Department of Human Biology, Zoological Institute, Christian-Albrechts-University, Am Botanischen Garten 9, 24118 Kiel, Germany.
Exp Gerontol. 2021 Aug;151:111422. doi: 10.1016/j.exger.2021.111422. Epub 2021 May 24.
BMAL1 is a core clock gene that positively regulates circadian rhythms. In animals, BMAL1 further acts as transcription factor for the SOD1 gene which encodes the major antioxidant enzyme superoxide dismutase. SOD1 protects against oxidative damage that is a major factor for human aging. Mice lacking mBmal1 exhibit premature aging phenotypes which might be related to a reduced expression of SOD1. The purpose of this study was to explore the circadian relationship between BMAL1, SOD1, oxidative RNA/DNA damage and aging in healthy humans.
In a cross-sectional study design, buccal mucosa cells and saliva samples were obtained from 21 young (23.7 ± 2.3 yrs) and 21 older women (66.8 ± 5.7 yrs) within 24 h at 4-h intervals (08:00, 12:00, 16:00, 20:00, 24:00 and 04:00 h). Transcript levels of BMAL1 and SOD1 were measured by real-time quantitative PCR. Protein levels of SOD1 were determined by immunoblotting and densitometry. Levels of oxidative RNA/DNA damage and melatonin were quantified by enzyme immunosorbent assays.
Transcript levels of BMAL1 and SOD1 mRNAs as well as protein levels of SOD1 and melatonin exhibited significant 24-h variation in each age group (P < 0.010, Friedman tests). The mRNA expression patterns of BMAL1 and SOD1 showed similar 24-h rhythmicity and positive relationships were found. Strongest relationships occurred in young women at 12:00 h (rs = 0.81, P = 0.005) and in older women at 08:00 h (r = 0.84, P < 0.001). Maximum levels of SOD1 mRNA appeared within 24 h in both age groups at 24:00 h. In both age groups, the timing of maximum level for SOD1 protein was delayed relative to the timing of maximum level for SOD1 mRNA. This delay was larger in older women (8 h) compared to young women (4 h). Older women showed higher oxidative RNA/DNA damage at all time-points compared to young women (P < 0.020). The oxidative RNA/DNA damage decreased continuously from 08:00 to 20:00 h (P < 0.001) in both age groups. Although oxidative damage and SOD1 protein levels declined simultaneously, only weak and non-significant relationships were noted.
This study demonstrates that transcript levels of SOD1 and protein level of SOD1 follow a circadian pattern of expression in healthy young and older women. The time-shift found between the 24-h maximum levels of SOD1 mRNA and SOD1 protein could be explained by the time needed for translation of SOD1 protein. The positive relationship found between expression levels of SOD1 mRNA and BMAL1 mRNA is in line with animal studies showing that BMAL1 acts as transcription factor and regulates the circadian synthesis of SOD1 mRNA. Differences between young and older women found in the timing of 24-h maxima as well as differences observed in 24-h relationships between characters might be due to age-related alterations in the circadian system. Absence of relationships between levels of SOD1 and oxidative RNA/DNA damage argue for a more complex interaction between the antioxidant system and the circadian system.
BMAL1 是一种核心时钟基因，它正向调节昼夜节律。在动物中，BMAL1 进一步作为 SOD1 基因的转录因子发挥作用，该基因编码主要的抗氧化酶超氧化物歧化酶。SOD1 可防止氧化损伤，氧化损伤是人类衰老的一个主要因素。缺乏 mBmal1 的小鼠表现出衰老表型提前，这可能与 SOD1 表达降低有关。本研究旨在探索健康人体中 BMAL1、SOD1、氧化 RNA/DNA 损伤与衰老之间的昼夜节律关系。
在一项横断面研究设计中，在 24 小时内每隔 4 小时（08:00、12:00、16:00、20:00、24:00 和 04:00 h）从 21 名年轻（23.7±2.3 岁）和 21 名老年女性（66.8±5.7 岁）的口腔颊黏膜细胞和唾液样本中获取。通过实时定量 PCR 测量 BMAL1 和 SOD1 的转录水平。通过免疫印迹和密度测定法测定 SOD1 蛋白水平。通过酶联免疫吸附试验定量测定氧化 RNA/DNA 损伤和褪黑素水平。
在每个年龄组中，BMAL1 和 SOD1 mRNA 的转录水平以及 SOD1 和褪黑素的蛋白水平均表现出显著的 24 小时变化（P<0.010，Friedman 检验）。BMAL1 和 SOD1 的 mRNA 表达模式表现出相似的 24 小时节律性，并且发现它们之间存在正相关关系。在年轻女性中，最强的相关性出现在 12:00 h（rs=0.81，P=0.005），在老年女性中，最强的相关性出现在 08:00 h（r=0.84，P<0.001）。在两个年龄组中，SOD1 mRNA 的最高水平出现在 24 小时内的 24:00 h。在两个年龄组中，SOD1 蛋白的最高水平出现在 SOD1 mRNA 最高水平之后。老年女性的延迟时间（8 h）比年轻女性（4 h）更长。与年轻女性相比，老年女性在所有时间点的氧化 RNA/DNA 损伤均更高（P<0.020）。在两个年龄组中，氧化 RNA/DNA 损伤从 08:00 到 20:00 连续下降（P<0.001）。尽管氧化损伤和 SOD1 蛋白水平同时下降，但只注意到微弱且无统计学意义的关系。
本研究表明，在健康的年轻和老年女性中，SOD1 的转录水平和 SOD1 蛋白水平呈现出昼夜节律表达模式。在 SOD1 mRNA 和 SOD1 蛋白的 24 小时最大值之间发现的时间移位可以用 SOD1 蛋白翻译所需的时间来解释。发现 SOD1 mRNA 和 BMAL1 mRNA 之间表达水平的正相关与表明 BMAL1 作为转录因子并调节 SOD1 mRNA 的昼夜合成的动物研究一致。在 24 小时最大值的时间和在 24 小时内观察到的特征之间的关系方面，年轻女性和老年女性之间存在差异，这可能是由于与年龄相关的昼夜节律系统改变所致。SOD1 水平与氧化 RNA/DNA 损伤之间缺乏关系表明抗氧化系统和昼夜节律系统之间存在更复杂的相互作用。
