Figure 7
From: Cardiac Pacemaker Cells Generate Cardiomyocytes from Fibroblasts in Long-Term Cultures
Changes in the properties of purified GCFs in various culture environments. (A) Expression of desmin in GCFs co-cultured with SANCs without cell–cell contact (CC/I); desmin was not detected when GCFs were monocultured without SANCs in normal medium (NM); cTnT expression was not detected under either condition. Oxytocin (100 nM, O), 5-azacytidine (10 µM, 5-A) and trichostatin A (10 pg/mL, T) failed to induce desmin expression in monocultured GCFs even after 3 weeks culture. A, atrium. Each lane contained a total of 50 µg of protein. Full-length blots are given in Supplementary Fig. S10. (B) Typical phase-contrast images of GCFs cultured in normal medium (NM) (a) or co-cultured with SANCs in the same dish, but with physical isolation by cellulose membranes with 1-µm pores (co-culture with isolation, CC/I) (b) for 3 weeks; C, Immunocytochemical detection of cTnT protein (red) in EGFP-labelled (green) GCFs cultured under various conditions for 1 week; a) co-cultures with pre-fixed SANCs in SANC-conditioned medium; cTnT was detected only in pre-fixed SANCs (arrows). (b,c) Culture with repetitive electrical field stimulation in normal (b) and SANC-conditioned (c) medium; (d) the same condition as in (c) but with pre-fixed SANCs (arrow). bars, 50 µm.
