Induction of Pluripotent Stem Cell-Derived Cardiomyocyte Toxicity by Supernatant of Long Term-Stored Red Blood Cells in Vitro

Abstract

Background/Aims: Preserved red blood cells (RBCs) in vitro undergo a series of morphological, functional and metabolic changes during storage. RBC metabolites accumulate over time during storage, the toxicity of the supernatants of RBCs (SSRBCs) on tissue cells is largely unknown. Here, we aimed to study cardiomyocyte toxicity by supernatant of long term-stored RBCs in vitro and to discover elements involved in the mechanism. Methods: Using human-induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) and real-time cell analyzing (RTCA), we analyzed the cardiotoxicity of d0, d14 and d35 SSRBCs. To analyze the cardiotoxicity of potassium (K) and lactic acid (LA) in SSRBCs, solutions containing the same concentrations of K and LA were respectively prepared and co-cultured with hiPS-CMs. Immunofluorescence and Gene Expression Array of hiPS-CMs were performed to evaluate the effects of d35 K and d35 SSRBCs. Results: The beating of hiPS-CM was stopped by d14, d35 SSRBCs, or d35 K solution. Beating resumed within 48 hours in the presence of d14 SSRBC or d35 K but not d35 SSRBC; d0, d14 and d35 LA solution had no effect on beating patterns. At 48h after treatment, the immunofluorescence results showed that the integrity of the filament and sarcomere were intact. Gene Expression Array results found 14 differentially expressed genes which were likely to play an important role in the cytotoxic effect. Conclusion: Our results demonstrated cardiomyocyte toxicity by long term-stored SSRBCs in vitro. Besides high K-induced cardiotoxicity, there must be other unknown components in long term-stored SSRBCs that are cytotoxic to hiPS-CMs.

Keywords

• Cardiotoxicity
• Human-induced pluripotent stem cell-derived cardiomyocytes
• Suspended red blood cells
• Real-time cell analyzing