Zhang Wanliu, Lu Jingqian, Gao Yan, Song Qianhong, Luo Shihua, Li Yi
Department of Cardiology, Yunnan Provincial Hospital of Traditional Chinese Medicine, The First Affiliated Hospital of Yunnan University of Chinese Medicine, Kunming, Yunnan Province, PR China.
Histol Histopathol. 2025 May;40(5):773-784. doi: 10.14670/HH-18-820. Epub 2024 Sep 24.
From a clinical standpoint, myocardial ischemia/reperfusion injury (MIRI) has always been an enormous challenge for the treatment of acute myocardial infarction (AMI). Molecular targeting therapy may help overcome this challenge. The present work aimed to elucidate the possible involvement of Yin-Yang 1 (YY1)/nuclear receptor-interacting protein 1 (NRIP1) and discover the molecular mechanism of MIRI.
Herein, a cardiomyocyte ischemia/reperfusion (I/R) model was established via oxygen-glucose deprivation/re-oxygenation (OGD/R) damage in H9c2 cardiomyocytes. Reverse transcription-quantitative PCR and western blotting were conducted to measure the levels of YY1 and NRIP1 at the RNA and protein levels, respectively. H9c2 cell viability and apoptosis were assayed using the Cell Counting Kit-8, flow cytometry, and western blotting. In addition, superoxide dismutase, glutathione peroxidase, and malondialdehyde levels were analyzed as markers of oxidative stress. Additionally, mitochondrial membrane potential, which was measured via JC-1 staining, ATP content, Complex I activity, mitochondrial DNA copy number, and mitochondrial permeability transition pore (mPTP) opening rate were analyzed to evaluate mitochondrial activity. Moreover, luciferase reporter and chromatin immunoprecipitation assays experimentally validated the predicted affinity of YY1 with the NRIP1 promoter according to the HumanTFDB online tool.
YY1/NRIP1 were both highly expressed in OGD/R-injured H9c2 cardiomyocytes. Downregulation of NRIP1 improved cell viability, whereas it inhibited cell apoptosis and oxidative stress, and suppressed mitochondrial dysfunction in OGD/R-injured H9c2 cardiomyocytes. Importantly, it was verified that YY1 could bind to the NRIP1 promoter to positively regulate NRIP1 expression. The protective effects of NRIP1 knockdown against cardiomyocyte damage and mitochondrial dysfunction in OGD/R-injured H9c2 cardiomyocytes were partly abolished through overexpression of YY1.
NRIP1 emerged as a downstream target of YY1 in promoting OGD/R-induced H9c2 cardiomyocyte injury and mitochondrial dysfunction, providing novel ideas for targeted treatments to alleviate MIRI.
从临床角度来看，心肌缺血/再灌注损伤（MIRI）一直是急性心肌梗死（AMI）治疗中的巨大挑战。分子靶向治疗可能有助于克服这一挑战。本研究旨在阐明阴阳1（YY1）/核受体相互作用蛋白1（NRIP1）可能的参与情况，并发现MIRI的分子机制。
在此，通过对H9c2心肌细胞进行氧糖剥夺/复氧（OGD/R）损伤建立心肌细胞缺血/再灌注（I/R）模型。分别通过逆转录定量PCR和蛋白质免疫印迹法检测YY1和NRIP1在RNA和蛋白质水平的表达。使用细胞计数试剂盒-8、流式细胞术和蛋白质免疫印迹法检测H9c2细胞活力和凋亡情况。此外，分析超氧化物歧化酶、谷胱甘肽过氧化物酶和丙二醛水平作为氧化应激指标。另外，通过JC-1染色测量线粒体膜电位、ATP含量、复合体I活性、线粒体DNA拷贝数以及线粒体通透性转换孔（mPTP）开放率，以评估线粒体活性。此外，根据HumanTFDB在线工具，通过荧光素酶报告基因和染色质免疫沉淀实验验证了YY1与NRIP1启动子的预测亲和力。
YY1/NRIP1在OGD/R损伤的H9c2心肌细胞中均高表达。下调NRIP1可提高细胞活力，抑制细胞凋亡和氧化应激，并抑制OGD/R损伤的H9c2心肌细胞的线粒体功能障碍。重要的是，证实了YY1可与NRIP1启动子结合以正向调节NRIP1表达。通过过表达YY1，部分消除了NRIP1敲低对OGD/R损伤的H9c2心肌细胞的心肌损伤和线粒体功能障碍的保护作用。
NRIP1作为YY1的下游靶点，在促进OGD/R诱导的H9c2心肌细胞损伤和线粒体功能障碍中发挥作用，为减轻MIRI的靶向治疗提供了新思路。
