The study cohort comprised 42 stillbirths (SBs) with brainstem lesions, including 32 cases that also displayed cardiac conduction system (CCS) defects, along with 32 control fetuses lacking these anomalies. DNA extracted from brainstem tissues was analyzed in both groups. In addition, unaffected tissues from the SB cases were examined for intra-individual comparison. Subgroup stratification was performed according to sex, severity of brainstem lesions, and the presence of concomitant CCS defects. NGS-based mtDNA sequencing revealed an increased mutational burden in cases without CCS anomalies. MtDNA-CN was significantly higher in affected brainstem tissues compared with both control brainstem tissues ( p <0.0001) and unaffected tissues from the same cases ( p =0.005), particularly in SBs with milder brainstem lesions. Displacement loop (D-loop) instability was observed in 37% of cases, with significantly increased D-loop methylation in affected brainstem tissues compared with both control brainstem tissues ( p <0.0001) and unaffected tissues ( p =0.0001). • Mt-DNA was analysed in the affected brainstem tissues from 42 stillbirths. • A higher mtDNA copy number was observed in cases compared with controls. • D-loop methylation levels were higher in affected brainstem tissues compared with controls. • Mitochondrial dysfunction is a key contributor to fetal demise, and mtDNA-CN is a potential biomarker for SB. Stillbirth (SB) accounts for over 60% of perinatal deaths in high-income countries, with a significant portion of cases remaining unexplained following thorough anatomopathological investigation. Mitochondrial DNA (mtDNA) alterations were analyzed in 42 SB cases with brainstem and cardiac conduction system (CCS) anomalies and in 32 control fetuses without these anomalies. DNA extracted from brainstem tissues was analyzed in both groups. In addition, unaffected tissues from the SB cases were examined for intra-individual comparison. The analysis included mtDNA sequencing, haplogroup determination, copy number (CN) quantification, and evaluation of displacement loop (D-loop) instability and methylation. Across the entire SB cohort, a total of 158 variants were identified, with a significant enrichment of variants observed in cases without CCS anomalies ( p = 0.024). Affected brainstem tissues exhibited significantly higher mtDNA-CN compared with both control brainstem tissues ( p < 0.0001) and unaffected tissues ( p = 0.005), with levels higher in mild lesions than in severe lesions ( p = 0.02). D-loop instability was identified in 37% of cases, and D-loop methylation levels were consistently higher in affected brainstem tissues compared with both control brainstem tissues ( p < 0.0001) and unaffected tissues ( p = 0.0001). These findings support mitochondrial dysfunction as a key contributor to fetal demise and mtDNA-CN as a potential biomarker for SB.
Lecca et al. (Wed,) studied this question.