Aquaporin5 Deficiency Aggravates ROS/NLRP3 Inflammasome-Mediated Pyroptosis in the Lacrimal Glands.
Aquaporin5 Deficiency Aggravates ROS/NLRP3 Inflammasome-Mediated Pyroptosis in the Lacrimal Glands.
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DOI：
10.1167/iovs.64.1.4
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发表时间：
2023-01-03
影响因子：
4.4
通讯作者：
中科院分区：
文献类型：
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作者：
在干眼症中，泪腺(LGs)的发病机制是由Nacht、LRR和NLRP3炎性小体介导的炎症促进的。本研究旨在探讨水通道蛋白5(Aquaporin 5，AQP5)通过抑制活性氧(ROS)和NLRP3炎症小体对LGs的保护作用。采用AQP5基因敲除小鼠(AQP5−/−)观察LGS的病理变化。用二氯二氢荧光素二乙酸酯比色法检测ROS的产生。油红O染色检测脂质代谢。用JC-1荧光探针试剂盒检测线粒体膜电位的逆转。用N-乙酰L半胱氨酸或MCC950进行药物干预，观察AQP5对NLRP3/Caspase-1/Gasdermin-D(GSDMD)介导的下垂的影响。AQP5缺失显著增加了AQP5−/−LGs的ROS生成、脂代谢紊乱、TUNEL阳性细胞数和线粒体膜电位的反转。在AQP5−/−小鼠LGs和原代LG上皮细胞中，NLRP3表达上调，caspase-1和GSDMD活性增加，IL-1 NLRP3释放增加。MCC950显著抑制AQP5缺乏诱导的LGs和原代LG上皮细胞NLRP3炎症性下垂。此外，我们还发现，用N-乙酰L半胱氨酸预先刺激AQP5−/−原代LG上皮细胞后，NLRP3的表达、Caspase-1和GSDMD活性水平以及IL-1β的释放均降低。我们的结果表明，AQP5的丢失通过产生ROS促进了NLRP3炎症体的激活。抑制ROS或NLRP3炎性小体显著减轻AQP5缺陷LG上皮细胞的损伤和下垂，这可能为干眼病提供新的见解。
The pathogenesis of the lacrimal glands (LGs) is facilitated by inflammation mediated by the NACHT, LRR, and NLRP3 inflammasomes in dry eye disease. This research aimed to explore the protective effects of Aquaporin 5 (AQP5) on LGs by inhibiting reactive oxygen species (ROS) and the NLRP3 inflammasome. AQP5 knockout (AQP5−/−) mice were used to evaluate pathological changes in LGs. ROS generation was detected with a dichlorodihydro–fluorescein diacetate assay. Lipid metabolism was assessed by Oil Red O staining. The reversal of the mitochondrial membrane potential was detected using a JC-1 fluorescent probe kit. The effect of AQP5 on NLRP3/caspase-1/Gasdermin-D (GSDMD)-mediated pyroptosis was examined using pharmacological treatment of N-acetyl L-cysteine or MCC950. AQP5 loss significantly increased ROS generation, lipid metabolism disorders, TUNEL-positive cells, and reversal of the mitochondrial membrane potential in the AQP5−/− LGs. NLRP3 upregulation, increased caspase-1 and GSDMD activity, and enhanced IL-1β release were detected in the AQP5−/− mouse LGs and primary LG epithelial cells. MCC950 significantly suppressed NLRP3 inflammasome-related pyroptosis induced by AQP5 deficiency in LGs and primary LG epithelial cells. Furthermore, we discovered that prestimulating the AQP5−/− primary LG epithelial cells with N-acetyl L-cysteine decreased NLRP3 expression, caspase-1 and GSDMD activity levels, and IL-1β release. Our results revealed that AQP5 loss promoted NLRP3 inflammasome activation through ROS generation. Inhibiting the ROS or NLRP3 inflammasome significantly alleviated the damage and pyroptosis of AQP5-deficient LG epithelial cells, which could provide new insights into dry eye disease.
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