VEGF Contributes to Mesenchymal Stem Cell-Mediated Reversion of Nor1-Dependent Hypertrophy in iPS Cell-Derived Cardiomyocytes

Figure 6

MSC-derived VEGF is essential for hypertrophy regression in iPS-CM. (a) VEGF was quantified in culture supernatants of MSCs stimulated with 30 ng/ml IFN-γ and 3 ng/ml IL-1β for 24 h and those cultured for additional 24 h in serum-free medium (48 h). In addition, VEGF levels in fractionated MSC-conditioned medium (concentrates and filtrates) were determined by ELISA. . (b) PE-treated hypertrophied iPS-CM were cultured in the presence of 20% MSC-conditioned medium (CM) for 24 h. As a control, cells were cultured in medium supplemented with >30 kDa concentrates. For VEGF neutralization, 50 ng/ml or 150 ng/ml neutralizing anti-VEGF antibodies was added to the culture medium. The cell surface area was quantified by F-actin staining. Representative images are displayed. Bar: 100 μm. . (c) Nor1 gene and protein expression was quantified in iPS-CM cultured in the presence of MSC-CM supplemented with VEGF-neutralizing antibodies. . (d) Intracellular VEGF levels were quantified in hypertrophied iPS-CM cultured in the presence of 20% CM for 24 h. MSCs with and without preconditioning served as a control. . (e) VEGF levels were quantified in culture supernatants of PE-treated iPS-CM cultured in medium supplemented with 20% CM for 24 h. . , , and .

(a)

(b)

(c)

(d)

(e)