The Regulatory Role Of Wnt/β-catenin Signaling Pathway On Scleral Remodeling In Experimentally-induced Myopia

Background: To investigate Wnt/β-catenin signaling pathway expression and its regulation of scleral remodeling in form-deprivation myopia(FDM)guinea pig model.Methods: Guinea pigs were used to develop FDM by wearing a translucent diffuser.Wnt isoforms were examined using genome microarrays.In vitro scleral fibroblasts were primary cultured and identified by vimentin and keratin.Wnt isoforms,β-catenin,TGF-β1,and type I collagen expression levels were examined with or without Wnt/β-catenin pathway antagonist DKK-1 or TGF-β1 neutralizing antibody by Western-blot,ELISA and immunofluorescence.Guinea pigs for the experiment in vivo were divided into three groups: control group,FDM group,DKK-1 intervention group and TGF-β1 neutralizing antibody intervention group.Axial length and myopic refraction errors were observed in the development process of FDM.The sclera ultra structure and collagen fibers were examined by transmission electron microscope(TEM).Wnt isoforms,β-catenin,and type I collagen expression levels were examined by western-blot.Results: In the third and fourth week,in the FDM group,axial length increased significantly and myopic degrees increased significantly compared with those in control group.For genome microarrays,the expression of Wnt3 in FDM group was significantly greater than that in control group as confirmed by Western-blot in retinal and scleral tissue.Primary scleral fibroblasts were spindle-shaped.Scleral cells cultured were positively stained for vimentin and negatively stained for keratin and PBS.Experiment in vitro: in the FDM group,the expressions of Wnt3 and β-catenin significantly increased,and TGF-β1 expression level decreased significantly.Along with morphological misalignment inside and outside cells,the amount of type I collagen decreased significantly.In FDM group with DKK-1 added,the expressions of Wnt3 and β-catenin significantly decreased,TGF-β1 and type I collagen expression level increased significantly,while type I collagen was sequentially distributed.In FDM group with TGF-β1 neutralizing antibody added,TGF-β1 and type I collagen expression level decreased,moreover,type I collagen rearranged more disorder.Experiment in vivo: in FDM group,the expression levels of Wnt3 and β-catenin in retina and sclera were increased significantly,collagen fibers in sclera were in disorder and scattered,and type I collagen decreased significantly in sclera tissue.In FDM group with DKK-1 injected,axial length was significantly shorter and myopic refraction errors decreased significantly compared with the FDM group without DKK-1.The expression levels of Wnt3 and β-catenin in retina reduced significantly.Collagen fibers were in order,and type I collagen increased significantly.There were no significant changes with TGF-β1 neutralizing antibody added.Conclusions: From experiments in vitro and in vivo,the study suggested that in guinea pig FDM model,the Wnt3/β-catenin signaling pathway could further regulate the expression and arrangement of type I collagen by the decrease of TGF-β1.This pathway regulated the remodeling of scleral matrix to affect the progress of myopia ultimately,which can be reversed by DKK-1.