A Study Of The Protective Effect Of Aminogunidine On Retinal Ganglion Cells After Optic Nerve Injury In Rabbits

ObjectiveOptic nerve injury is a common eye disease, Which is usually coincidenced on skull and brain injury, If the medical prognosis was bleak, it may lead to patient blindness. As the pathogenesis of optic nerve injury has not yet been fully clear,So far the treatment is still a hang-up in ophthalmology group of internal and external. The aim of the study,through establishing the model of optic nerve injury , and Aminogunidine treats early it after optic nerve injury ,to observe the pathomorphologic characteristics of retina and the expression of NO/iNOS,MDA/SOD.Through this study,Which will explore a new method of treatmenting optic nerve damages and provide the experiment evidence for the clinical treatment. To save the Visual function of the patients.Materials and methodSixty-six healthy adult rabbits approximate 2.5 kilograms without eye diseases,were randomly divided into three groups:Normal control group(n=6),injury treatment group(n=30),Injury control group(n=30). According to the surviving time after injury,the injury group were divided into 5 subgroup by sacrificed time:1d,3d,7d ,14d and21d,there are 12 eyes at each subgroup. The animals of the injury treatment group and injury control group received optic nerve clipping injury with the same artery clamp(15cmH₂O)forceps to the nerve at 3mm behind the globe for 20s. The injury treatment group were injected 2%AG(80mg/Kg)through ear-border vein,once every day.The injury control group were were injected equal 0.9%NS through ear-border vein,once every day. All animals were killed respectively at 1d,3d,7d 14d and21d after optic nerve injury,and the eyes were enucleated.Half of them,retinal slice stained HE and TUNEL to locate and calculate apoptosis cells. Half of them,take biochemistry way to determine the Nitric oxide (NO) ,s content , inducible nitric oxide synthase (iNOS),s content , Superoxide Dismutase (SOD),s vitality and the Maleic Dialdehyde (MDA),s content in retina paste. All the data were statisticed with the SPSS13.0 software.Result1.HE stain of retina:Normal control group: layer of retina is smooth and complete,there are three layer cells in the retina. The layer of RGC is monolayer. RGC are divided into two types according to the size of nucleus: big nucleuses are light-colored and small nucleuses are deep- dyed. Intranuclear caryotin is well distributed. The cells in interior and external plexiform layer are compacted and even at thickness and dyeing.The injury control group: After injured 1d, retina were slight edema and complete.After injured 3d ,retina had become more edema,vacuolar degeneration seldom. After injured 7d,14d, there are serious edema in every retinal layer, especially the nerve fiber layer , RGC layer and inner plexiform layer . Thickening every layer was pultaceous and cell spaces increasing. After injured 21d ,retinal edema was abatement . The number of RGC began to decrease and the RGC partly became vacuolar degeneration. The arrange of cells inner nuclear layer got more derangemen .Thickness of retina obviously became thinness.The injury treatment group: the pathology change(cell spaces,retina edema, RGC,s number) in treated group of each time-point were lighter than the injury group. 2.The locatio and calculation of TUNEL stained apoptosis cells:Cells with DNA fragmentations were detected using TUNEL staining. Apoptosis cells were not only seen in the ganglion cells layer, but also in the interior and external plexiform layer .Apoptosis cells were stained at nucleuses, nucleuses were yellow brown. Some may be loss of normal morphology because nuclear is condensation and distorted and breakdown , Sometime little apoptosis bodies could be seen.Few TUNEL-positive cells were detected at normal retina. In this study, only calculates apoptosis cells of retinal ganglion cells. The number of TUNEL-positive RGC cells in injury treatment groups were significantly reduced (P<0.05) at the same time points .3.The nitric oxide(NO) ,s content and the inducible nitric oxide synthase (iNOS),s vitality in retina: iNOS is expressed rarely in normal retinal tissue, but contains a small amount of NO. Both content increased gradually after injury. The NO ,s content and iNOS ,s vitality in retina of the injury treatment groups were significantly different (P<0.05)at the same time points .4.The Maleic Dialdehyde(MDA),s content and the Superoxide Dismutase (SOD) ,s vitality in retina: In normal retina paste,there were some MDA and SOD. The MDA,s content in retina of the injury treatment groups were significantly different (P<0.05) at the same time points .The Superoxide Dismutase (SOD) ,s vitality in retina of the injury treatment groups were significantly different (P<0.05) at various time points.Conclusion1. The pathology change in treated group of each time-point were lighter than the injury group. So AG ear-border vein injection in earlier period had a definite protective effect on retina after optic nerve clipping injury.2. INOS is expressed rarely in normal retinal tissue, but contains a small amount of NO.Both content increased gradually after injury.the results is consistent trends apoptosis of RGCs after optic nerve injury. Indicating that large number of NO and iNOS is a factor ,which will lead to apoptosis of RGCs after optic nerve injury.3.MDA,s content increased gradually in the Retina tissue after optic nerve injury.With tissue injury increase further,SOD,s vitality decreased gradually.Weakening its protective effect on the optic nerve and retina. Free radicals accumulation continued,leading to cell occurs damage irreversible.4. AG ear-border vein injection in earlier period can reduce the degree of apoptosis of RGCs After optic nerve injury. The experiment show AG can reduce NO synthesis by inhibiting the production of iNOS, and inhibit free radical formation, then protect the RGCs after optic nerve injury.