非眼组织中的证据表明,抗氧化剂谷胱甘肽(GSH)可以以昼夜节律方式被调节,导致晶状体中的GSH水平也可以以昼夜节律方式被控制以预测氧化应激的时期的想法。
■在24小时内每4小时收集雄性大鼠Wistar晶状体(6周),10am,2pm,6pm,晚上10点和凌晨2点和定量PCR,进行蛋白质印迹和免疫组织化学以检查核心时钟基因和蛋白质的表达(BMAL1,CLOCK,CRY1-2,PER1-3)及其在24小时内的亚细胞定位。还进行了晶状体的Western印迹以检查NRF2的表达,NRF2是一种参与调节GSH稳态和GSH相关酶的基因的转录因子(GCLC,GS和GR)在24小时内。最后,HLPC用于在24小时内每4小时测量房水和晶状体中的GSH水平。
■大鼠晶状体含有生物钟的核心分子成分,具有核心时钟蛋白的表达,NRF2和GSH相关酶在24小时内波动。BMAL1表达在白天最高,上午10点BMAL1定位于细胞核。NRF2表达在24小时内保持恒定,尽管似乎每4小时进出原子核。GSH相关的酶表达趋于在夜间开始时达到峰值,这与晶状体中GSH的高水平和房水中GSH的低水平有关。
■镜头包含昼夜节律钟的关键部件,GSH和参与维持GSH稳态的GSH相关酶的表达存在时间差异。大鼠晶状体中的GSH水平在夜间开始时最高,这代表了大鼠的活动阶段,此时可能需要高GSH水平来抵消细胞代谢引起的氧化应激。将时钟与晶状体中GSH水平的调节直接联系起来的未来工作对于确定时钟是否可用于帮助恢复晶状体中的GSH水平将是重要的。
UNASSIGNED: Evidence in non-ocular tissues indicate that the antioxidant glutathione (GSH) may be regulated in a circadian manner leading to the idea that GSH levels in the
lens may also be controlled in a circadian manner to anticipate periods of oxidative stress.
UNASSIGNED: Male rat Wistar lenses (6 weeks) were collected every 4 hours over a 24-hour period at 6am, 10am, 2pm, 6pm, 10pm and 2am and quantitative-PCR, western blotting and immunohistochemistry performed to examine the expression of core clock genes and proteins (BMAL1, CLOCK, CRY1-2, PER 1-3) and their subcellular localisation over a 24-hour period. Western blotting of lenses was also performed to examine the expression of NRF2, a transcription factor involved in regulating genes involved in GSH homeostasis and GSH related enzymes (GCLC, GS and GR) over the 24-hour period. Finally, HLPC was used to measure GSH levels in the aqueous humour and lenses every 4 hours over a 24-hour period.
UNASSIGNED: The rat
lens contains the core molecular components of a circadian clock with the expression of core clock proteins, NRF2 and GSH related enzymes fluctuating over a 24-hour period. BMAL1 expression was highest during the day, with BMAL1 localised to the nuclei at 10am. NRF2 expression remained constant over the 24-hour period, although appeared to move in and out of the nuclei every 4 hours. GSH related enzyme expression tended to peak at the start of night which correlated with high levels of GSH in the
lens and lower levels of GSH in the aqueous humour.
UNASSIGNED: The lens contains the key components of a circadian clock, and time-of-day differences exist in the expression of GSH and GSH related enzymes involved in maintaining GSH homeostasis. GSH levels in the rat
lens were highest at the start of night which represents the active phase of the rat when high GSH levels may be required to counteract oxidative stress induced by cellular metabolism. Future work to directly link the clock to regulation of GSH levels in the lens will be important in determining whether the clock can be used to help restore GSH levels in the
lens.