並列タイトル等バイサルファイト変換とARMS PCRを用いた膵β細胞傷害の新規定量法
Detecting pancreatic β-cell cfDNA
タイトル(掲載誌)Journal of Diabetes Investigation
一般注記Aims/Introduction: Several research groups have reported methods for quantifying pancreatic beta cell (β-cell) injury by measuring β-cell-specific CpG unmethylation of the insulin gene in circulation using digital droplet PCR or next-generation sequencing. However, these methods have certain disadvantages, such as the need to consider the background signal owing to the small number of target CpG sites and the need for unique equipment.
Materials and Methods: We established a novel method for detecting four CpG unmethylations of the insulin gene using two-step amplification refractory mutation system PCR. We applied it to type 1 diabetes (T1D) patients with a wide range of disease durations and to healthy adults.
Results: The assay showed high linearity and could detect a single copy of unmethylated insulin DNA in experiments using methylated and unmethylated plasmid DNA. The unmethylated insulin DNA level in the type 1 diabetes group, whose β-cell mass was considerably reduced, was similar to that of healthy adults. An inverse correlation was observed between copy number and disease duration in patients with unmethylated insulin DNA-positive type 1 diabetes.
Conclusions: We developed a novel method for detecting unmethylated insulin DNA in circulation that can be performed using a conventional real-time PCR system. This method would be useful for analyzing dynamic profiles of β-cells in human disease such as type 1 diabetes.
著作権情報This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License ( https://creativecommons.org/licenses/by-nc/4.0/ ), which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.
連携機関・データベース国立情報学研究所 : 学術機関リポジトリデータベース(IRDB)(機関リポジトリ)