Inhibitory effect of ursolic acid on high oxygen-induced mouse retinal neovascularization by intravitreal injection / 中华实验眼科杂志

ABSTRACT

Background Retinal neovascular diseases affect visual function.Although many drugs have been used to manage the visual diseases,their effectiveness is less than satisfactory.Studies showed that ursolic acid has multiple biological effects including anti-vascularization.However,the effect of ursolic acid on retinal neovascular diseases is unclear now.Objective This study was to observe the inhibitory effect of ursolic acid on the high oxygen-induced mouse retinal neovascularization after intravitreal injection.Methods Sixty clean 7-day-old C57BL/6J mice were divided into the blank control group,PBS control group,positive control group (triamcinolone) and low,moderate and high dose (1.5,3.0 and 6.0 μg) ursolic acid groups randomly.The blank control group mice were raised in normal environment,and the mice from other groups were fed in the environment with O2 concentration at (75±2)％ for 5 days together with the maternal mice.The mice then were back to the normal air environment to induce retinal neovascularization.Then,the drugs were intravitreally immediately injected in the mice of the different groups.The mice were sacrificed at the 17-day old for the preparation of retinal sections.Retinal new blood vessel was examined by haematoxylin and eosin stain under the light microscope,and the number of vascular endothelial cell nucleus breaking the inner limiting membrane was counted.The gene expressions of vascular endothelial growth factor (VEGF),cyclooxygenase-2 (COX-2) and matrix metalloproteinase-2 (MMP-2) in the mouse retinas were quantitatively assayed using reverse transcription PCR.Results The number of endothelial nuclei newly-generated vessel breaking internal limiting membrane in the mice of PBS control group was (18.65±3.24)/field,which was more than (0.78±0.11)/field of the blank control group obviously (t =2.24,P＜0.05).The number of endothelial nuclei newly-generated breaking internal limiting membrane in the moderate-or high-dose ursolic acid group was less than that of moderate group obviously,it was statistically significant(P＜0.05).The number of vascular endothelial cell nuclei breaking internal limiting membrane in high high-dose group was (13.32 ± 1.87)/field and (8.93 ± 1.09) /field,showing significant decreases in comparison with the PBS control group and low-dose ursolic acid group (18.65±3.24)/field (15.44±2.02)/field (all at P＜0.05).However,no significant difference were seen in the number of new vascular endothelial cell nucleus between the high-dose ursolic acid group and the positive control group(9.14±1.13)/field (t=1.17,P＞0.05).The relative expressions of COX-2 mRNA,VEGF mRNA and MMP-2 mRNA in the mouse retinas were higher in the PBS control group than those in the blank control group (t =13.45,12.49,14.32,all at P＜0.05),and those in the moderate-dose or high-dose ursolic acid group were lowed in comparison with the PBS control group and the low-dose ursolic acid group (all at P＜0.05),but there were no significant differences between the high-dose ursolic acid group and the positive control group (all at P＞0.05).Conclusions Ursolic acid can suppress retinal neovascularization by down-regulating the expressions of VEGF,COX-2 and MMP-2 in oxygen-induced retinopathy of mouse in dose-dependent manner.