Obscurin participates in the development of striated muscles and maintenance of the functional sarcoplasmic reticulum. However, the role of obscurin in arrhythmogenic right ventricular cardiomyopathy (ARVC) is not well understood. We aimed to study the novel obscurin mutations in the pathogenesis of ARVC and the underlying mechanisms./nMethods: We generated induced pluripotent stem cells (iPSC) through retroviral reprogramming of peripheral blood mononuclear cells isolated from a 46-year-old female diagnosed with ARVC, carrying a mutation in OBSCN. The cells differentiated into functional iPSC-based cardiomyocytes (iPSC-CMs), whose phenotype was determined by transmission electron microscopy, electrophysiological description, immunofluorescence staining, and Oil Red O staining. Molecular characterization was performed by bioinformatic analyses, and identification by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting./nResults: ARVC-iPSC-CMs mutation in OBSCN showed significant accumulation of lipids, increased pleomorphism, irregular Z-bands, and increased L type calcium currents. Functional enrichment analysis identified pathways involved in focal adhesion and structure formation; the adipocytokines and PPAR signaling pathways were also activated in the ARVC group. Moreover, our results from ultra-high-resolution microscopy, qRT-PCR and Western blotting confirmed that the mutant OBSCN protein and its anchor protein, Ank1.5, showed structural disorder and decreased expression, but there was increased expression of junctional protein N-Cadherin. Further analysis revealed the gene expression of other desmosomal proteins in ARVC-iPSC-CMs was also decreased but some adipogenesis pathway-related proteins (PPARγ, C/EBPα, and FABP4) were increased./nConclusion: A novel frameshift mutation in OBSCN caused phenotypic alteration accompanied by disrupted localization and decreased expression of its anchoring protein Ank1.5. Furthermore, there was an accumulation of lipids with an increase in fatty fibrosis area and myocardial structural disorder, possibly leading to dysrhythmia in calcium channel-related myocardial contraction. These observations suggested the possibility of attenuating ARVC progression by therapeutic modulation of OBSCN expression.

中文翻译：

---

心律失常性右心室心肌病中基于OBSCN突变体iPSC的心肌细胞的细胞内钙电流异常和疾病表型

Obscurin参与横纹肌的发育和功能性肌浆网的维持。然而，关于obscurin在致心律失常性右心室心肌病（ARVC）中的作用尚不清楚。我们的目的是研究ARVC发病机理中新的obscurin突变及其潜在机制。/n方法：通过逆转录病毒重编程，从46岁诊断为女性的女性外周血中分离出外周血单核细胞，从而产生诱导性多能干细胞（iPSC）。 ARVC，在OBSCN中携带突变。这些细胞分化为功能性的基于iPSC的心肌细胞（iPSC-CM），其表型通过透射电子显微镜，电生理学描述，免疫荧光染色和油红O染色确定。通过生物信息学分析进行分子表征，并通过定量实时聚合酶链反应（qRT-PCR）和Western印迹进行鉴定。/n结果：OBSCN中的ARVC-iPSC-CMs突变表现出大量脂质堆积，多态性增加，Z形带不规则以及L型钙电流增加。功能富集分析确定了参与粘着斑和结构形成的途径；ARVC组的脂肪细胞因子和PPAR信号通路也被激活。此外，我们从超高分辨率显微镜，qRT-PCR和Western印迹获得的结果证实，突变型OBSCN蛋白及其锚蛋白Ank1.5表现出结构紊乱并表达降低，但连接蛋白N-的表达增加钙黏着蛋白。进一步的分析表明，ARVC-iPSC-CMs中其他桥粒蛋白的基因表达也降低了，但一些与脂肪形成途径相关的蛋白（PPARγ，C /EBPα和FABP4）却增加了。OBSCN中的新型移码突变会导致表型改变，并伴随定位异常和锚蛋白Ank1.5表达降低。此外，存在脂质堆积，脂肪纤维化面积增加和心肌结构紊乱，可能导致钙通道相关的心肌收缩性节律失常。这些观察结果提示通过治疗性调节OBSCN表达来减慢ARVC进展的可能性。