Ou Qinghui, Abouleisa Riham R E, Tang Xian-Liang, Juhardeen Hamzah R, Meki Moustafa H, Miller Jessica M, Giridharan Guruprasad, El-Baz Ayman, Bolli Roberto, Mohamed Tamer M A
Institute of Molecular Cardiology, Department of Medicine, University of Louisville.
Department of Bioengineering, University of Louisville.
J Vis Exp. 2020 Mar 20(157). doi: 10.3791/60913.
Many novel drugs fail in clinical studies due to cardiotoxic side effects as the currently available in vitro assays and in vivo animal models poorly predict human cardiac liabilities, posing a multi-billion-dollar burden on the pharmaceutical industry. Hence, there is a worldwide unmet medical need for better approaches to identify drug cardiotoxicity before undertaking costly and time consuming 'first in man' trials. Currently, only immature cardiac cells (human induced pluripotent stem cell-derived cardiomyocytes [hiPSC-CMs]) are used to test therapeutic efficiency and drug toxicity as they are the only human cardiac cells that can be cultured for prolonged periods required to test drug efficacy and toxicity. However, a single cell type cannot replicate the phenotype of the complex 3D heart tissue which is formed of multiple cell types. Importantly, the effect of drugs needs to be tested on adult cardiomyocytes, which have different characteristics and toxicity responses compared to immature hiPSC-CMs. Culturing human heart slices is a promising model of intact human myocardium. This technology provides access to a complete multicellular system that mimics the human heart tissue and reflects the physiological or pathological conditions of the human myocardium. Recently, through optimization of the culture media components and the culture conditions to include continuous electrical stimulation at 1.2 Hz and intermittent oxygenation of the culture medium, we developed a new culture system setup that preserves viability and functionality of human and pig heart slices for 6 days in culture. In the current protocol, we are detailing the method for slicing and culturing pig heart as an example. The same protocol is used to culture slices from human, dog, sheep, or cat hearts. This culture system has the potential to become a powerful predictive human in situ model for acute cardiotoxicity testing that closes the gap between preclinical and clinical testing results.
许多新型药物在临床研究中失败，原因是心脏毒性副作用，因为目前可用的体外试验和体内动物模型难以预测人类心脏的不良反应，给制药行业带来了数十亿美元的负担。因此，在进行代价高昂且耗时的“首次人体试验”之前，全球迫切需要更好的方法来识别药物的心脏毒性。目前，只有未成熟的心脏细胞（人类诱导多能干细胞衍生的心肌细胞[hiPSC-CMs]）用于测试治疗效果和药物毒性，因为它们是唯一能够长时间培养以测试药物疗效和毒性的人类心脏细胞。然而，单一细胞类型无法复制由多种细胞类型组成的复杂三维心脏组织的表型。重要的是，药物的效果需要在成人心肌细胞上进行测试，与未成熟的hiPSC-CMs相比，成人心肌细胞具有不同的特征和毒性反应。培养人类心脏切片是一种很有前景的完整人类心肌模型。这项技术提供了一个完整的多细胞系统，该系统模拟人类心脏组织并反映人类心肌的生理或病理状况。最近，通过优化培养基成分和培养条件，包括以1.2Hz的频率进行连续电刺激和对培养基进行间歇性氧合，我们开发了一种新的培养系统设置，可在培养6天的时间内保持人类和猪心脏切片的活力和功能。在本方案中，我们将详细介绍以猪心脏切片和培养为例的方法。同样的方案用于培养来自人类、狗、羊或猫心脏的切片。这种培养系统有可能成为一种强大的预测急性心脏毒性的人体原位模型，弥合临床前和临床测试结果之间的差距。
