Effects Of Mitochondria-endoplasmic Reticulum Membrane In A Rat Model Of Chronic Glaucoma

Purpose: To observe and confirm the morphological and functional changes of mitochondrial-associated endoplasmic reticulum membrane(MAM)and investigate the effects of mitofusin2(Mfn2)on retinal ganglion cells(RGCs)apoptosis in experimental glaucoma model of rats.Methods:1.In this study,the chronic elevation of intraocular pressure(IOP)was induced by circumlimbal suture in rats.To identify the successful establishment of the chronic glaucoma model,HE staining of retinas was applied to observe the pathological changes of retina and optic nerve;RGCs were specifically labeled with Brn3 b antibody and counted;and the expression of apoptotic protein caspase3 was detected by western blot(WB).2.Transmission electron microscope(TEM)was used to observe the morphological changes of MAM in chronic glaucoma model,and WB was applied to detect the expression of mitochondrial and endoplasmic reticulum-related proteins.3.Adeno-associated virus-mediated MFN2 gene was transferred to retina by vitreous injection to interfere with the expression of Mfn2 in the retina of rats.The morphological changes of retina,the morphology and function of mitochondria and the changes of endoplasmic reticulum stress-related proteins in each group were observed to explore the role and mechanism of Mfn2 in the pathogenesis of glaucoma.Results:1.Circumlimbal suture could induce chronic elevation of intraocular pressure in rats,accompanied by characteristic glaucomatous pathological changes including decrease of retinal thickness,optic nerve degeneration and RGC apoptosis shown by HE staining and RGC counting.2.In circumlimbal suture model,distance between mitochondria and endoplasmic reticulum increased;mitochondrial morphology changed from long shape to small and round;mitochondrial dynamics were disrupted;and unfolded protein response(UPR)was activated.3.In chronic glaucoma model,MFN2 overexpression could cause the increase of retinal thickness and RGC number,decrease the expression of p-PERK and p-eIF2α,and activate caspase 3.Conclusion: In chronic glaucoma model induced by circumlimbal suture,decrease of MAM-related protein Mfn2 can cause RGC apoptosis by changing the morphology and function of MAM,which present as disordered mitochondrial dynamics and activation of UPR.And overexpression Mfn2 of can recover the retinal morphology and inhibition the apoptosis of RGCs at some extent by inhibiting the activation of PERK pathway.