Rouhi Leila, Cheedipudi Sirisha M, Cathcart Benjamin, Gurha Priyatansh, Marian Ali J
Center for Cardiovascular Genetics, Institute of Molecular Medicine, The University of Texas Health Science Center. Houston TX 77030, USA.
J Cardiovasc Aging. 2023;3(3). doi: 10.20517/jca.2023.20. Epub 2023 Jul 10.
The genome is constantly exposed to numerous stressors, which induce DNA lesions, including double-stranded DNA breaks (DSBs). DSBs are the most dangerous, as they induce genomic instability. In response to DNA damage, the cell activates nuclear DNA damage response (DDR) and the cytosolic DNA sensing protein (CDSP) pathways, the latter upon release of the DSBs to the cytosol. The CDSP pathway activates NFκB and IRF3, which induce the expression of the pro-inflammatory genes. There is scant data on the activation of the CDSP pathway in human hearts with dilated cardiomyopathy (DCM).
We aimed to determine expression levels of selected components of the CDSP pathway in human hearts with DCM.
The DNA strand breaks were detected by the single-cell gel electrophoresis or the comet assay and expression of selected proteins by immunoblotting. Transcript levels were quantified in the RNA-Seq data.
Single-cell gel electrophoresis showed an approximately 2-fold increase in the number of COMET cells in the DCM hearts. Immunoblotting showed increased levels of cyclic GMP-AMP synthase (CGAS), the canonical CDSP; TANK-binding kinase 1 (TBK1), an intermediary kinase in the pathway; and RELB, P52, and P50 components of the NFκB pathway in human heart samples from patients with DCM. Likewise, transcript levels of over 2 dozen genes involved in inflammatory responses were increased.
The findings provide the first set of evidence for the activation of the CDSP pathway in human hearts with DCM. The data in conjunction with the previous evidence of activation of the DDR pathway implicate the DSBs in the pathogenesis of human DCM.
基因组不断受到多种应激源的影响，这些应激源会导致DNA损伤，包括双链DNA断裂（DSB）。DSB是最危险的，因为它们会导致基因组不稳定。作为对DNA损伤的反应，细胞会激活核DNA损伤反应（DDR）和胞质DNA传感蛋白（CDSP）途径，后者在DSB释放到胞质溶胶时被激活。CDSP途径激活NFκB和IRF3，从而诱导促炎基因的表达。关于扩张型心肌病（DCM）患者心脏中CDSP途径激活的数据很少。
我们旨在确定DCM患者心脏中CDSP途径选定成分的表达水平。
通过单细胞凝胶电泳或彗星试验检测DNA链断裂，并通过免疫印迹检测选定蛋白质的表达。在RNA测序数据中对转录水平进行定量。
单细胞凝胶电泳显示DCM心脏中彗星细胞数量增加了约2倍。免疫印迹显示，在DCM患者的心脏样本中，典型的CDSP——环磷酸鸟苷-腺苷合酶（CGAS）水平升高；该途径的中间激酶——TANK结合激酶1（TBK1）水平升高；以及NFκB途径的RELB、P52和P50成分水平升高。同样，超过20种参与炎症反应的基因的转录水平也升高。
这些发现为DCM患者心脏中CDSP途径的激活提供了第一组证据。这些数据与之前DDR途径激活的证据一起表明DSB参与了人类DCM的发病机制。
