Expression And Significance Of YAP In Rabbit Corneal Neovascularization Induced By Alkali Burn

Objective:To investigate the expression and significance of Yes-associated protein（Yes-associated protein,YAP）in rabbit corneal neovascularization induced by alkali burn,so as to provide a new exploration for the treatment of corneal neovascularization.Methods:A total of 17 healthy adult New Zealand white rabbits with 34eyes were selected.The rabbits were divided randomly into control group（without any treatment）with 2 rabbits with 4 eyes and experimental group（alkali burn model）with 15 rabbits with 30 eyes.In the experimental group,corneal neovascularrization（CNV）model induced by alkali burn was established in both eyes at the same time.The growth of CNV in rabbits was observed under microscope at 1 day,4 day,7 day,10 day and 14 days after alkali burn.Then the area and length of CNV were measured by image analysis software.After observation,rabbit corneal specimens were taken at each time point to make frozen sections,and the expression of vascular endothelial growth factor（Vascularendothelialgrowthfactor,VEGF）in corneal tissues was detected by immunofluorescence.Corneal tissue protein was extracted and the expression of YAP and VEGF was detected by Western blot.The mRNA of corneal tissue was extracted and the expression of YAP,VEGF,large tumor suppressor factor（Large tumor suppressor,LATS）and TAZ-tea domain family member（TEA domain family member,TEAD）was detected by real-time fluorescence quantitative PCR.Pearson correlation analysis was used to analyze the correlation between YAP and VEGF expression.Results:The length and area of corneal neovascularization in experimental group changed curvilinear with the increase of time after alkali burn.One day after alkali burn,slit lamp microscope showed that limbal vasodilation could be seen in the experimental group,and there was no significant difference in the length and area of CNV(P_length>0.05,P_area>0.05).On the 4th days after alkali burn,CNV grew along the limbus to the center of the cornea in the experimental group,and the length and area of CNV were significantly higher than those on the 1st day after alkali burn(P_length<0.05,P_area<0.05).On the 7th days after alkali burn,the length of CNV in experimental group were close to the edge of the alkali burn area,and the length and area of CNV were significantly higher than those on the 4th day after alkali burn(P_length<0.05,P_area<0.05).On the 10th days after alkali burn,the length of CNV in experimental group reached the edge of alkali burn area,and the length and area of CNV were significantly higher than those on the 7th day after alkali burn(P_length<0.05,P_area<0.05).On the 14th days after alkali burn,the length of CNV in the experimental group crossed the edge of the alkali burn area,and the length and area of CNV were significantly higher than those on the 10th day after alkali burn(P_length<0.05,P_area<0.05).Immunofluorescence detection showed that the expression of VEGF in CNV of the experimental group was higher than that of the control group,and the expression of VEGF in neovascularization was consistent with the area and length of CNV at each time point.In the experimental group,the expression of YAP and VEGF protein increased significantly after alkali burn,reached the peak on the 7th day,and then decreased gradually,and the expression trend of YAP was consistent with that of VEGF.Pearson correlation analysis was used to verify the correlation between YAP protein and VEGF protein expression.The expression of YAP and VEGF in 1 day,4 days,7 days,10 days and 14 days after alkali burn was significantly different from that in the control group（all P<0.05）.The expression of YAP,VEGF and TEAD mRNA was higher than that of the control group,and the expression trend was consistent with the growth trend of CNV,while the expression of LATS mRNA was lower than that of the control group,and the expression trend was opposite to that of CNV.Pearson correlation analysis was used to verify the correlation between the expression of YAP mRNA and VEGF mRNA.After alkali burn,the mRNA expressions of YAP,VEGF,TEAD and LATS at 1 day,4 days,7 days,10 days and 14 days were significantly different from those in the control group（all P<0.05）.Conclusion:Hippo/YAP signal pathway may be involved in the formation of rabbit corneal neovascularization induced by alkali burn and is related to the expression of VEGF.