Comparison of gelatinolytic activities and gene expression of MMP-2 and MMP-9 in CD34⁺ cells obtained from various sources. (A) Zymogram of media conditioned by CD34⁺ cells from steady-state bone marrow (BM, lane 1), steady-state peripheral blood (PB, lane 2), G-CSF–mobilized PB (lane 3), G-CSF plus chemotherapy-mobilized PB (lane 4), and CD34⁻ MNC from PB (lane 5) and BM (lane 6). The cells were incubated in serum-free IMDM at 37°C and 5% CO₂for 24 hours and the cell-conditioned media were electrophoresed in 10% acrylamide containing 2 mg/mL gelatin. The data presented here are representative of 12 BM, 2 unmobilized PB, 3 G-CSF–mobilized PB, and 12 G-CSF plus chemotherapy-mobilized PB experiments. (B) Effect of the MMP inhibitor o-phenanthroline on the expression of gelatinases by PB CD34⁺ cells. Media conditioned by KG-1 cells, known to secrete MMP-9 and MMP-2, was used as the standard (Std). The gels were incubated in the absence (left gel) and in the presence of 1.0 mmol/L o-phenanthroline (right gel) after electrophoresis. (C) RT-PCR analysis of MMP-9 and MMP-2 mRNA transcripts expressed by steady-state BM (lane 1) and PB (lane 2) CD34⁺ cells, G-CSF–mobilized PB CD34⁺ cells (lane 3) and PB MNC (C, lane 4). PCR products were electrophoresed on 2% agarose gels containing 0.1 g/mL ethidium bromide. GAPDH was used as the mRNA internal control to ensure equivalence of loading.