to be associated with a multitude of comorbidities including cardiovascular disease, liver disease, smoking, and other substance use disorders, as well as psychiatric disorders, such as major depressive disorder and bipolar I disorder1. Previous studies suggest that comorbidities and other lifestyle factors, in combination with the medication used to treat them, are likely to influence epigenetic aging and should be controlled for in future analyses13,15.

Given that the epigenetic clock is valid across different tissue types, collecting blood, liver, and postmortem brain tissue samples from the same individuals would not only remedy this problem of clinical heterogeneity, but also allow us to take full advantage of the capabilities of the epigenetic clock. Furthermore, it would be beneficial to acquire data on race and ethnicity and specific drinking patterns/abstinence in these samples, as well as to create a more equal balance of males and females, as recent research has shown that both race and sex may play a role in the epigenetic aging process13,38. Finally, future studies of epigenetic aging in ALC might consider analysis of longitudinal data, especially after a period of treatment or sobriety, to investigate the stability of age acceleration in different tissue types in this population.

In conclusion, the present study provided evidence that epigenetic aging differs in blood and liver tissue of individuals with ALC compared to healthy volunteers. Although there were limitations in this study, these results warrant further investigation into the role of chronic, heavy alcohol consumption in the aging process across various tissue types.