Distinct effects of genetic and pharmacological inhibition of galectin-3 on cardiac fibrosis and function in a mouse model of dilated cardiomyopathy

Background: Galectin-3 is upregulated in heart disease, and its inhibition has been reported to confer benefits on cardiac remodeling and dysfunction. It remains unknown whether galectin-3 plays a biological role in a setting of dilated cardiomyopathy (DCM). We determined galectin-3 expression in transgenic (TG) mice with cardiac-restricted overexpression of mammalian sterile 20-like kinase 1 (Mst1-TG), and studied the effects of pharmacological and genetic inhibition of galectin-3 on the DCM phenotype. Methods and results: The DCM phenotype of male Mst1-TG mice was assessed at 2, 3 and 6 months of age. Galectin-3 deletion in Mst1-TG mice was achieved by cross-breeding Mst1-TG with galectin-3 knockout (KO) mice to produce genotypes of non-TG (nTG), KO, Mst1-TG, and Mst1-TG mice with partial or complete deletion of the galectin-3 gene. Pharmacological inhibition of galectin-3 was tested by treating Mst1-TG mice with modified citrus pectin for 4 months. Changes in the DCM phenotype were assessed by serial echocardiography, ECG recording and biochemical assays. Mst1-TG mice exhibited upregulated levels of cardiac galectin-3 expression by 40~50-fold, atrial dilatation, reduced left ventricular (LV) ejection fraction, increased LV volume at systole and diastole, atrial conduction delay, severe cardiac fibrosis, and upregulated fibrosis-related genes. Galectin-3 gene deletion in Mst1-TG mice attenuates increased LV volume at systole and diastole, atrial conduction delay and cardiac fibrogenesis. Treatment with modified citrus pectin in Mst1-TG mice did not confer benefit on the DCM phenotype, possibly due to the very high expression of galectin-3. Conclusion: Galectin-3 expression is markedly increased in Mst1-TG hearts exhibiting DCM. Galectin-3 gene deletion effectively suppressed cardiac fibrotic signaling and LV dilatation, and was associated with better function. Thus, suppression of galectin-3 expression may represent a potential therapeutic approach in DCM. The development of new galectin-3 inhibitors are warranted. We performed RNA-seq on left ventricle samples from control and Gal-3 knock-out mice in two genetic backgrounds: wild type C57BL6 and the Mst-1 overexpressing mouse that is frequently used as a model for dilated cardiomyopathy. Between 5 and 8 replicates from each group were processed.背景：半乳糖凝集素3（Galectin-3）在心脏疾病中表达上调，已有研究证实其抑制可改善心肌重构与心功能障碍。目前尚不明确半乳糖凝集素3在扩张型心肌病（dilated cardiomyopathy, DCM）发生发展中是否发挥生物学作用。本研究检测了心肌特异性过表达哺乳动物不育系20样激酶1（mammalian sterile 20-like kinase 1, Mst1-TG）的转基因（transgenic, TG）小鼠体内半乳糖凝集素3的表达水平，并探讨了半乳糖凝集素3的药物与基因抑制手段对扩张型心肌病表型的影响。 方法与结果：本研究于2、3及6月龄时评估雄性Mst1-TG小鼠的扩张型心肌病表型。通过将Mst1-TG小鼠与半乳糖凝集素3敲除（knockout, KO）小鼠杂交，获得非转基因（non-TG, nTG）、KO、Mst1-TG以及部分或完全敲除半乳糖凝集素3基因的Mst1-TG小鼠，以此实现Mst1-TG小鼠中半乳糖凝集素3的基因敲除。采用修饰柑橘果胶（modified citrus pectin）处理Mst1-TG小鼠4个月，以评估半乳糖凝集素3的药物抑制效果。通过系列超声心动图、心电图记录及生化检测评估扩张型心肌病表型的变化。结果显示，Mst1-TG小鼠的心肌半乳糖凝集素3表达水平上调40~50倍，同时出现心房扩张、左心室（left ventricular, LV）射血分数降低、收缩期与舒张期左心室容积增加、心房传导延迟、严重心肌纤维化以及纤维化相关基因表达上调。在Mst1-TG小鼠中敲除半乳糖凝集素3基因，可减轻其收缩期与舒张期左心室容积增加、心房传导延迟及心肌纤维化进程。对Mst1-TG小鼠施以修饰柑橘果胶治疗，未对其扩张型心肌病表型产生获益，这可能与半乳糖凝集素3的高表达水平有关。 结论：本研究发现，呈现扩张型心肌病表型的Mst1-TG小鼠心脏中，半乳糖凝集素3的表达显著上调。敲除半乳糖凝集素3基因可有效抑制心肌纤维化信号通路与左心室扩张，并改善心功能。因此，抑制半乳糖凝集素3的表达可能成为扩张型心肌病的潜在治疗策略，开发新型半乳糖凝集素3抑制剂具有重要意义。此外，本研究对两种遗传背景下的对照组与半乳糖凝集素3敲除小鼠的左心室样本进行了RNA测序（RNA-seq）：分别为野生型C57BL/6小鼠，以及常用作扩张型心肌病模型的Mst1过表达小鼠。每组设置5~8个生物学重复样本进行测序处理。