申請者はこれまで、DNAメチル化やヒストン修飾などのエピゲノム変化の腎臓病病態への影響に注目して検討し報告してきた（JCI 2014, Kidney Int 2015）。また最近ではDNA損傷修復がエピゲノム変化形成の原因の一つであることを、糖尿病性腎症モデルを用いて明らかにした（Cell Rep 2019）。本研究では、最近報告されているエピゲノムワイド解析において、腎機能と血球細胞のDNAメチル化が関連していることに注目し、腎臓のDNA損傷・DNAメチル化変化と血球細胞のDNAメチル化の関連性について明らかにすることを目的とした。最近、老化研究領域で注目されているEpigenetic clock (EC)は機械学習で特定された重要領域のDNAメチル化レベルから計算されるDNAメチル化年齢であり、複数の疾患で促進が報告されているがCKDとの関連は不明であることから、まずECと腎臓のDNA損傷との関連を検討した。当院予防医療センター受診者26名（男性16名、女性10名、年齢62±10、eGFR 63±9）を対象に採血検体を用いてEPICアレイで85万ヶ所のCpGを解析し、既法のECであるWeidner's clockを算出した。その結果、ECは暦年齢と有意に相関（r=0.39, p=0.0143）し、CKD群（eGFR 45-59）ではEC-暦年齢が有意に増加し、生物学的な加齢との関連が示唆された。また、long-distance PCR法により尿中脱落細胞のDNA二本鎖切断（DSB）を評価すると、CKD群ではポドサイトDSBが増加しており、特男性では暦年齢よりもEC-暦年齢で表される生物学的な加齢と相関する傾向にあった。以上の結果、早期CKDと生物学的加齢との関連が示唆された。また、尿中脱落細胞のDSBが生物学的加齢を検出するマーカーとなりうる可能性があると考えられた。今後腎機能低下症例に関して更に解析を予定している。
Prevalence of chronic kidney disease (CKD) is now increasing over the world due to the increase in the population of diabetes and hypertension patients, and in Japan, one in eight adults has CKD. There is not an effective therapy for CKD regression, therefore the establishment of novel strategies for conquering CKD is an urgent issue.
We have investigated the 'memory effect' in lifestyle-related diseases, which is a sustained effect of a transient treatment or insult, focusing the memory contained in the kidney. Recently we have demonstrated that epigenetic alterations in glomerular podocytes is involved in the memory effect in CKD (J Clin Invest 2014, Kidney Int 2015). In addition, the epigenetic alterations are associated with DNA damage repair, which is related to the pathogenesis of diabetic nephropathy (Cell Rep 2019). In this study, we aimed to clarify the relationship between kidney DNA damage and altered DNA methylation in blood cells, as the recently reported epigenome-wide analysis suggests the relationship between renal function and DNA methylation in blood cells. In addition, epigenetic clock (EC) has recently attracted attention in the field of aging research, which is a DNA methylation age calculated from DNA methylation levels in critical CpGs identified by machine learning. Its promotion has been reported in several diseases, but the association with CKD is unknown. Therefore, we first investigated the relationship between the EC and kidney DNA damage. We analyzed 850,000 CpG sites by EPIC array using blood samples from 26 patients (16 males and 10 females, age 62±10, eGFR 63±9) who visited the center for preventive medicine of Keio University Hospital, and calculated ECs. EC (Weider's clock) was significantly correlated with chronological age (r=0.39, p=0.0143), and EC was significantly increased in the CKD group (eGFR 45-59), suggesting an association with biological aging. Furthermore, evaluation of DNA double-strand breaks (DSBs) in urine derived cells by long-distance PCR showed increased podocyte DSBs in the CKD group, which tended to correlate with biological aging rather than chronological age, especially in men. These results suggest an association between CKD even in early stage and biological aging. We consider that DSBs of urine derived cells may be a potential marker for detecting biological aging. Further analysis is planned for patients with advanced CKD.