Heart failure is associated with myocardial fibrosis,a pivotal histopathological feature arising from β-adrenergic receptor(β-AR) stimulation through sympathetic nervous system activation.Augmented glutaminolysis wi...Heart failure is associated with myocardial fibrosis,a pivotal histopathological feature arising from β-adrenergic receptor(β-AR) stimulation through sympathetic nervous system activation.Augmented glutaminolysis with increased bioavailability of α-ketoglutarate(α-KG) is suggested to contribute to fibrogenesis and changes in cellular gene expression.K_(Ca)3.1 is a calcium-activated potassium channel expressed in fibroblasts and has been implicated in mediating fibrosis,yet the putative interactions between glutaminolysis and K_(Ca)3.1 in β-ARmediated cardiac fibrosis remain poorly understood.Here,we performed a series of in vitro and in vivo experiments to investigate how α-KG might influence the expression of K_(Ca)3.1 in the context of experimental myocardial fibrosis driven by β-AR activation.In cultured adult mouse cardiac fibroblasts,α-KG exposure resulted in the upregulation of K_(Ca)3.1 m RNA and protein levels that were commensurate with the dose and duration of exposure,and also led to increased K_(Ca)3.1 channel currents.Exposure to α-KG led to a significant decrease in levels of histone methylation(H3K27me3) within the K_(Ca)3.1 promoter,a decrease in the association of the transcription repressor REST from this site,as well as an enrichment of transcription activator AP-1 binding.The exacerbated fibrotic signaling induced by α-KG in cultured fibroblasts was suppressed by functional inhibition of K_(Ca)3.1 or by genetic knockdown of Kcnn4.Moreover,β-AR activation by isoproterenol significantly augmented glutaminolysis mediated by glutaminase 1(GLS1) and significantly increased α-KG levels detected in the supernatant of cultured fibroblasts and cardiomyocytes.In addition,isoproterenol-induced K_(Ca)3.1 expression in fibroblasts was curtailed by treatment with the GLS1 inhibitor CB-839,or by GLS1 gene knockdown,or by treatment with the selective β_2-AR antagonist,ICI118551.In mouse models of established cardiac fibrosis evoked by isoproterenol-stimulation or β_2-AR overexpression,treatment with CB-839 for 4 weeks suppressed the phenotypic features of fibrosis,and this was associated with a decline in α-KG tissue content,a lack of histone demethylation at the K_(Ca)3.1 promoter,as well as suppression of K_(Ca)3.1 expression.Taken together,our study demonstrates for the first time that glutaminolysis contributes to β-AR activation-induced myocardial fibrosis via α-KG-stimulated K_(Ca)3.1 expression.We anticipate that treatments which target the β-AR/GLS1/α-KG/K_(Ca)3.1 signaling pathway might be effective for cardiac fibrosis.展开更多
基金supported by the National Natural Science Foundation of China (82170298,82070393,32171103,82270327)。
文摘Heart failure is associated with myocardial fibrosis,a pivotal histopathological feature arising from β-adrenergic receptor(β-AR) stimulation through sympathetic nervous system activation.Augmented glutaminolysis with increased bioavailability of α-ketoglutarate(α-KG) is suggested to contribute to fibrogenesis and changes in cellular gene expression.K_(Ca)3.1 is a calcium-activated potassium channel expressed in fibroblasts and has been implicated in mediating fibrosis,yet the putative interactions between glutaminolysis and K_(Ca)3.1 in β-ARmediated cardiac fibrosis remain poorly understood.Here,we performed a series of in vitro and in vivo experiments to investigate how α-KG might influence the expression of K_(Ca)3.1 in the context of experimental myocardial fibrosis driven by β-AR activation.In cultured adult mouse cardiac fibroblasts,α-KG exposure resulted in the upregulation of K_(Ca)3.1 m RNA and protein levels that were commensurate with the dose and duration of exposure,and also led to increased K_(Ca)3.1 channel currents.Exposure to α-KG led to a significant decrease in levels of histone methylation(H3K27me3) within the K_(Ca)3.1 promoter,a decrease in the association of the transcription repressor REST from this site,as well as an enrichment of transcription activator AP-1 binding.The exacerbated fibrotic signaling induced by α-KG in cultured fibroblasts was suppressed by functional inhibition of K_(Ca)3.1 or by genetic knockdown of Kcnn4.Moreover,β-AR activation by isoproterenol significantly augmented glutaminolysis mediated by glutaminase 1(GLS1) and significantly increased α-KG levels detected in the supernatant of cultured fibroblasts and cardiomyocytes.In addition,isoproterenol-induced K_(Ca)3.1 expression in fibroblasts was curtailed by treatment with the GLS1 inhibitor CB-839,or by GLS1 gene knockdown,or by treatment with the selective β_2-AR antagonist,ICI118551.In mouse models of established cardiac fibrosis evoked by isoproterenol-stimulation or β_2-AR overexpression,treatment with CB-839 for 4 weeks suppressed the phenotypic features of fibrosis,and this was associated with a decline in α-KG tissue content,a lack of histone demethylation at the K_(Ca)3.1 promoter,as well as suppression of K_(Ca)3.1 expression.Taken together,our study demonstrates for the first time that glutaminolysis contributes to β-AR activation-induced myocardial fibrosis via α-KG-stimulated K_(Ca)3.1 expression.We anticipate that treatments which target the β-AR/GLS1/α-KG/K_(Ca)3.1 signaling pathway might be effective for cardiac fibrosis.
