Fig. 2

JIB-04 inhibited HASMC viability and proliferation. A. Microscopic observation of HASMCs in 6-cm dishes after treatment with DMSO or JIB-04 (0.5 μM) for 0, 24, 48, and 72 h. Scale bar, 100 μm. B. Cell counts of HASMCs in 6-cm dishes after treatment with DMSO or JIB-04 (0.5 μM) for 72 h (n = 3 per group). C. Absorbance at 450 nm of HASMCs after treatment with DMSO or JIB-04 (0.5 μM) for 0, 24, 48, and 72 h (n = 5 per group). D-E. The EdU incorporation assay showed the proliferative potential in HASMCs after treatment with DMSO or JIB-04 (0.5 μM) for 24 h, in which nuclei were stained with DAPI (blue) and EdU incorporation appeared in red. The positive EdU rate was measured (n = 3 per group). Scale bar, 50 μm. F-G. Immunofluorescence staining of proliferating nuclear antigen Ki67 revealed the proliferative capacity of HASMCs after treatment with DMSO or JIB-04 (0.5 μM) for 24 h. Nuclei were stained with DAPI (blue), and Ki67 was stained red (n = 3 per group). Scale bar, 50 μm. H-I, Western blot showed the proliferation-related protein levels of PCNA and phosphorylated H3 (p-H3) in HASMCs after treatment with DMSO or JIB-04 (0.5 μM) for 48 h (n = 4 per group). β-Actin served as a loading control. *P < 0.05, **P < 0.01