外周血线粒体DNA D环区甲基化及拷贝数与帕金森病相关性研究

袁瑞; 赵艳青; 张中奇; 王丽珍; 杨志甫

doi:10.12435/j.issn.2095-5227.2023.069

外周血线粒体DNA D环区甲基化及拷贝数与帕金森病相关性研究

Correlation between methylation and copy number of mitochondrial DNA D-loop region in peripheral blood and Parkinson's disease

摘要

摘要:
背景帕金森病(Parkinson's disease，PD)是一种多发于中老年的神经退行性疾病，线粒体DNA(mtDNA)甲基化可能参与PD的发生和发展，目前相关研究极少。
目的观察PD患者外周血mtDNA D环区的甲基化水平及其拷贝数的变化，分析D环区甲基化与mtDNA拷贝数的相关性，探讨PD的发病机制。
方法选取2020 - 2022年内蒙古科技大学包头医学院第一附属医院PD患者(PD组)和健康体检者(对照组)各30例(66.63 ± 7.95)岁 vs (63.50 ± 4.96)岁，每组男性、女性各15例，采用质谱法(MassARRAY)检测外周血mtDNA D环区的甲基化水平，采用实时定量PCR法(qRT-PCR)检测mtDNA拷贝数。
结果 PD组与对照组的外周血mtDNA中D环区总体甲基化率及各CpG位点甲基化率差异无统计学意义(P＞0.05)，早期PD患者与中晚期PD患者的外周血mtDNA中D环区总体甲基化率及各CpG位点甲基化率差异无统计学意义(P＞0.05)。PD患者外周血mtDNA拷贝数低于对照组(P=0.02)，中晚期PD患者外周血mtDNA拷贝数低于早期PD患者，差异有统计学意义(P=0.012)。mtDNA D环区第1、15个CpG位点(即人mtDNA序列第112、413位的核苷酸)甲基化率与mtDNA拷贝数呈负相关(r=-0.257，P=0.047；r=-0.517，P＜0.001)。Logistic回归分析显示，mtDNA拷贝数越高，PD发生概率越低(OR=0.119，P=0.023)。
结论 PD患者外周血mtDNA D环区甲基化水平未发生显著改变，但其外周血mtDNA拷贝数降低，且随着PD病程的进展呈动态变化，mtDNA拷贝数有望成为预测PD发生及病程进展的指标。D环区的甲基化可能会负向调控mtDNA的复制与转录。

Abstract:
Background Parkinson's disease (PD) is a neurodegenerative disease that occurs mostly in middle-aged and elderly people. Mitochondrial DNA (mtDNA) methylation may be involved in the pathogenesis and progression of PD, but there are few related studies.
Objective To observe the changes in methylation level of the D-loop region and the mtDNA copy number in peripheral blood of PD patients, analyze the correlation between them, so as to explore the potential involvement of mtDNA methylation in the pathogenesis of PD.
Methods Totally 30 PD patients and 30 age and sex-matched normal controls (66.63 ± 7.95 years vs 63.50 ± 4.96 years) were selected from the First Affiliated Hospital of Baotou Medical College from 2020 to 2022, including 15 males and 15 females in this case-control study. The D-loop methylation analyses in peripheral blood were performed using mass spectrometry (MassARRAY), and the mtDNA copy number was detected by quantitative real-time PCR analysis (qRT-PCR).
Results There was no significant difference in the overall methylation level and the methylation level of each CpG site of the D-loop region in peripheral blood between PD group and control group, and no significant difference was found between early and advanced PD patients either (P>0.05). The mtDNA copy number in peripheral blood of PD group was lower than that of controls (P=0.02), and the mtDNA copy number of advanced PD patients was lower than that of early PD patients (P=0.012). The methylation levels of CpG sites 1, 15 in the D-loop region were negatively correlated with the mtDNA copy number (r=-0.257, P=0.047; r=-0.517, P＜0.001). Binary Logistic regression analysis showed that more mtDNA copy number was associated with lower likelyhood of PD (OR=0.119, P=0.023).
Conclusion The methylation level of D-loop region in peripheral blood of PD patients does not change significantly, but the mtDNA copy number in peripheral blood of PD patients decreases dynamically with the progression of PD, which will be an index to predict the pathogenesis and progression of PD. The methylation of the D-loop region may negatively regulate mtDNA replication and transcription.

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