SNRK facilitates cardiac repair associated with nonischemic fibrosis: regulating transforming growth factor-beta1 levels in atrial cardiomyocytes

Thirugnanam, Karthikeyan1; Rizvi, Farhan2; Jahangir, Arshad2; Homar, Peter2; Shabnam, Fathima3; Palecek, Sean P.3; Kumar, Suresh N.4; Pan, Amy5; Bai, Xiaowen6; Sekine, Hidekazu7; Ramchandran, Ramani1

1Department of Pediatrics, Division of Neonatology, Developmental Vascular Biology Program, Children’s Research Institute (CRI), Milwaukee, WI, USA

2Aurora Cardiovascular and Thoracic Services, Advocate Health, Milwaukee, WI, USA

3Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI, USA

4Department of Pathology, Division of Pediatric Pathology, Medical College of Wisconsin, Milwaukee, WI, USA

5Department of Pediatrics, Division of Bioinformatics and Quantitative Child Health, Milwaukee, WI, USA

6Department of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, WI, USA

7Institute of Advanced Biomedical Engineering and Science, Tokyo Women’s Medical University, Tokyo, Japan

*Correspondence to: Ramani Ramchandran, PhD, rramchan@mcw.edu.

Abstract

Heart failure is a pressing clinical condition that is expected to increase as our population ages and thus requires better treatment options. Identifying the precise mechanisms that underlie fibrosis and inflammation, two key features associated with cardiac repair and regeneration during ischemic and nonischemic heart failure, is likely to provide effective strategies for the clinical intervention of heart failure. This study investigated a metabolic serine threonine kinase gene, sucrose nonfermenting-related kinase (SNRK), which we previously reported to control cardiac metabolism and function. Conditional knockout of Snrk in mouse cardiomyocytes (Snrk cmcKO) leads to deleterious fibrosis, inflammation, and, subsequently, heart failure. The precise mechanism underlying cardiomyocyte SNRK-driven repression of deleterious cardiac fibrosis in nonischemic heart failure-mediated cardiac repair and regeneration is not known. Here, using mouse, rat, and human tissues, we demonstrated that SNRK expression is increased in the atrial chamber, especially in left atrial cardiomyocytes. Using a nonischemic heart failure mouse model, we showed that fibrosis in the atria, particularly the left atria, is associated with cardiac functional decline. To elucidate the mechanistic pathway responsible for the SNRK-mediated repression of cardiac fibrosis, we focused on the profibrotic protein transforming growth factor-β1. Transforming growth factor-β1 levels in Snrk siRNA-knockdown HL-1 adult immortalized mouse atrial cells were higher compared with control siRNA-knockdown HL-1 cells. Coculture of HL-1 cardiomyocytes (-/+ Snrk) with cardiac fibroblasts in vitro revealed that SNRK represses transforming growth factor-β1 signaling (Smad2/3) in cardiac fibroblasts and cardiac fibroblast activation (alpha-smooth muscle actin marker). We conclude that under nonischemic heart failure conditions, increased SNRK expression in the atria is associated with a cardioprotective mechanism by controlling the release of the profibrotic transforming growth factor-β1 factor. These studies illuminate a potential deleterious fibrosis pathway for intervention during cardiac repair and regeneration in nonischemic heart failure.

SNRK促进非缺血性纤维化相关的心脏修复：调节心房心肌细胞中的 TGFβ 水平

摘要

纤维化和炎症是缺血性和非缺血性心力衰竭期间心脏修复和再生的两个关键特征，找出这两个特征的确切机制可能会为心力衰竭的临床干预提供有效的策略。实验旨在了解与控制心脏代谢和功能相关的代谢丝氨酸苏氨酸激酶基因--非发酵相关的蛋白激酶（SNRK）促进非缺血性纤维化相关的心脏修复的机制。在小鼠心肌细胞中条件性敲除 Snrk（Snrk cmcKO）会导致有害的纤维化、炎症和随后的高房颤。在非缺血性高频介导的心脏修复和再生过程中，心肌细胞 SNRK 驱动的抑制有害心脏纤维化的确切机制尚不清楚。实验利用小鼠、大鼠和人体组织证明，SNRK 在心房腔中表达较高，尤其是在左心房心肌细胞中。实验在非缺血性高频小鼠模型中发现心房，尤其是左心房的纤维化与心脏功能衰退有关。为了阐明 SNRK 介导的抑制心脏纤维化的机制途径，实验重点研究了促纤维化蛋白转化生长因子β1水平。与对照组siRNA 敲除的HL-1 细胞相比，Snrk siRNA 敲除的 HL-1 成体永生小鼠心房细胞中的转化生长因子β1 含量更高。体外将 HL-1 心肌细胞（-/+ Snrk）与心成纤维细胞共培养显示，SNRK 可抑制 HL-1 细胞中的转化生长因子β1信号传导（Smad 2/3），并阻止旁分泌性心成纤维细胞活化（α-SMA 水平）。这些结果表明，在非缺血性高房颤动情况下，心房中较高的 SNRK 表达与通过控制促纤维化转化生长因子β1因子的释放来保护心脏的机制有关。这阐明了在非缺血性高房颤动的心脏修复和再生过程中进行干预的潜在有害纤维化途径。