Effects Of Sunitinib On Proliferation And Migration Of Cultured Rhesus Macaque Choroid-Retinal Endothelial Cells In Vitro

Objective To evaluate the effects of sunitinib on proliferation and migration, and the effect of sunitinib on the expression of kinase insert domain-containing receptor (KDR/VEGFR-2) mRNA of cultured RF/6A cells (Rhesus macaque choroid-retinal endothelial cells).Methods The cultured cells were treated with different doses of sunitinib for 24 h and 48 h respectively. SRB (Sulforhodamine B) assay was used to examine the effect of sunitinib on cellular proliferation of RF/6A, concentrations of sunitinib used in the assay were 0 mg·L-1,0.00125 mg·L-1,0.0025 mg·L-1,0.005 mg·L-1,0.01 mg·L-1,0.02 mg·L-1 and 0.04 mg·L-1 respectively. Wound-healing assay was used to assess the effect of sunitinib on cell migration of RF/6A cells, concentrations of sunitinib used in the assay were 0.0025 mg·L-1,0.005 mg·L-1,0.01 mg·L-1 and 0.02 mg·L-1 respectively. The KDRmRNA on RF/6A was measured by Reverse Transcriptase-Polymerase chain reaction (RT-PCR), concentrations of sunitinib used in the assay were 0.0025 mg·L-1,0.005 mg·L-1,0.01 mg·L-1 and 0.02 mg·L-1 respectively. SPSS13.0 package was used for statistical analysis. Values are given in mean± SD. Statistical significance was evaluated using the one-way ANOVA followed by LSD-test for multiple comparisons. Differences at P<0.05 were considered to be statistically significant.Results 1.The results of SRB assay for the effects of sunitinib on the proliferation of RF/6A cells vere investigated by showed as follows:Sunitinib can inhibit the proliferation of RF/6A cells. The inhibitory effect was increased in both time-dependent and dose-dependent manner. Compared with that of group without sunitinib, sunitinib of all concentrations could inhibit the proliferation of RF/6A cells in both time-dependent and dose-dependent manner, the inhibitory rate were (12.009±0.038)%,(21.440±0.007)%,(35.434±0.015)%,(43.125±0.002)%,(53.700±0.001)% and (60.971±0.003)%, respectively at 24 h of cells cultivation (P<0.01).While at 48 h of cells cultivation the inhibitory rate were(36.872±0.006)%,(40.673±0.013)%,(47.313±0.004)%,(55.910±0.003)%,(63.120±0.003)% and (69.975±0.014)%, respectively (P<0.01).2. The results of wound-healing assay showed as follows:The migration distance at 24 h in cultured medium with 0 mg·L-1,0.0025 mg·L-1,0.005 mg·L-1,0.01 mg·L-1 and 0.02 mg·L-1 sunitinib were (203.3±2.2)μm,(145.4±4.4)μm,(123.9±2.6)μm,(96.1±3.1)μm and (46.6±2.9)μm, respectively. While at 48 h, the migration distance were (313.1±4.1)μm,(213.9±2.8)μm,(193.9±4.2)μm,(134.5±3.2)μm and (109.9±5.7)μm respectively. After the cells treated with sunitinib for 24 h and 48 h, the migration distance of RF/6A in groups with sunitinib were significantly shorter than that in group without sunitinib, and the distances decreased gradually along with the increase of sunitinib concentrations (P< 0.01).3. The RT-PCR results for the effects of sunitinib on KDRmRNA expression were showed as follows:The expression of KDRmRNA at 24 h in cultured medium with 0 mg·L-1,0.0025 mg·L-1,0.005 mg·L-1,0.01 mg·L-1 and 0.02 mg·L-1 sunitinib were (0.583±0.004),(0.570±0.008),(0.553±0.007),(0.531±0.003) and (0.513±0.005), respectively. While at 48h, the expression of KDRmRNA were (0.628±0.005),(0.610±0.002),(0.588±0.002),(0.564±0.005) and (0.525±0.004), respectively. After the cells treated with sunitinib for 24 h and 48 h, the expression of KDRmRNA in groups with sunitinib were significantly lower than that in group without sunitinib, and the expression of KDRmRNA decreased gradually along with the increase of sunitinib concentrations (P<0.01).Conelusions 1 These results suggested that sunitinib can inhibit the proliferation and the migration of RF/6A cells. And sunitinib can inhibit the expression of VEGFR mRNA.2 The effects of growth inhibition of proliferation and migration of RF/6A cells maybe associated with down-regulating the expression of the receptor of vascular endothelial growth factor (VEGFR).