Effect of rhHAPO on the growth of purified CD34⁺KDR⁺ cells. (A) Effect of rhHAPO on the growth pattern of CD34⁺KDR⁺ cells. CD34⁺KDR⁺ cells were cultured at 5 × 10³ cells per well in a 24-well plate. The number of viable cells was measured by counting trypan blue-negative cells. The log growth phase of cells was between day 3 and day 9, followed by the growth arrest phase when rhHAPO was added. Without rhHAPO, the number of CD34⁺KDR⁺ cells is decreased. (B) Hematopoietic and endothelial proliferation and differentiation of CD34⁺KDR⁺ cells in a liquid culture system. Original magnification × 200. In the absence of rhHAPO, a few cells grew in culture on day 7 (i). In contrast, significant cell proliferation with colony formation was observed in the presence of rhHAPO (ii). The suspended cells were aspirated away, and adherent cells were cultured in fresh medium with or without rhHAPO for 3 additional days. The cells grew well but displayed different morphology in the presence or absence of rhHAPO. In comparison with control culture (iii), the addition of rhHAPO into the culture significantly promoted the proliferation of endothelial-like cells (iv). When single CD34⁺KDR⁺ cells were cultured in 96-well plates in culture system C, as described in “Materials and methods,” for 2 weeks, these cells displayed different morphology in the presence or absence of rhHAPO. In comparison with control culture without rhHAPO (v), the addition of rhHAPO into the culture significantly promoted the proliferation of endothelial-like cells and formed capillary-like structures (vi).