High-throughput screening of drugs with human induced pluripotent stem cell–derived cardiomyocytes reveals a “cardiac safety index.”

Background: Cell-based regeneration therapies have great potential for application in new areas in clinical medicine, although some obstacles still remain to be overcome for a wide range of clinical applications. One major impediment is the difficulty in large-scale production of cells of interest with reproducibility. Current protocols of cell therapy require a time-consuming and laborious manual process. To solve this problem, we focused on the robotics of an automated and high-throughput cell culture system. Automated robotic cultivation of stem or progenitor cells in clinical trials has not been reported till date. The system AutoCulture® used in this study can automatically replace the culture medium, centrifuge cells, split cells, and take photographs for morphological assessment. We examined the feasibility of this system in a clinical setting. Results: We observed similar characteristics by both the culture methods in terms of the growth rate, gene expression profile, cell surface profile by fluorescence-activated cell sorting, surface glycan profile, and genomic DNA stability. These results indicate that AutoCulture® is a feasible method for the cultivation of human cells for regenerative medicine. Conclusions: An automated cell-processing machine will play important roles in cell therapy and have widespread use from application in multicenter trials to provision of off-the-shelf cell products.

Aims: Doxorubicin (DOX) is one of the most effective chemotherapeutic agents for various types of cancers. However, DOX often causes cardiotoxicity, which is referred to as DOX-induced cardiomyopathy (DIC). Despite extensive research, only a limited number of effective treatments are currently available. In this study, we aimed to identify a potential therapeutic target for DIC by preventing DOX-induced cell injury in cardiomyocytes. Methods and results: We performed a kinome-wide CRISPR gene knockout screen in human cardiomyocytes derived from pluripotent stem cells (hPSC-CMs) and identified a member of the STE20 kinase family, thousand and one amino acid protein kinase 1 (TAOK1) as a potential regulator of DOX-induced cardiomyocyte death. Using CRISPR-mediated gene knockout and small interfering RNA-mediated gene knockdown, we demonstrated that TAOK1 suppression improved DOX-induced cardiomyocyte death and dysfunction, including sarcomere disarray, contractile dysfunction, DNA damage, and mitochondrial dysfunction in hPSC-CMs. Transcriptome analysis using RNA-seq also showed that DOX-induced mitochondrial dysfunction was attenuated by TAOK1 suppression. In contrast to the protective role of TAOK1 against DOX toxicity in cardiomyocytes, TAOK1 suppression did not induce DOX resistance in human cancer cell lines. DOX-induced activation of p38 mitogen-activated protein kinase (MAPK) was markedly attenuated in TAOK1-knockout hPSC-CMs. Furthermore, DOX-induced cardiomyocyte death and disruption of mitochondrial membrane potential were augmented by TAOK1 overexpression, which was partially attenuated by an inhibitor or knockdown of p38 MAPK or an apoptosis inhibitor. Finally, we demonstrated that TAOK1 suppression using adeno-associated virus (AAV)-mediated gene silencing attenuated DOX-induced myocardial damage, including myocardial fibrosis, apoptosis, and cardiomyocyte atrophy, resulting in improved cardiac function in a mouse model of DIC. Conclusion: Our results indicate that TAOK1 suppression is a promising therapeutic approach for treating DIC in patients with cancer and highlight the advantages of hPSC-CMs as a platform to study drug-induced cardiotoxicity.

Cell-based therapies hold great promise for the treatment of peripheral arterial disease (PAD), especially in patients presenting with severe limb ischemia, although the optimal strategy remains to be explored. In this study, we evaluated the therapeutic effect of intravenous administration of human Muse cells, a unique subpopulation of mesenchymal stem cells (MSC), using a mouse model of hindlimb ischemia (HLI) without an immunosuppressant. Compared with the phosphate buffered saline (PBS) or non-Muse MSC groups, the Muse group showed significantly higher laser doppler blood flow in the ischemic limb at days 7 and 14 after HLI. Increased microvascular density [percent area of CD31(+) cells] and reduced interstitial fibrosis in the ischemic limb muscle were also observed in the Muse group. mCherry-expressing Muse cells were found in the ischemic border zone and expressed CD31 but did not in the non-ischemic limb. Muse cells produced higher amounts of vascular endothelial growth factor (VEGF) than non-Muse cells under normoxic and hypoxic conditions in vitro. In the ischemic muscle, tissue VEGF concentration and angiogenesis-related genes such as Vegfa, Angpt1, Pdgfb, and Igf1 were significantly higher in the Muse group than in the other two groups. In addition, the proportion of M2 macrophages to total macrophages and the ratio of anti-inflammatory-related genes such as IL-10, Arg1, and CD206 per iNOS were significantly higher in the Muse group than in the other two groups. In summary, Muse cells exert pleiotropic effects in a mouse model of HLI, and therefore may provide a novel therapeutic approach for the treatment of PAD patients with severe limb ischemia.