Ivabradine (IVA) reduces heart rate by inhibiting hyperpolarization-activated cyclic nucleotide-gated channels (HCNs), which play a role in the promotion of pacemaker activity in cardiac sinoatrial node cells. HCNs are highly expressed in neural and myocardial tissues and are involved in the modulation of inflammatory neuropathic pain. However, whether IVA exerts any effect on myocardial inflammation in the pathogenesis of heart failure is unclear. We employed single-cell RNA sequencing (scRNA-seq) in porcine cardiac myosin-induced experimental autoimmune myocarditis rat model to determine the effects and mechanisms of IVA. Lewis rats (n = 32) were randomly divided into the normal, control, high-dose-IVA, and low-dose-IVA groups. Heart rate and blood pressure were measured on days 0 and 21, respectively. Echocardiography was performed on day 22, and inflammation of the myocardium was evaluated via histopathological examination. Western blot was employed to detect the expression of HCN1-4, MinK-related protein 1 (MiRP1), matrix metalloproteinase 2 (MMP-2), MMP-9, and transforming growth factor-β (TGF-β). Furthermore, enzyme-linked immunosorbent assay was performed to measure serum IL-1, IL-6, and TNF-α. The relative mRNA levels of collagen I, collagen III, and α-smooth muscle actin (α-SMA) were determined via qRT-PCR. We found that IVA reduced the total number of cells infiltrated into the myocardium, particularly in the subset of fibroblasts, endocardia, and monocytes. IVA administration ameliorated cardiac inflammation and reduced collagen production. Results of the echocardiography indicated that left ventricular internal diameter at end-systole LVIDs increased whereas left ventricular ejection fraction and left ventricular fractional shortening decreased in the control group. IVA improved cardiac performance. The expression of HCN4 and MiRP1 protein and the level of serum IL-1, IL-6, and TNF-α were decreased by IVA treatment. In conclusion, HCNs and the helper proteins were increased in the profile of myocardial inflammation. HCNs may be involved in the regulation of myocardial inflammation by inhibiting immune cell infiltration. Our findings can contribute to the development of IVA-based combination therapies for the future treatment of cardiac inflammation and heart failure.

伊伐布雷定 (IVA) 通过抑制超极化激活的环核苷酸门控通道 (HCN) 来降低心率，HCN 在促进心脏窦房结细胞的起搏器活动中发挥作用。 HCN 在神经和心肌组织中高表达，并参与炎症性神经病理性疼痛的调节。然而，IVA是否对心力衰竭发病机制中的心肌炎症产生影响尚不清楚。我们在猪心肌肌球蛋白诱导的实验性自身免疫性心肌炎大鼠模型中采用单细胞 RNA 测序 (scRNA-seq) 来确定 IVA 的作用和机制。 Lewis大鼠（n = 32）被随机分为正常组、对照组、高剂量IVA组和低剂量IVA组。分别在第 0 天和第 21 天测量心率和血压。第22天进行超声心动图检查，并通过组织病理学检查评估心肌炎症。 Western blot检测HCN1-4、MinK相关蛋白1（MiRP1）、基质金属蛋白酶2（MMP-2）、MMP-9、转化生长因子-β（TGF-β）的表达。此外，采用酶联免疫吸附法测定血清IL-1、IL-6和TNF-α。通过 qRT-PCR 测定胶原蛋白 I、胶原蛋白 III 和 α-平滑肌肌动蛋白 (α-SMA) 的相对 mRNA 水平。我们发现 IVA 减少了渗入心肌的细胞总数，特别是在成纤维细胞、心内膜和单核细胞亚群中。 IVA 给药可改善心脏炎症并减少胶原蛋白的产生。超声心动图结果显示，对照组收缩末期 LVID 时左心室内径增大，而左心室射血分数和左心室缩短分数降低。 IVA 改善心脏功能。 IVA治疗降低了HCN4和MiRP1蛋白的表达以及血清IL-1、IL-6和TNF-α的水平。总之，HCN 和辅助蛋白在心肌炎症中有所增加。 HCN 可能通过抑制免疫细胞浸润来调节心肌炎症。我们的研究结果有助于开发基于 IVA 的联合疗法，用于未来治疗心脏炎症和心力衰竭。