Lv Zi, Lv Ding-Yi, Meng Jia-Yu, Sha Xiao-Yan, Qian Xue-Ya, Chen Yun-Shan, Pan Xiu-Yu, Yu Guang-Yuan, Liu Hui-Shu
Department of Obstetrics, First Affiliated Hospital of Jinan University, 9 Jinsui Road, Guangzhou, China; Department of Obstetrics, Guangzhou Women and Children's Medical Center, Guangzhou Medical University, 9 Jinsui Road, Guangzhou, China.
Department of Cardiology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China.
Int Immunopharmacol. 2023 Jan;114:109523. doi: 10.1016/j.intimp.2022.109523. Epub 2022 Dec 9.
Preeclampsia (PE) is characterised by systemic vascular endothelium dysfunction. Circulating trophoblastic secretions contribute to endothelial dysfunction, resulting in PE; however, the underlying mechanisms remain unclear. Herein, we aimed to determine the potential correlation between the release of trophoblastic mitochondrial deoxyribonucleic acid (DNA) (mtDNA) and endothelium damage in PE.
Umbilical cord sera and tissues from patients with PE were investigated for inflammasome activation. Following this, trophoblastic mitochondria were isolated from HTR-8/SVneo trophoblasts under 21 % oxygen (O) or hypoxic conditions (1 % O for 48 h) for subsequent treatments. Primary human umbilical veinendothelial cells (HUVECs) were isolated from the human umbilical cord and then exposed to a vehicle (phosphate-buffered saline [PBS]), mtDNA, hypo-mtDNA, or hypo-mtDNA with INF39 (nucleotide oligomerisation domain-like receptor family pyrin domain containing 3 [NLRP3]-specific inhibitor) for 12 h before flow cytometry and immunoblotting. The effects of trophoblastic mtDNA on the endothelium were further analysed in vivo using enzyme-linked immunosorbent assay (ELISA) and vascular reactivity assay. The effects of mtDNA on vascular phenotypes were also tested on NLRP3 knockout mice.
Elevated interleukin (IL)-1β in PE sera was accompanied by NLRP3 inflammasome activation in cord tissues. In vitro and in vivo experiments revealed that the release of trophoblastic mtDNA could damage the endothelium via NLRP3 activation, resulting in the overexpression of NLRP3, caspase-1 p20, IL-1β p17, and gasdermin D (GSDMD); reduced endothelial nitric oxide synthase (eNOS) levels; and impaired vascular relaxation. Flow cytometric analysis confirmed that extensive cell death was induced by mtDNA, and simultaneously, a more pronounced pro-apoptotic effect was caused by hypoxia-treated trophoblastic mtDNA. The NLRP3 knockout or pharmacologic NLRP3 inhibition partially reversed tumour necrosis factor-α (TNF-α) and IL-1β levels and endothelium-dependent vasodilation in mice.
These findings demonstrate that trophoblastic mtDNA induced NLRP3/caspase-1/IL-1β signalling activation, eNOS-related endothelial injury, and vasodilation dysfunction in PE.
子痫前期（PE）的特征是全身血管内皮功能障碍。循环中的滋养层分泌物导致内皮功能障碍，进而引发子痫前期；然而，其潜在机制仍不清楚。在此，我们旨在确定滋养层线粒体脱氧核糖核酸（DNA）（mtDNA）释放与子痫前期内皮损伤之间的潜在关联。
对PE患者的脐带血清和组织进行炎性小体激活检测。此后，在21%氧气（O₂）或低氧条件（1% O₂，持续48小时）下，从HTR-8/SVneo滋养层细胞中分离出滋养层线粒体，用于后续处理。从人脐带中分离出原代人脐静脉内皮细胞（HUVECs），然后将其暴露于溶媒（磷酸盐缓冲盐水 [PBS]）、mtDNA、低剂量mtDNA或添加INF39（含核苷酸寡聚化结构域样受体家族吡啉结构域3 [NLRP3]特异性抑制剂）的低剂量mtDNA中12小时，之后进行流式细胞术检测和免疫印迹分析。使用酶联免疫吸附测定（ELISA）和血管反应性测定在体内进一步分析滋养层mtDNA对内皮的影响。还在NLRP3基因敲除小鼠上测试了mtDNA对血管表型的影响。
PE血清中白细胞介素（IL）-1β升高，同时脐带组织中NLRP3炎性小体激活。体外和体内实验表明，滋养层mtDNA的释放可通过激活NLRP3损伤内皮，导致NLRP3、半胱天冬酶-1 p20、IL-1β p17和gasdermin D（GSDMD）过度表达；降低内皮型一氧化氮合酶（eNOS）水平；并损害血管舒张功能。流式细胞术分析证实，mtDNA可诱导广泛的细胞死亡，同时，低氧处理的滋养层mtDNA可产生更明显的促凋亡作用。NLRP3基因敲除或药理学抑制NLRP3可部分逆转小鼠体内肿瘤坏死因子-α（TNF-α）和IL-1β水平以及内皮依赖性血管舒张功能。
这些发现表明，滋养层mtDNA可诱导子痫前期中NLRP3/半胱天冬酶-1/IL-1β信号激活、与eNOS相关的内皮损伤以及血管舒张功能障碍。
