Biologic effects of different concentrations of putrescine on human umbilical vein endothelial cells

Objective To explore the effects of different concentrations of putrescine on proliferation, migration, and apoptosis of human umbilical vein endothelial cells (HUVECs). Methods HUVECs were routinely cultured in vitro. The 3rd to the 5th passage of HUVECs were used in the following experiments. (1) Cells were divided into 500, 1 000, and 5 000 μg/mL putrescine groups according to the random number table (the same grouping method was used for following grouping), with 3 wells in each group, which were respectively cultured with complete culture solution containing putrescine in the corresponding concentration for 24 h. Morphology of cells was observed by inverted optical microscope. (2) Cells were divided into 0.5, 1.0, 5.0, 10.0, 50.0, 100.0, 500.0, 1 000.0 μg/mL putrescine groups, and control group, with 4 wells in each group. Cells in the putrescine groups were respectively cultured with complete culture solution containing putrescine in the corresponding concentration for 24 h, and cells in control group were cultured with complete culture solution with no additional putrescine for 24 h. Cell proliferation activity (denoted as absorption value) was measured by colorimetry. (3) Cells were divided (with one well in each group) and cultured as in experiment (2), and the migration ability was detected by transwell migration assay. (4) Cells were divided (with one flask in each group) and cultured as in experiment (2), and the cell apoptosis rate was determined by flow cytometer. Data were processed with one-way analysis of variance, Kruskal-Wallis test, and Dunnett test. Results (1) After 24-h culture, cell attachment was good in 500 μg/mL putrescine group, and no obvious change in the shape was observed; cell attachment was less in 1 000 μg/mL putrescine group and the cells were small and rounded; cells in 5 000 μg/mL putrescine group were in fragmentation without attachment. (2) The absorption values of cells in 0.5, 1.0, 5.0, 10.0, 50.0, 100.0, 500.0, 1 000.0 μg/mL putrescine groups, and control group were respectively 0.588±0.055, 0.857±0.031, 0.707±0.031, 0.662±0.023, 0.450±0.019, 0.415±0.014, 0.359±0.020, 0.204±0.030, and 0.447±0.021, with statistically significant differences among them (χ²=6.86, P=0.009). The cell proliferation activity in 0.5, 1.0, 5.0, and 10.0 μg/mL putrescine groups was higher than that in control group (P<0.05 or P<0.01). The cell proliferation activity in 500.0 and 1 000.0 μg/mL putrescine groups was lower than that in control group (with P values below 0.01). The cell proliferation activity in 50.0 and 100.0 μg/mL putrescine groups was close to that in control group (with P values above 0.05). (3) There were statistically significant differences in the numbers of migrated cells between the putrescine groups and control group (F=138.662, P<0.001). The number of migrated cells was more in 1.0, 5.0, and 10.0 μg/mL putrescine groups than in control group (with P value below 0.01). The number of migrated cells was less in 500.0 and 1 000.0 μg/mL putrescine groups than in control group (with P value below 0.01). The number of migrated cells in 0.5, 50.0, and 100.0 μg/mL putrescine groups was close to that in control group (with P values above 0.05). (4) There were statistically significant differences in the apoptosis rate between the putrescine groups and control group (χ²=3.971, P=0.046). The cell apoptosis rate was lower in 0.5, 1.0, 5.0, and 10.0 μg/mL putrescine groups than in control group (with P values below 0.05). The cell apoptosis rate was higher in 500.0 and 1 000.0 μg/mL putrescine groups than in control group (with P values below 0.01). The cell apoptosis rates in 50.0 and 100.0 μg/mL putrescine groups were close to the cell apoptosis rate in control group (with P values above 0.05). Conclusions Low concentration of putrescine can remarkably enhance the ability of proliferation and migration of HUVECs, while a high concentration of putrescine can obviously inhibit HUVECs proliferation and migration, and it induces apoptosis.