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Background: Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a typical neurodegenerative disease associated with mitochondrial dysfunction. Methylation of the D-loop region and mitochondrial DNA copy number (mtDNAcn) play a critical role in the maintenance of mitochondrial function. However, the association between D-loop region methylation, mtDNAcn and CADASIL remains unclear. Methods: Overall, 162 individuals were recruited, including 66 CADASIL patients and 96 age- and sex-matched controls. After extracting genomic DNA from the peripheral white blood cells, levels of D-loop methylation and mtDNAcn were assessed using MethylTarget sequencing and real-time PCR, respectively. Results: We observed increased mtDNAcn and decreased D-loop methylation levels in CADASIL patients compared to the control group, regardless of gender stratification. Besides, we found a negative correlation between D-loop methylation levels and mtDNAcn. Mediation effect analysis shows that the proportion of the association between mtDNAcn and CADASIL that is mediated by D-loop methylation is 11.6% (95% CI 5.6, 22.6). After gender stratification, the proportions of such associations that are mediated by D-loop methylation in males and females were 7.2% (95% CI 2.4, 19.8) and 22.0% (95% CI 7.4, 50.1), respectively. Conclusion: Decreased methylation of the D-loop region mediates increased mtDNAcn in CADASIL, which may be caused by a compensatory mechanism of mitochondrial dysfunction in patients with CADASIL. [ABSTRACT FROM AUTHOR]

Background: Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) caused by NOTCH3 mutations is the most common monogenic hereditary pattern of cerebral small vessel disease. The aggregation of the mutant NOTCH3 may play a cytotoxic role in CADASIL. However, the main mechanism of this process remains unclear.Objective: We aimed to investigate the possible pathogenesis of the mutant NOTCH3 in CADASIL.Methods: The clinical information of two pedigrees were collected and analyzed. Furthermore, we constructed cell lines corresponding to this mutation in vitro. The degradation of the extracellular domain of NOTCH3 (NOTCH3ECD) was analyzed by Cycloheximide Pulse-Chase Experiment. Flow cytometry and cell counting kit-8 assay were performed to observe the effects of the NOTCH3 mutation on mitochondrial function and apoptosis.Results: We confirmed a de novo heterozygous missense NOTCH3 mutation (c.1690G > A, p. A564T) in two pedigrees. In vitro, the NOTCH3ECD aggregation of A564T mutant may be related to their more difficult to degrade. The mitochondrial membrane potential was attenuated, and cell viability was significant decreased in NOTCH3ECD A564T group. Interestingly, BAX and cytochrome c were significantly increased, which are closely related to the mitochondrial-mediated pathway to apoptosis.Conclusion: In our study, the aggregation of NOTCH3ECD A564T mutation may be associated with more difficult degradation of the mutant, and the aggregation may produce toxic effects to induce apoptosis through the mitochondrial-mediated pathway. Therefore, we speculated that mitochondrial dysfunction may hopefully become a new breakthrough point to explain the pathogenesis of cysteine-sparing NOTCH3 mutations. [ABSTRACT FROM AUTHOR]

Background: Exosomes are nano-sized extracellular vesicles which are secreted by cells and usually found in body fluids. Previous research has shown that exosomal secretion and autophagy-lysosomal pathway synergistically participates in intracellular abnormal protein elimination. The main pathological manifestations of Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is abnormal accumulation of mutant NOTCH3, and CADASIL vascular smooth muscle cells have been found with autophagy-lysosomal dysfunction. However, whether plasma exosomes change in CADASIL patients is still unclear.Objective: We are aimed to investigate the differences of plasma exosomes between CADASIL patients and healthy controls.Methods: The subjects included 30 CADASIL patients and 30 healthy controls without NOTCH3 mutation. The severity of white matter lesions (WMLs) of CADASIL patients was quantified by Fazekas score. Transmission electron microscopy and nanoparticle tracking analysis were performed to characterize plasma exosomes. In addition, NOTCH3, Neurofilament light and Aβ42 levels in plasma exosomes were quantified by enzyme-linked immunosorbent assays.Results: We found that exosomes from CADASIL patients were lower in quantity. In addition, CADASIL plasma exosomes had significantly lower levels of NOTCH3 and significantly increased levels of NFL than those of matched healthy subjects. Interestingly, plasma exosome NOTCH3 levels of CADASIL patients significantly correlated with severity of WMLs.Conclusion: The exosome NOTCH3 may be related to the pathological changes of CADASIL, which provides a basis for the pathogenesis research of CADASIL. In addition, plasma exosome NOTCH3 and NFL levels may act as biomarkers to monitor and predict disease progression and measure therapeutic effectiveness in the future clinical trials. [ABSTRACT FROM AUTHOR]