Degradation of methylation signals in cryopreserved DNA.

IF 5.7 2区 医学 Q1 Medicine Clinical Epigenetics Pub Date : 2023-09-11 DOI:10.1186/s13148-023-01565-y
Ning Yuan Lee, Melissa Hum, Guek Peng Tan, Ai Choo Seah, Patricia T Kin, Ngiap Chuan Tan, Hai-Yang Law, Ann S G Lee
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Abstract

Background: Blood-based DNA methylation has shown great promise as a biomarker in a wide variety of diseases. Studies of DNA methylation in blood often utilize samples which have been cryopreserved for years or even decades. Therefore, changes in DNA methylation associated with long-term cryopreservation can introduce biases or otherwise mislead methylation analyses of cryopreserved DNA. However, previous studies have presented conflicting results with studies reporting hypomethylation, no effect, or even hypermethylation of DNA following long-term cryopreservation. These studies may have been limited by insufficient sample sizes, or by their profiling of methylation only on an aggregate global scale, or profiling of only a few CpGs.

Results: We analyzed two large prospective cohorts: a discovery (n = 126) and a validation (n = 136) cohort, where DNA was cryopreserved for up to four years. In both cohorts there was no detectable change in mean global methylation across increasing storage durations as DNA. However, when analysis was performed on the level of individual CpG methylation both cohorts exhibited a greater number of hypomethylated than hypermethylated CpGs at q-value < 0.05 (4049 hypomethylated but only 50 hypermethylated CpGs in discovery, and 63 hypomethylated but only 6 hypermethylated CpGs in validation). The results were the same even after controlling for age, storage duration as buffy coat prior to DNA extraction, and estimated cell type composition. Furthermore, we find that in both cohorts, CpGs have a greater likelihood to be hypomethylated the closer they are to a CpG island; except for CpGs at the CpG islands themselves which are less likely to be hypomethylated.

Conclusion: Cryopreservation of DNA after a few years results in a detectable bias toward hypomethylation at the level of individual CpG methylation, though when analyzed in aggregate there is no detectable change in mean global methylation. Studies profiling methylation in cryopreserved DNA should be mindful of this hypomethylation bias, and more attention should be directed at developing more stable methods of DNA cryopreservation for biomedical research or clinical use.

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冷冻保存DNA中甲基化信号的降解。
背景:基于血液的DNA甲基化作为一种生物标志物在多种疾病中显示出巨大的前景。血液中DNA甲基化的研究通常使用冷冻保存数年甚至数十年的样本。因此,与长期冷冻保存相关的DNA甲基化的变化可能会引入偏见或以其他方式误导冷冻保存DNA的甲基化分析。然而,先前的研究结果与报告长期冷冻保存后DNA低甲基化、无作用甚至高甲基化的研究结果相矛盾。这些研究可能受到样本量不足的限制,或者仅在总体范围内对甲基化进行分析,或者仅对少数CpG进行分析。结果:我们分析了两个大型前瞻性队列: = 126)和验证(n = 136)队列,其中DNA被冷冻保存长达四年。在这两个队列中,随着DNA储存时间的增加,平均全局甲基化没有可检测的变化。然而,当对单个CpG甲基化水平进行分析时,在q值下,两个队列都表现出比高甲基化CpG更多的低甲基化CpGs 结论:几年后DNA的冷冻保存导致在单个CpG甲基化水平上对低甲基化的可检测的偏向,尽管在总体分析时,平均全局甲基化没有可检测的变化。分析冷冻保存DNA中甲基化的研究应注意这种低甲基化倾向,并应更多地关注开发更稳定的DNA冷冻保存方法,用于生物医学研究或临床应用。
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来源期刊
Clinical Epigenetics
Clinical Epigenetics Biochemistry, Genetics and Molecular Biology-Developmental Biology
CiteScore
8.90
自引率
5.30%
发文量
150
审稿时长
12 weeks
期刊介绍: Clinical Epigenetics, the official journal of the Clinical Epigenetics Society, is an open access, peer-reviewed journal that encompasses all aspects of epigenetic principles and mechanisms in relation to human disease, diagnosis and therapy. Clinical trials and research in disease model organisms are particularly welcome.
期刊最新文献
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