背景:除了观察到的细胞结构和线粒体的改变,在受desmin突变影响的患者中,将罕见的基因突变与心力衰竭发展联系起来的机制仍不清楚,部分原因是,缺乏相关的人类心肌细胞模型。
方法:为了阐明线粒体在这些机制中的作用,我们研究了来自人诱导性多能干细胞的心肌细胞,这些干细胞携带有杂合的DESE439K突变,这些干细胞要么从患者体内分离,要么通过基因编辑产生.为了增加生理相关性,心肌细胞要么在各向异性微图案化表面上培养,以获得细长和对齐的心肌细胞,或者作为心脏球体来创建微组织。此外,适用时,来自心肌细胞的结果与心脏活检证实的突然死亡的患者的同一家庭有DESE439K突变,和五个对照健康捐献者的死后心脏样本。
结果:杂合DESE439K突变导致心肌细胞的整体细胞结构发生巨大变化,包括细胞大小和形态。最重要的是,突变心肌细胞显示改变的线粒体结构,线粒体呼吸能力和代谢活动让人想起在患者心脏组织中观察到的缺陷。最后,挑战病理机制,我们将正常线粒体转移到突变心肌细胞内,并证明这种治疗能够恢复心肌细胞的线粒体和收缩功能.
结论:这项工作突出了DESE439K突变的有害作用,证明线粒体异常在结蛋白相关心肌病的病理生理学中的关键作用,并为这种疾病开辟了新的潜在治疗前景。
Beyond the observed alterations in cellular structure and mitochondria, the mechanisms linking rare genetic mutations to the development of heart failure in patients affected by desmin mutations remain unclear due in part, to the lack of relevant human cardiomyocyte models.
To shed light on the role of mitochondria in these mechanisms, we investigated cardiomyocytes derived from human induced pluripotent stem cells carrying the heterozygous DESE439K mutation that were either isolated from a patient or generated by gene editing. To increase physiological relevance, cardiomyocytes were either cultured on an anisotropic micropatterned surface to obtain elongated and aligned cardiomyocytes, or as a cardiac spheroid to create a micro-tissue. Moreover, when applicable, results from cardiomyocytes were confirmed with heart biopsies of suddenly died patient of the same family harboring DESE439K mutation, and post-mortem heart samples from five control healthy donors.
The heterozygous DESE439K mutation leads to dramatic changes in the overall cytoarchitecture of cardiomyocytes, including cell size and morphology. Most importantly, mutant cardiomyocytes display altered mitochondrial architecture, mitochondrial respiratory capacity and metabolic activity reminiscent of defects observed in patient\'s heart tissue. Finally, to challenge the pathological mechanism, we transferred normal mitochondria inside the mutant cardiomyocytes and demonstrated that this treatment was able to restore mitochondrial and contractile functions of cardiomyocytes.
This work highlights the deleterious effects of DESE439K mutation, demonstrates the crucial role of mitochondrial abnormalities in the pathophysiology of desmin-related cardiomyopathy, and opens up new potential therapeutic perspectives for this disease.