Abstract 11789: Exosomes Released by Human Cardiac Fibroblasts Promote the Maturation of Human Induced-Pluripotent Stem Cells Derived Cardiomyocyte via AMPK Signal Pathway

Background: Human induced-pluripotent stem cell derived cardiomyocytes (hiPS-CMs) is phenotypically less matured than adult cardiomyocytes. The aim of this study was to explore the potential mechanism by which human cardiac fibroblast-derived exosomes (hc-FB-EXOs) enhance the maturation of hiPS-CMs. Method: Exosomes were extracted from human cardiac fibroblasts (hc-FBs) which were cultured with exosome-free FBS medium. The cultured hiPS-CMs were randomly devided into two groups: co-culture with (EXO) and without (NC) hc-FB-EXOs. The morphology of hiPS-CMs was assessed via immunostaining. Mitochondrial functions assay were measured by Seahorse XF. Potential mechanism was assessed via mRNA-Seq, real-time PCR and western blot. Results: Morphologically, the cells treated in EXO group was significantly larger than the in NC group in terms of cell area and sarcomere length (Panel A). In cellular metabolism, the mitochondrial oxygen consumption rate of EXO group was significantly higher than that of NC group, including basal respiration, proton leak, maximal respiration and spare respiratory capacity (Panel B). The expression of cardiac Troponin I (cTnI), Myosin Heavy Chain 7 (MYH7), Cyclin Dependent Kinase Inhibitor 2C (CDKN2C or p18), phospho-WEE1 and CAV3 (caveolin3) were significant higher, while the expression of of Myosin Heavy Chain 6 (MYH6) was significant lower in EXO group than in NC group (Panel C). Furthermore, qPCR showed that TNNI1, TNNT2, TNNI3/TNNI1, total titin, N2B, MYH7, LPL, ND1, N2BA, and MYH6 were significantly upregulated in EXO group compared with the NC group. Furthermore, western blot validation of RNA-Seq data confirmed that significantly upregulated of phospho-AMPK and downregulated of ACC1 and SCD1 were detected in EXO group compared with the NC group (Panel D). Conclusion: hc-FB-EXOs could significantly facilitate maturation of hiPS-CMs via the phosphorylation of AMPK.