cfDNA is increased in patients with SCD. (A) Quantification of cf-nDNA in the cross-sectional cohort (upper panel) and paired samples (lower panel) by real-time qPCR assays of human genomic targets GAPDH and TERT. Upper panels: HCs, n = 13; SCD-Baseline, n = 72, and SCD-Crisis, n = 20. Data are sample mean ± SD. *P < .05, nonparametric Kruskal-Wallis test with Dunn’s multiple-comparison test. Lower panels: SCD-Baseline and SCD-Crisis pairs, n = 18. Circles represent the mean of each sample. *P < .05, **P < .01, nonparametric Wilcoxon matched-pairs signed-rank test. (B) qPCR assays were performed for human mitochondrial targets (MT-ND1 and MT-ND6), as well as for nuclear targets (GAPDH and TERT), and the cf-mtDNA/cf-nDNA ratio was calculated by normalizing the cycle threshold (ct) values with the mean ct value of GAPDH from HCs. Quantitation of cf-mtDNA/cf-nDNA ratio using the following mitochondrial and nuclear targets: MT-ND1/GAPDH, MT-ND1/TERT, MT-ND6/GAPDH, and MT-ND6/TERT. Error bars represent the sample mean ± SD. *P < .05, ***P < .0005, ****P < .0001, nonparametric Kruskal-Wallis test with Dunn’s multiple comparison test.