Objective: The study was to investigate the impact of cord blood CD(3)AK cell culture supernatant (CS) on proliferation, differentiation and apoptosis of HL-60 cells.
Methods: HL-60 cells were treated with different concentrations of CS (10%, 15%, 20%) for 3 days, 6 days and 9 days, and the same cells of control group were not treated with CS. The growth of induced cells was assessed with Trypan blue staining and cell counting with cytometer. The differentiation marker CD(11b) on the cell surface and cell-cycle was analyzed by flow cytometry (FCM), cell morphology (Wright-Giemsa staining) and NBT test to determine the extent of differentiation. Meanwhile, the changes of the apoptosis of the cells induced by 20% CS at different time points (3, 6 and 9 days) were analyzed by TUNNEL-POD, and the apoptotic characteristics of cells were observed.
Results: The growth of HL-60 cell was inhibited as CS-inducing time and the dose of CS increased. At the same time, but HL-60 cell number in G(0)/G(1) phase of cell-cycle increased, but HL-60 cell number in S phase decreased compared with untreated group. The HL-60 cells induced by 20% CS for 9 days showed that (52.7 +/- 1.8)% of cells were at G(0)+G(1) phase and (43.8 +/- 1.1)% were at S phase (P < 0.05), which demonstrated that HL-60 cells induced by 20% CS underwent G(0)/G(1) phase cell-cycle arrest. The volume of the differentiated cells was enlarged gradually as CS-inducing time prolonged. After 3 days the differentiating cells began to express differentiating marker CD(11b) on the cell surface and the nuclei morphology of the differentiated cells was also changed and NBT-stained cells increased in number with the increased dose of CS increased. Three days after induction by 20% CS, the induced cells began to show signs of apoptosis and the apoptotic percentage of induced cells gradually increased with CS-induction time. The rate of apoptosis of cells was (33.3 +/- 2.3)% at 9 days (P < 0.01).
Conclusion: CS could not only inhibit the growth of HL-60 cells but also induce the differentiation and apoptosis in HL-60 cells.