hBMSC transplantation augments myocardial cell proliferation, reduces myocardial apoptosis, and affects multiple paracrine factors. (A–F) BrdU imunohistochemistry. (A–D) Numerous BrdU-positive cells (green fluorescence) were observed in both host myocardial cells and transplanted cells in hBMSC-transplanted hearts (D), in contrast to TBMC-treated (E) or PBS-treated (F) hearts. (G–J) Double staining for BrdU (H) and ILB4 (I) shows proliferating capillary ECs (arrows). (K–N) Double staining for BrdU (L) and cTnI (M) shows proliferating cells (arrows) (N) in hBMSC-transplanted hearts. (O–Q) hBMSC-treated (O), TBMC-treated (P), or PBS-treated (Q) myocardium with anti–α-sarcomeric actinin mAb and TUNEL to identify apoptotic myocardial cells (green fluorescence). Fewer apoptotic cells are evident in hBMSC-transplanted hearts compared with those receiving TBMCs or PBS. (R and S) Staining with mAb against α-sarcomeric actinin (red) and TUNEL (green) demonstrates CMC apoptosis (arrows) in PBS-injected heart tissue. (T and U) Staining with ILB4 (red) and TUNEL (green) demonstrates EC apoptosis (arrows) in the PBS-injected heart. Scale bar: 100 μm (A–F and O–U), 50 μm (G–N). (V–Y) Heart samples (n = 3 per group) were harvested 4 weeks after MI and cell transplantation. Quantification of protein expression normalized to tubulin expression (V) demonstrates significant upregulation of VEGF, bFGF, and HGF in hBMSC-transplanted hearts compared with TBMC- or PBS-injected hearts. mRNA expression of angiogenic cytokines (W), cardiac transcription factors (X), and other factors (IGF and SDF1α) (Y) normalized to GAPDH expression shows significant upregulation of all factors in the hBMSC-transplanted hearts. Data were obtained from 3 separate experiments and are presented as amount relative to controls. *P < 0.05, **P < 0.01 vs. PBS; ⁺P < 0.01, ⁺⁺P < 0.001, ^#P < 0.0001 vs. TBMC and PBS. Ang, angiopoietin.