Background:The sino atrial node (SAN) is characterized by the microenvironment of pacemaker cardiomyocytes (PCs) encased with fibroblasts. An altered microenvironment leads to rhythm failure. Operable cell or tissue models are either generally lacking or difficult to handle. The biological process behind the milieu of SANs to evoke pacemaker rhythm is unknown. We explored how fibroblasts interact with PCs and regulate metabolic reprogramming and rhythmic activity in the SAN.Methods:Tbx18 (T-box transcription factor 18)-induced PCs and fibroblasts were used for cocultures and engineered tissues, which were used as the in vitro models to explore how fibroblasts regulate the functional integrity of SANs. RNA-sequencing, metabolomics, and cellular and molecular techniques were applied to characterize the molecular signals underlying metabolic reprogramming and identify its critical regulators. These pathways were further validated in vivo in rodents and induced human pluripotent stem cell-derived cardiomyocytes.Results:We observed that rhythmicity in Tbx18-induced PCs was regulated by aerobic glycolysis. Fibroblasts critically activated metabolic reprogramming and aerobic glycolysis within PCs, and, therefore, regulated pacemaker activity in PCs. The metabolic reprogramming was attributed to the exclusive induction of Aldoc (aldolase c) within PCs after fibroblast-PC integration. Fibroblasts activated the integrin-dependent mitogen-activated protein kinase-E2F1 signal through cell-cell contact and turned on Aldoc expression in PCs. Interruption of fibroblast-PC interaction or Aldoc knockdown nullified electrical activity. Engineered Tbx18-PC tissue sheets were generated to recapitulate the microenvironment within SANs. Aldoc-driven rhythmic machinery could be replicated within tissue sheets. Similar machinery was faithfully validated in de novo PCs of adult mice and rats, and in human PCs derived from induced pluripotent stem cells.Conclusions:Fibroblasts drive Aldoc-mediated metabolic reprogramming and rhythmic regulation in SANs. This work details the cellular machinery behind the complex milieu of vertebrate SANs and opens a new direction for future therapy.

中文翻译：

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成纤维细胞驱动起搏器心肌细胞的代谢重编程

背景：窦房结（SAN）的特点是起搏器心肌细胞（PCs）包裹着成纤维细胞的微环境。改变的微环境会导致节律失常。可操作的细胞或组织模型通常缺乏或难以处理。SAN 环境背后引发起搏器节律的生物学过程尚不清楚。我们探讨了成纤维细胞如何与 PC 相互作用并调节 SAN 中的代谢重编程和节律活动。方法：将 Tbx18（T-box 转录因子 18）诱导的 PC 和成纤维细胞用于共培养和工程组织，用作体外模型探索成纤维细胞如何调节 SAN 的功能完整性。RNA测序，代谢组学，并应用细胞和分子技术来表征代谢重编程的分子信号并确定其关键调节因子。这些通路在啮齿类动物体内得到了进一步验证，并诱导了人多能干细胞衍生的心肌细胞。结果：我们观察到 Tbx18 诱导的 PC 的节律性受有氧糖酵解的调节。成纤维细胞严重激活了 PC 内的代谢重编程和有氧糖酵解，因此调节了 PC 中的起搏器活性。代谢重编程归因于成纤维细胞-PC 整合后 PC 内 Aldoc（醛缩酶 c）的独家诱导。成纤维细胞通过细胞间接触激活整合素依赖性丝裂原活化蛋白激酶-E2F1 信号，并开启 PC 中的 Aldoc 表达。成纤维细胞-PC 相互作用的中断或 Aldoc 敲低使电活动无效。生成工程 Tbx18-PC 组织片以概括 SAN 内的微环境。Aldoc 驱动的节律机制可以在组织片内复制。类似的机制在成年小鼠和大鼠的从头 PC 以及从诱导多能干细胞衍生的人类 PC 中得到了忠实的验证。结论：成纤维细胞在 SAN 中驱动 Aldoc 介导的代谢重编程和节律调节。这项工作详细介绍了脊椎动物 SAN 复杂环境背后的细胞机制，并为未来的治疗开辟了新方向。类似的机制在成年小鼠和大鼠的从头 PC 以及从诱导多能干细胞衍生的人类 PC 中得到了忠实的验证。结论：成纤维细胞在 SAN 中驱动 Aldoc 介导的代谢重编程和节律调节。这项工作详细介绍了脊椎动物 SAN 复杂环境背后的细胞机制，并为未来的治疗开辟了新方向。类似的机制在成年小鼠和大鼠的从头 PC 以及从诱导多能干细胞衍生的人类 PC 中得到了忠实的验证。结论：成纤维细胞在 SAN 中驱动 Aldoc 介导的代谢重编程和节律调节。这项工作详细介绍了脊椎动物 SAN 复杂环境背后的细胞机制，并为未来的治疗开辟了新方向。