Effect of chondroitin sulfate on the proliferation and differentiation of HL60 cells / 中国组织工程研究

ABSTRACT

BACKGROUND: Chondroitin sulfate is the important component of cell matrix, it can accelerate the proliferation of tumor cells and restrain its ransfer.OBJECTIVE: To observe the effects of chondroitin sulfate on the proliferation and differentiation of HL60 cells under the action of adriamycin.DESIGN: An open experiment.SETTING: Department of Biochemistry and Molecular Biology, Medical College of Qingdao University.MATERIALS: The experiments were carried out in the Research Room of Biochemistry and Molecular Biology, Medical College of Qingdao University of September 2003 to December 2004. Experimental materials and reagents: HL60 cell strains, which were the cells from promylocytic leukemia, were purchased from Shanghai Cell Bank, Chinese Academy of Medical Sciences; Bovine cartilage chondroitin sulfate (Sigma) was also used.METHODS: ① After the passage and culture, the cells at the logarithmic proliferative phase were dispensed into cell suspension of 1×108 L-1 with RPMI1640 culture medium containing inactivated fetal bovine serum of 0.1in volume fraction, and then filled into the culture bottles with 4 mL in each bottle for a total of 45 bottles. ② Chondroitin sulfate was added to 15 bottles filled with cell suspension according to the concentrations of 0, 5,25, 50 and 75 mg/L respectively, and 3 bottles for each concentration, and 0.01 mol/L phosphate buffer (pH7.2) was added in the blank control group. Then the density of HL60 cells was determined by cell counting after treatment of chondroitin sulfate. ③ Thirty bottles filled with cell suspension were divided into chondroitin sulfate+adriamycin group and chondroitin sulfate group, 15 bottles in each group. Chondroitin sulfate of 0, 5,25, 50 and 75 mg/L was added to the two groups, and 3 bottles for each concentration, and 0.01 mol/L phosphate buffer (pH7.2) wass adokd in the blank control group. Then the survival rate of chondroitin sulfate treated HL60 was detected after adding adriamycin. ④ Chondroitin sulfate was added to 15 bottles filled with cell suspension according to the concentrations of 0, 5, 25, 50 and 75 mg/L respectively, and 3 bottles for each concentration, 0.01 mol/L phosphate buffer (pH7.2) was added in the blank control group. The activity of acid phosphatase was detected with enzymelinked immunosorbant assay (ELISA) in each group, and the effect of the cell differentiation was observed.MAIN OUTCOME MEASURES: ① Effects of chondroitin sulfate on the proliferation of HL60 cells; ② Effects of chondroitin sulfate plus adriamycin on the survival rate of HL60 cells; ③ Effect of chondroitin sulfate on the activity of acid phosphatase of HL60 cells.RESULTS: Totally 45 bottles of cell suspension were prepared, and all were involved in the analysis of results. ① As compared with the blank control group, the densities of HL60 cells at 24 hours after treated with chondroitin sulfate of different concentrations were all significantly increased (P ＜ 0.01), which were increased more obviously in the 50 and 75 mg/L chondroitin sulfate treated groups, and there was no significant difference between the two groups (P ＞ 0.05), which indicated that chondroitin sulfate within the range of concentration did not accelerate the growth of cells greatly. ② As compared with the blank control group, the survival rates of HL60 cells in the chondroitin sulfate+adriamycin groups were decreased to different extents after chondroitin sulfate of different concentrations were added, and it decreased obviously when the concentration of chondroitin sulfate was higher than 25 mg/L (P ＜ 0.01). ③ As compared with the blank control group, the A values of acid phosphatase of the HHL60 cells were all obviously increased in the 5, 25 and 50 mL chondroitin sulfate treated groups (1.268±0.038, 1.305±0.101, 1.321±0.021,1.354±0.013, P ＜ 0.01 or 0.05), especially that it reached 1.406±0.113 in the 75 mL chondroitin sulfate treated group, which was extremely and significantly different from that in the blank control group (P ＜ 0.001).CONCLUSION: Chondroitin sulfate with a proper concentration can accelerate the proliferation of HL60 cells, and it can increase the sensibility of HL60 cells to adriamycin, and promote the differentiation of HL-60 cells.