Assessing myofilament phenotype and contractile response to pharmacotherapies in a human induced pluripotent stem cell cardiomyocyte (hiPSC-CM) model of hypoplastic left heart syndrome (HLHS)

项目摘要 尽管在手术技术和术后护理方面取得了巨大进步，但患有发育不良的儿童 左心综合征(HLHS)仍然是系统性右室(RV)收缩衰竭的终生风险。在……里面 HLHS患者，传统的成人心力衰竭药物在预防病理方面总体无效 心脏重塑或保留收缩功能。因此，一旦患有HLHS的儿童出现心力衰竭 症状学继发于心功能下降，心脏移植通常是唯一长期的 治疗选择。 这一建议的中心假设是HLHS心肌细胞在 可能导致心力衰竭倾向并改变收缩反应的肌丝水平 医用心力衰竭疗法。初步数据显示钙敏感性和 终末期高血压病患者心肌细胞肌钙蛋白I磷酸化水平降低 失败了。然而，有必要利用HLHS模式系统对这些研究进行扩展，以进一步 评估这些肌丝变化的影响。因此，这项建议的目标是发展一个人 诱导多能干细胞心肌细胞模型的建立 肌丝对HLHS存在的收缩异常的贡献并评估 肌丝对HF治疗的收缩反应的差异。特定患者的HLHS hiPSC-CM将是 来自3名患有终末期心力衰竭的HLHS患者，肌丝收缩表型将 通过对去皮HiPSC-CM的力学研究，肌丝蛋白亚型的定量， 以及对肌丝翻译后修饰的评估。这些发现将与 同一受试者移植的HLHS心脏组织中显示了肌丝表型。另外， 肌丝评估将在HLHS hiPSC-CM治疗后进行 心力衰竭的药物治疗，以直接衡量药物治疗对HLHS的影响 肌丝。 最终，加强对HLHS存在的收缩功能障碍的机械学理解 心肌细胞有可能通过告知准确的心力衰竭风险来改善HLHS患者的预后。 分层范式。此外，优先考虑HLHS对收缩反应的具体评估 药物治疗可能导致个性化的药理学策略，以保护全身RV功能。

Project Summary Despite tremendous advances in surgical technique and post-operative care, children with hypoplastic left heart syndrome (HLHS) remain at lifelong risk for systolic failure of the systemic right ventricle (RV). In HLHS patients, traditional adult heart failure medications have been overall ineffective in preventing pathologic cardiac remodeling or preserving contractile function. Thus, once children with HLHS develop heart failure symptomatology secondary to decreased cardiac function, heart transplantation is often the only long-term therapeutic option. The central hypothesis of this proposal is that HLHS cardiomyocytes have inherent alterations at the level of the myofilament that may confer a heart failure predisposition and alter the contractile response to medical heart failure therapies. Preliminary data demonstrates abnormalities in calcium sensitivity and decreased cardiac troponin I phosphorylation in cardiomyocytes from HLHS subjects with end-stage heart failure. Nevertheless, there is a need to expand upon these studies utilizing a HLHS model system to further evaluate the impact of these myofilament changes. Therefore, the goal of this proposal is to develop a human induced pluripotent stem cell cardiomyocytes (hiPSC-CM) model derived from HLHS subjects to define the myofilament contribution to the contractile abnormalities present in HLHS and assess the impact of myofilament differences on the contractile response to HF therapies. Patient-specific HLHS hiPSC-CM will be generated from 3 HLHS subjects with end-stage heart failure, and the myofilament contractile phenotype will be defined through mechanical studies of skinned hiPSC-CM, quantification of myofilament protein isoforms, and assessment of myofilament post-translational modifications. These findings will be paired with the myofilament phenotype demonstrated in the explanted, HLHS heart tissue from the same subject. Additionally, myofilament assessments will be performed following treatment of HLHS hiPSC-CM with current and emerging HF pharmacotherapies in order to directly measure the consequences of drug therapy on the HLHS myofilament. Ultimately, augmenting mechanistic understanding of the contractile dysfunction present in HLHS cardiomyocytes has the potential to improve outcomes for HLHS patients by informing an accurate HF risk- stratification paradigm. Furthermore, prioritizing an HLHS-specific appraisal of the contractile response to pharmacotherapies may result in personalized pharmacologic strategies to preserve systemic RV function.