Mitochondrial DNA Copy Number as a Marker and Mediator of Stroke Prognosis: Observational and Mendelian Randomization Analyses

Background: Low buffy coat mitochondrial DNA copy number (mtDNA-CN) is associated with incident risk of stroke and post-stroke mortality; however, its prognostic utility has not been extensively explored. Objective: To investigate whether low buffy coat mtDNA-CN is a marker and causal determinant of post-stroke outcomes using epidemiological and genetic studies. Methods: First, we performed association testing between baseline buffy coat mtDNA-CN measurements and 1-month post-stroke outcomes in 3498 acute, first stroke cases from 25 countries from the international, multicenter case-control study, “Importance of Conventional and Emerging Risk Factors of Stroke in Different Regions and Ethnic Groups of the World” (INTERSTROKE). Then, we performed two-sample Mendelian Randomization analyses to evaluate potential causative effects of low mtDNA-CN on 3-month modified Rankin Scale (mRS). Genetic variants associated with mtDNA-CN levels were derived from the UKBiobank study (N=383476), and corresponding effects on 3-month mRS were ascertained from the Genetics of Ischemic Stroke funCtional Outcome study (GISCOME; N=6021). Results: A 1-standard deviation (SD) lower mtDNA-CN at baseline was associated with stroke severity (baseline mRS; OR=1.27; 95% CI, 1.19-1.36; P=4.7x10 -12 ). Independent of baseline stroke severity, lower mtDNA-CN was associated with increased odds of greater 1-month disability (ordinal mRS; OR=1.16; 95% CI, 1.08-1.24; P=4.4x10 -5 ), poor functional outcome status (mRS 3-6 vs. 0-2; OR=1.21; 95% CI, 1.08-1.34; P=6.9x10 -4 ), and mortality (OR=1.35; 95% CI, 1.14-1.59; P=3.9x10 -4 ). Subgroup analyses demonstrated consistent effects across stroke type, sex, age, country income level, and education level. In addition, mtDNA-CN significantly improved reclassification of poor functional outcome status (Net Reclassification Index (NRI)=0.16; 95% CI, 0.08-0.23; P=3.6x10 -5 ) and mortality (NRI=0.31; 95% CI, 0.19-0.43; P=1.7x10 -7 ) beyond known prognosticators. Using independent datasets, Mendelian Randomization revealed that a 1 SD decrease in genetically determined mtDNA-CN was associated with increased odds of greater 3-month disability quantified by ordinal mRS (OR=2.35; 95% CI, 1.13-4.90; P=0.02) and poor functional outcome status (OR=2.68; 95% CI, 1.05-6.86; P=0.04). Conclusions: Buffy coat mtDNA-CN is a novel and robust marker of post-stroke prognosis that may also be a causal determinant of post-stroke outcomes. Classification of Evidence: This study provides class II evidence that low buffy coat mtDNA-CN (>1-standard deviation) was associated with worse baseline severity and 1-month outcomes in patients with ischemic or hemorrhagic stroke.