Study On Expression And Function Of Nogo-A And NgR In The Retina And Optic Nerve In Rat Model With Acute And Chronic Ocular Hypertension

BackgroundThe pathological mechanism of glaucomatous optic neuropathy is progressive death of retina ganglion cells and optic nerve fiber degeneration,which leads to irreversible damage.The regeneration of damaged central nervous system,imcluding optic nerve, was difficulty achieved since series of factors,such as inhibitors of axonal regeneration which are present in myelin,a lack of neurotrophic factors and formation of the glia scar. Some studies have showed that the regeneration of central nervous system was influenced by myelin associated protein Nogo-A,which functioned by connecting with its receptor-complex NgR/p75/Lingo-1.Thus,finding the expression of Nogo-A and NgR in the retina and optical nerve of normal and glaucomatous models would make a new way of the rehabilitation of glaucomatous optic nerve.Objective1.To investigate the expression and distribution of Nogo-A and its receptor NgR in the retina and optic nerve of normal rats.2.To evaluate the expressive variation of Nogo-A and NgR in the retina and optic nerve in the retina and optic nerve in rat model with acute and chronic ocular hypertension.3.To explore the effect in promoting axon regeneration of optic nerve by intravitreal injection siRNA knocking down the NgR protein expression in rat with chronic ocular hypertension.Materials and Methods1.Immunohistochemistry technique was used to assay the expression and distribution of Nogo-A and its receptor NgR in the retina and optic nerve.2.Establish rat model with acute and chronic ocular hyperrention.3.Reverse-transcription-polymerase chain reaction(RT-PCR)and Western Blot methods were used to evaluate the expressive varieties of Nogo-A and NgR in the retina and optic nerve in the retina and optic nerve in rat model with acute and chronic ocular hypertension.4.The NgR mRNA and protein were evaluated by PT-PCR and Western Blot after the intravitreal injection of NgR siRNA,Western Blot was also used to evaluate the expression of GAP-43 protein. Results1.The expression of Nogo-A was found in the optic nerve,retinal ganglion cells layer, nerve fiber layer,inner nuclear layer,inner plexiform layer,outer plexiform layer and outer nuclear layer.The expression of NgR was found in the optic nerve,retinal ganglion cells layer,nerve fiber layer and inner plexiform layer.2.Rat models with acute and chronic ocular hypertension were successfully established.3.In the retina and optic nerve in rat model with acute ocular hypertension,the significant upregulation of the level of Nogo-A mRNA and protein was found at 1d,3d after the model establishment(P＜0.05 in comparing with control group),and then decrease to the nomal level at 7d and 14d after the model establishment.In the retina and optic nerve in rat model with chronic ocular hypertension,the level of Nogo-A mRNA and protein were found to increase at 3d and 7d after the model establishment,and the increase remained significantly at 14d and 28d after the model establishment(P＜0.05 in comparing with control group).The level of NgR in the ratina and optic nerve in rat with the acute and chronic ocular hypertension was not found to change significantly.4.In the retina in rat model with chronic ocular hypertension,the expression of NgR mRNA was knocked down and NgR protein was suppressed after the intravitreal injection of NgR siRNA demonstrated by using RT-PCR and Western Blot.The expression of GAP-43 protein increased after the intravitreal injection of NgR siRNA compared with control.Conclusions1.Nogo-A and NgR were widely distributed in the retina and optic nerve of normal rats.2.The change of expression of Nogo-A mRNA and protein in the retina was associated with the elevated ocular pressure.The dramatically increased Nogo-A indicated that Nogo-A may play a primary role in obstructing regeneration of optic nerve.3.The expression of NgR in the retina and optic nerve in rat model with acute and chronic ocular hypertension was not significantly changed.In the retina in rat model with chronic ocular hypertension,the level of NgR mRNA expression was knocked down by the siRNA,and NgR protein was suppressed too.NgR may also play an important effect in the mechanism of inhibiting the axon regeneration after the ocular hypertension.NgR siRNA could improve the expression GAP-43 which may promote the axon regeneration of injured optic nerve.NgR siRNA would be an effective method in the treatment of glaucomatous optic neuropathy.