Department of Neurosurgery, Robert Wood Johnson Medical School, Rutgers University, Piscataway, NJ, 08854, USA.
Department of Psychology and Neuroscience Program, Trinity College, Hartford, CT, 06106, USA.
Sci Rep. 2022 Jan 10;12(1):380. doi: 10.1038/s41598-021-04346-w.
Epigenetic modifications are crucial for normal development and implicated in disease pathogenesis. While epigenetics continues to be a burgeoning research area in neuroscience, unaddressed issues related to data reproducibility across laboratories remain. Separating meaningful experimental changes from background variability is a challenge in epigenomic studies. Here we show that seemingly minor experimental variations, even under normal baseline conditions, can have a significant impact on epigenome outcome measures and data interpretation. We examined genome-wide DNA methylation and gene expression profiles of hippocampal tissues from wild-type rats housed in three independent laboratories using nearly identical conditions. Reduced-representation bisulfite sequencing and RNA-seq respectively identified 3852 differentially methylated and 1075 differentially expressed genes between laboratories, even in the absence of experimental intervention. Difficult-to-match factors such as animal vendors and a subset of husbandry and tissue extraction procedures produced quantifiable variations between wild-type animals across the three laboratories. Our study demonstrates that seemingly minor experimental variations, even under normal baseline conditions, can have a significant impact on epigenome outcome measures and data interpretation. This is particularly meaningful for neurological studies in animal models, in which baseline parameters between experimental groups are difficult to control. To enhance scientific rigor, we conclude that strict adherence to protocols is necessary for the execution and interpretation of epigenetic studies and that protocol-sensitive epigenetic changes, amongst naive animals, may confound experimental results.
表观遗传修饰对于正常发育至关重要，并与疾病发病机制有关。尽管表观遗传学在神经科学中仍是一个迅速发展的研究领域，但实验室间数据可重复性相关的未解决问题仍然存在。将有意义的实验变化与背景变异性区分开来是表观基因组研究中的一个挑战。在这里，我们展示了即使在正常基线条件下，看似微小的实验变化也会对表观基因组的测量结果和数据解释产生重大影响。我们使用几乎相同的条件，检查了来自三个独立实验室的野生型大鼠海马组织的全基因组 DNA 甲基化和基因表达谱。降低代表性的亚硫酸氢盐测序和 RNA-seq 分别鉴定出实验室之间存在 3852 个差异甲基化和 1075 个差异表达的基因，即使在没有实验干预的情况下也是如此。难以匹配的因素，如动物供应商以及部分饲养和组织提取程序，在三个实验室中的野生型动物之间产生了可量化的变化。我们的研究表明，即使在正常基线条件下，看似微小的实验变化也会对表观基因组的测量结果和数据解释产生重大影响。这对于动物模型中的神经科学研究尤其有意义，因为实验组之间的基线参数很难控制。为了提高科学严谨性，我们得出结论，严格遵守方案对于执行和解释表观遗传学研究是必要的，并且在天真动物中，对方案敏感的表观遗传变化可能会使实验结果复杂化。
