Coronary artery disease (CAD) remains a leading cause of illness, disability, and mortality worldwide, despite advances in prevention and treatment strategies (1). Alarmingly, recent trends have shown a rise in premature coronary artery disease, also called early-onset CAD (EOCAD), among younger populations (2, 3). Individuals with EOCAD often exhibit a high prevalence of modifiable traditional risk factors such as dyslipidemia, smoking, diabetes mellitus, hypertension, and overweight/obesity (4–8). Non-classical risk factors including inflammation, infections, sleep deprivation, and exposure to air pollution and environmental noise also warrant consideration (1, 9).

Moreover, it is well known that premature CAD is characterized by a rapid progression of the disease and is associated with a poorer long-term prognosis (10, 11). Thus, further research is essential to better understand the risk profile, behaviors, and underlying biological mechanisms contributing to premature CAD in younger individuals (12, 13).

Interestingly, recent research has identified a significant association for the mitochondrial mtDNA 16223T variant and heteroplasmy with an increased presence of cardiovascular risk factors and premature CAD (14).

Indeed, several studies have highlighted the direct role of mtDNA-mediated mitochondrial dysfunction in the initiation and progression of atherosclerosis (15), and the importance of preserving the mitochondrial genome to maintain healthy vessel and to delay vascular aging (16).

Mitochondria, essential cellular organelles, are primarily responsible for energy production through ATP generation via oxidative phosphorylation. They also play key roles in various cellular processes, including modulation of oxidation–reduction status, regulation of senescence and apoptosis. Mitochondria possess their own circular genome, known as mitochondrial DNA (mtDNA), with each mitochondrion containing multiple copies of this genome, referred to as mtDNA copy number (mtDNA-CN). The mtDNA-CN is positively correlated with mitochondrial enzyme activity and ATP production, making it a surrogate biomarker for assessing mitochondrial function (17).

Lower mtDNA-CN has shown associations to several aging-related diseases and all-cause mortality, including cardiovascular disease (18).

Additionally, mtDNA is particularly vulnerable to damage from reactive oxygen species (ROS) due to the absence of histones and a less efficient repair mechanism. Besides rare pathogenic variants, the accumulation of mutations in the mitochondrial genome can further accelerate the development of atherosclerosis (19). Among various mtDNA defects, a large-scale deletion of 4,977 bp, known as the mtDNA⁴⁹⁷⁷ common deletion, results in the loss of nearly one-third of the mitochondrial genome. This leads to the accumulation of dysfunctional mitochondrial transcripts and a consequent increase in ROS production (20).

In light of this, we hypothesized that reduced mtDNA integrity and mitochondrial dysfunction may predispose individuals to accelerated atherosclerosis and the premature development of CAD. To test this hypothesis, we investigated the association of leukocyte mtDNA⁴⁹⁷⁷ deletion and mtDNA-CN with early-onset coronary artery disease.

To the best of our knowledge, this is the first attempt to explore the interaction between mitochondrial damage and dysfunction in blood, and early-onset of coronary artery disease. We identified an independent, significant association between both mtDNA⁴⁹⁷⁷ deletion and mtDNA-CN and incident EOCAD after accounting for traditional risk factors. In addition, our results showed significant associations between traditional cardiovascular risk factors and mitochondrial dysfunction, suggesting that the mtDNA damage and dysfunction induced by atherosclerosis-related risk factors may play a crucial role in the early initiation and progression of atherosclerosis (25). In addition, our results agree with previous findings suggesting that mitochondrial dysfunction may have a gender-specific component influenced by hormonal and metabolic factors. In fact, women's mitochondria are better able to deal with stressful situations and conditions, and are more resistant to DNA damage and mutations (17, 18).

Previous studies have found that lower mtDNA-CN in blood is consistently associated with higher risk of CAD, reflecting mitochondrial dysfunction and oxidative stress, both of which are critical in CAD pathogenesis (26).

In particular, three large prospective studies suggested that mtDNA-CN in circulating leukocytes may have clinical utility in improving CVD risk classification (27).

Interestingly, a case-control study also found a significant joint effect between low levels of mtDNA-CN and risk factors like plasma LDL-C or smoking with increasing the incidence of CAD (28).

Moreover, cardiometabolic risk factors such as high blood pressure, obesity, and dyslipidemia have been shown to impact mtDNA-CN (29–31).

Regarding mtDNA damage, previous studies showed that mtDNA⁴⁹⁷⁷ deletion accumulates with age in normal hearts (19, 20) and that the levels of mtDNA⁴⁹⁷⁷ deletion are increased in human atherosclerotic lesions (32).

Our previous study also reported significant associations between lower mtDNA⁴⁹⁷⁷ deletion levels and poorer CVD outcomes as well as increased all-cause mortality in patients with stable CAD, confirming the critical role of mitochondrial dysfunction in atherosclerosis (24).

Remarkably, experimental studies in animal models further support the role of mtDNA damage in atherosclerosis. For instance, mtDNA damage has been observed during the early stages of atherogenesis in ApoE-/- mice and correlates with the severity of the disease (33). In double knock-out ATM+/-/ApoE-/- mice, an increased frequency of the mouse mitochondrial “common” deletion was associated with impaired respiratory complex activity (34). These mitochondrial alterations accelerated the development of atherosclerosis and were linked to key metabolic syndrome features such as hypertension, hypercholesterolemia, and obesity (35).

In response to reactive oxygen species, human vascular endothelial and smooth muscle cells accumulate more mtDNA damage than nuclear DNA, impairing key cellular processes like growth, signaling, and apoptosis. This cell dysfunction can, in turn, contribute to the initiation and progression of atherosclerotic lesions (35).

In addition, point mtDNA mutations in blood cells, especially rare variants and heteroplasmy levels (a mixture of both wild-type and mutant species of mtDNA), are being actively investigated in relation to atherosclerosis, coronary heart disease and various atherosclerosis-related diseases such as arterial hypertension and diabetes mellitus, and the list of identified mutations continues to grow (14, 36, 37). These mutations cause structural and functional changes in the mtDNA OXPHOS-related and mt-tRNA genes, which in turn can affect mitochondrial protein synthesis and lead to mitochondrial dysfunction.

Altogether, our findings, along with the evidence summarized above, strongly suggest that mtDNA plays a major role in the pathogenesis of atherosclerosis directly regulating the onset and progression of early vascular aging (16).

In patients with early-onset CAD, we hypothesize that the cumulative burden of traditional clinical risk factors may lead to more aggressive premature atherosclerosis through mechanisms such as oxidative stress and sustained inflammation, which are central to the progression of the mtDNA damage and dysfunction.

These observations raise significant clinical questions regarding broader preventive strategies to reduce the risk of premature coronary artery disease. First, our study confirms a high rate of concomitant modifiable cardiovascular risk factors in individuals with EOCAD (4–8), emphasizing the importance of increased efforts to reduce the burden of these risk factors and premature clinical events (11).

Second, ROC curve analysis suggested that both mitochondrial damage and dysfunction might be considered a biomarker reflecting the early diagnosis and prediction of EOCAD The evaluation of mtDNA could also serve as an easy and accessible biomarker that could be exploited for cardiovascular prognosis or for assessing responses to therapies (24). The incorporation of mitochondrial biomarkers into traditional risk assessments may have the potential to improve the accuracy of prediction. For example, it may help to reclassify intermediate risk patients into higher or lower risk categories. However, their integration into clinical practice will depend on standardization of measurement techniques and confirmation of their predictive value by robust evidence.

Third, targeting of mitochondrial dysfunction may be a feasible strategy to provide a more personalized approach to vascular risk managemen (38). Potential therapies include mitochondrial antioxidative therapies and lifestyle interventions (e.g., exercise, diet) aimed at mitigating mitochondrial dysfunction and ROS production. In addition, mitochondrial biogenesis enhancers may offer promising avenues for treating vascular damage caused by mitochondrial dysfunction by repairing mtDNA, such as increasing the activity of mitochondrial-specific enzymes (e.g., OGG1), or enhancing mitochondrial biogenesis. In addition, new gene-editing techniques such as CRISPR hold promise to correct mtDNA mutations (38).

Our study has some potential limitations. First, the sample size of patients is relatively small, which limits the precision of the associations. As result, the estimated ORs with the wide confidence intervals for risk factors should be interpreted cautiously. Second, the case-control study design comes with inherent sources of confounding and unmeasured biases. For instance, we did not measure family history or potential social and psychological biases. Third, the cohort comprised patients admitted to the hospital for coronary angiography, and some already had existing CAD or previous myocardial infarction and were receiving pharmacological treatment. Finally, mtDNA⁴⁹⁷⁷ deletion and mtDNA-CN were measured only at enrollment, without capturing potential variability over time. Indeed, there is potential uncertainty about the temporal variability of biomarkers when relying on measurements from a single point in time. Mitochondrial biomarkers may fluctuate with circadian rhythms, transient physiological states or in response to environmental exposures, which could affect their predictive utility. Future studies should, therefore, be focused on assessing mitochondrial biomarkers longitudinally in order to evaluate their temporal dynamics and improve the accuracy of risk prediction.

In conclusion, this study revealed that premature EOCAD is characterized by a high prevalence of modifiable cardiovascular risk factors and elevated mitochondrial damage and dysfunction. This underscores the need for more comprehensive efforts to reduce the burden of traditional risk factors and highlights the potential for innovative mitochondrial-targeted therapies to address vascular damage resulting from mitochondrial dysfunction.