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2-DG通过抑制NLRP3介导的细胞焦亡改善大鼠肺缺血/再灌注损伤

石璐¹, 黄曼², 陈思安², 徐俊鹏², 张淇昊², 曹文傑², 田云娜², 王肖婷², 王万铁^2,*

¹宜宾学院体育与大健康学院，宜宾 644000；²温州医科大学缺血/再灌注损伤研究所，温州 325035

摘要

本文旨在探讨2-脱氧葡萄糖(2-deoxyglucose, 2-DG)是否通过抑制核苷酸结合寡聚化结构域样受体蛋白3 (nucleotide- binding oligomerization domain-like receptor protein 3, NLRP3)介导的细胞焦亡改善大鼠肺缺血/再灌注(ischemia/reperfusion, I/R)损伤。雄性Sprague-Dawley大鼠随机分为对照组、2-DG组、肺I/R损伤组(I/R组)和2-DG+肺I/R损伤组(2-DG+I/R组)，每组6只。2-DG于造模前1 h腹腔注射(0.7 g/kg)；光镜下观察组织结构；检测肺损伤指标；商用试剂盒检测丙二醛(malondialdehyde, MDA)、髓过氧化物酶(myeloperoxidase, MPO)、乳酸含量；ELISA检测IL-1β、IL-18水平；Western blot、qRT-PCR和免疫荧光染色检测糖酵解、细胞焦亡相关指标表达变化。结果显示：与对照组相比，2-DG组各项指标无差异；I/R组肺损伤指标显著升高，氧化应激反应和乳酸含量显著增加，IL-1β、IL-18含量显著升高；糖酵解和细胞焦亡相关指标己糖激酶2 (hexokinase 2, HK2)、丙酮酸激酶2 (pyruvate kinase 2, PKM2)、NLRP3、Gasdermin 超家族成员(GSDMD-N)、活性半胱氨酸天冬氨酸蛋白酶1 (cleaved-Caspase1)、cleaved-IL-1β、cleaved-IL-18蛋白表达水平上调，HK2、PKM2、NLRP3基因表达水平升高，HK2、NLRP3免疫荧光表达增加；与I/R组相比，2-DG+I/R组肺泡结构及炎性浸润明显改善，各项肺损伤指标减轻，I/R诱发的糖酵解和细胞焦亡相关指标下调。上述结果提示，2-DG通过抑制NLRP3介导的细胞焦亡改善肺I/R引起的大鼠肺组织损伤。

关键词： 2-脱氧葡萄糖; 糖酵解; 细胞焦亡; 肺缺血/再灌注损伤

2-DG improves lung ischemia/reperfusion injury by inhibiting NLRP3-mediated pyroptosis in rats

SHI Lu¹, HUANG Man², CHEN Si-An², XU Jun-Peng², ZHANG Qi-Hao², CAO Wen-Jie², TIAN Yun-Na², WANG Xiao-Ting², WANG Wan-Tie^2,*

¹College of Physical Education and Health, Yibin University, Yibin 644000, China；²Institute of Ischemia/Reperfusion Injury, Wenzhou Medical University, Wenzhou 325035, China

Abstract

The aim of this study was to investigate whether the protective effect of 2-deoxyglucose (2-DG) on lung ischemia/reperfusion (I/R) injury is mediated by inhibiting nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3)-mediated pyroptosis in rats. Male Sprague-Dawley rats were randomly divided into control group, 2-DG group, lung I/R injury group (I/R group) and 2-DG+I/R group. 2-DG (0.7 g/kg) was intraperitoneally injected 1 h prior to lung ischemia. The tissue structure was measured under light microscope. Lung injury parameters were detected. The contents of malondialdehyde (MDA), myeloperoxidase (MPO) and lactate were determined by commercially available kits. ELISA was used to detect the levels of IL-1β and IL-18. Western blot, qRT-PCR and immunofluorescence staining were used to measure the expression changes of glycolysis and pyroptosis related indicators. The results showed that there was no significant difference in the parameters between the control group and the 2-DG group. However, the lung injury parameters, oxidative stress response, lactic acid content, IL-1β, and IL-18 levels were significantly increased in the I/R group. The protein expression levels of glycolysis and pyroptosis related indicators including hexokinase 2 (HK2), pyruvate kinase 2 (PKM2), NLRP3, Gasdermin superfamily member GSDMD-N, cleaved-Caspase1, cleaved-IL-1β and cleaved-IL-18, and the gene expression levels of HK2, PKM2 and NLRP3 were markedly up-regulated in the I/R group compared with those in the control group. The expression of HK2 and NLRP3 was also increased detected by immunofluorescence staining. Compared with the I/R group, the 2-DG+I/R group exhibited significantly improved alveolar structure and inflammatory infiltration, reduced lung injury parameters, and decreased expression of glycolysis and pyroptosis related indicators. These results suggest that 2-DG protects against lung I/R injury possibly by inhibiting NLRP3-mediated pyroptosis in rats.

Key words: 2-deoxyglucose; glycolysis; pyroptosis; lung ischemia/reperfusion injury

收稿日期：　　录用日期：

通讯作者：王万铁　　E-mail:

DOI: 10.13294/j.aps.2024.0050

引用本文：

石璐, 黄曼, 陈思安, 徐俊鹏, 张淇昊, 曹文傑, 田云娜, 王肖婷, 王万铁. 2-DG通过抑制NLRP3介导的细胞焦亡改善大鼠肺缺血/再灌注损伤[J]. 生理学报 2024; 76 (4): 517-525.

SHI Lu, HUANG Man, CHEN Si-An, XU Jun-Peng, ZHANG Qi-Hao, CAO Wen-Jie, TIAN Yun-Na, WANG Xiao-Ting, WANG Wan-Tie. 2-DG improves lung ischemia/reperfusion injury by inhibiting NLRP3-mediated pyroptosis in rats. Acta Physiol Sin 2024; 76 (4): 517-525 (in Chinese with English abstract).