Cylindromatosis protects against myocardial senescence in mice via the tumor necrosis factor-α pathway
滕敬华 1胡恋 1程飞 1何茜 1王洁 1丁妍 1王治校 1王馨
扫码查看
点击上方二维码区域，可以放大扫码查看
作者信息
1. 十堰市太和医院 湖北医药学院附属太和医院心血管内科，十堰 442000
折叠
摘要
目的 探讨去泛素化酶（CYLD）在小鼠心肌细胞衰老中的作用及机制。 方法 选取野生型FVB（WT）小鼠和CYLD过表达（CYLD+/+）小鼠构建自然衰老模型。2月龄雄性WT小鼠（WT-2M，n=10）和CYLD+/+小鼠（CYLD-2M，n=10）作为年轻组，普通饲养17.5月（WT-17.5M，CYLD-17.5M，n=10）作为年老组。小动物超声检测心功能[左室短轴缩短率（LVFS）、左室射血分数（LVEF）]，蛋白免疫印迹检测心肌组织衰老相关蛋白p53、p21、p16表达水平，实时荧光定量PCR（qPCR）检测心肌组织衰老相关分泌表型（SASP） [细胞因子白介素-1β（IL-1β）、基质金属蛋白酶3（MMP3）、趋化因子CXCL1（CXCL1）]转录水平。转录组测序检测CYLD调控心肌细胞衰老的信号通路，对肿瘤坏死因子α（TNF-α）通路行qPCR验证。 结果 与WT-17.5M小鼠比较，CYLD-17.5M小鼠心功能LVEF和LVFS值增高（均P<0.05）；p53、p21、p16蛋白表达水平减低（均P<0.05），IL1B、MMP3、CXCL1转录水平也下降（均P<0.05）。与WT-2M小鼠比较，WT-17.5M小鼠p53、p21、p16蛋白表达明显升高（均P<0.01），IL1B、MMP3及CXCL1转录水平上调（均P<0.01）。转录组测序发现WT-17.5M小鼠较WT-2M小鼠表达上调的信号通路64个，CYLD-17.5M小鼠较WT-17.5M小鼠表达下调的信号通路37个，二者共有的差异信号通路29个。选取TNF-α通路行qPCR验证，WT-17.5M较WT-2M小鼠TNF-α转录水平明显上调（P<0.05），且高于CYLD-17.5M小鼠（P<0.05）。 结论 CYLD对心肌自然衰老起保护作用，TNF-α通路介导可能是CYLD保护心肌衰老的作用机制之一。 Objective To investigate the role of the deubiquitinase Cylindromatosis (CYLD) in aging of mouse cardiomyocytes and the underlying mechanism. Methods Wild-type (WT) FVB mice and CYLD-overexpressing (CYLD+/+) mice were used to establish mice models of natural aging. Then, 2-month-old male wild-type FVB mice (WT-2M, n=10) and CYLD+/+ mice (CYLD-2M, n=10) were assigned to the young age group, and those routinely raised for 17.5 months (WT-17.5M, CYLD-17.5M, n=10 each) to the old age group. Small animal ultrasonography was used to measure the cardiac function (LVEF, LVFS), Western blotting was used to examine the expression of aging-related proteins p53, p21, and p16 in myocardial tissues, and qPCR was used to determine the transcription of senescence associated secretory phenotypes (SASP) interlenkin-1β (IL-1β), matrix metalloproteinase 3 (MMP3) and CXC chemokine ligand 1 (CXCL1) in myocardial tissues. Transcriptome sequencing was used to detect the signaling pathways by which CYLD regulates cardiomyocyte senescence. Tumor necrosis factor-α (TNF-α) pathway was verified by qPCR. Results Compared with WT-17.5M mice, CYLD-17.5M mice presented better cardiac function with higher values of LVEF and LVFS (all P<0.05), lower expression levels of p53, p21, and p16 proteins (allP<0.05), and less transcription of IL1B, MMP3, and CXCL1 (allP<0.05). Compared with WT-2M mice, the WT-17.5M mice had significantly higher expression of p53, p21 and p16 proteins (allP<0.01), and up-regulated transcription levels of IL1B, MMP3 and CXCL1 (allP<0.01). Transcriptome sequencing showed 64 up-regulated signaling pathways in WT-17.5M mice when comparing to WT-2M mice, and 37 down-regulated signaling pathways in CYLD-17.5M mice when comparing to WT-17.5M mice there were 29 pathways with differential signaling when comparing CYLD-17.5M to WT-17.5M mice. qPCR with TNF-α pathway selected for verification showed significantly increased TNF-α transcription in WT-17.5M mice than in WT-2M mice and CYLD-17.5M mice (allP<0.05) . Conclusion CYLD plays a protective role in natural cardiomyocyte senescence. Mediation of the TNF-α pathway may be one of mechanisms underlying the effect of CYLD against myocardial aging.
Abstract
Objective To investigate the role of the deubiquitinase Cylindromatosis (CYLD) in aging of mouse cardiomyocytes and the underlying mechanism. Methods Wild-type (WT) FVB mice and CYLD-overexpressing (CYLD+/+) mice were used to establish mice models of natural aging. Then, 2-month-old male wild-type FVB mice (WT-2M, n=10) and CYLD+/+ mice (CYLD-2M, n=10) were assigned to the young age group, and those routinely raised for 17.5 months (WT-17.5M, CYLD-17.5M, n=10 each) to the old age group. Small animal ultrasonography was used to measure the cardiac function (LVEF, LVFS), Western blotting was used to examine the expression of aging-related proteins p53, p21, and p16 in myocardial tissues, and qPCR was used to determine the transcription of senescence associated secretory phenotypes (SASP) interlenkin-1β (IL-1β), matrix metalloproteinase 3 (MMP3) and CXC chemokine ligand 1 (CXCL1) in myocardial tissues. Transcriptome sequencing was used to detect the signaling pathways by which CYLD regulates cardiomyocyte senescence. Tumor necrosis factor-α (TNF-α) pathway was verified by qPCR. Results Compared with WT-17.5M mice, CYLD-17.5M mice presented better cardiac function with higher values of LVEF and LVFS (all P<0.05), lower expression levels of p53, p21, and p16 proteins (allP<0.05), and less transcription of IL1B, MMP3, and CXCL1 (allP<0.05). Compared with WT-2M mice, the WT-17.5M mice had significantly higher expression of p53, p21 and p16 proteins (allP<0.01), and up-regulated transcription levels of IL1B, MMP3 and CXCL1 (allP<0.01). Transcriptome sequencing showed 64 up-regulated signaling pathways in WT-17.5M mice when comparing to WT-2M mice, and 37 down-regulated signaling pathways in CYLD-17.5M mice when comparing to WT-17.5M mice there were 29 pathways with differential signaling when comparing CYLD-17.5M to WT-17.5M mice. qPCR with TNF-α pathway selected for verification showed significantly increased TNF-α transcription in WT-17.5M mice than in WT-2M mice and CYLD-17.5M mice (allP<0.05) . Conclusion CYLD plays a protective role in natural cardiomyocyte senescence. Mediation of the TNF-α pathway may be one of mechanisms underlying the effect of CYLD against myocardial aging.
万方数据