| The optic nerve injury is one of the common kinds of ocular trauma, offen resulting in severe vision deterioration even permanent blindness. It causes irreversibly primary injury and reversibly secondary injury. Optic nerve is composed with axons of retinal ganglion cells(RGC). Previous studies have suggested that apoptosis is a main type of retinal ganglion cells' secondary death after optic nerve injury. Many deleterious factors triggered following optic nerve injury including deprevivation of nerve growth factors , the calcium overload and free radicals can change the RGC micro-enviroment, and activate the expression of bcl-2 gene families and then induce RGC apoptosis. High dose methylprednosolone (MP) and optic nerve decompression surgery are the common therapeuticmethods. But recent years therapeutic results are controversial about high dose MP treatment on acute optic nerve injury, and few experimental study was reported. It has not been seen whether high dose MP can inhibit RGC apoptosis after optic nerve crush, and prevent RGC from secondary degeneration.In the study, the model of optic nerve crush was established and given high dose MP at the early stage, apoptosis cells, Bcl-2 and Bax-positive cells were located and calculated to investigate the therapeutic effect and mechanism by the terminal-deoxynucleotidyl transferase mediated DUTP-biotin nick end labeling (TUNEL) reaction and immunohistochemical method with Combination of retinal whole-mount and section technique .Materials and methodOne hundred and twenty six healthy adult Wistar rats without eye diseases were divided into three groups randomly: normal control group(n=18, 36 eyes) , crush control group(n=54, 54 eyes) and treatment group(n=54, 54 eyes). According to the surviving time after crush, the groups were sub-divided into 3 timepoints: 4 d, 7 d and 14 d, there are 12 eyes at each timepoint of normal control group, and 18 eyes at each timepoint in the crush group and treatment group. The animal of the crush group and treatment group received optic nerve crush in unilateral eye by applying the lips of a cross-action forceps to the nerve at 3 mm behind the globe for 10 s. Thetreatment group were given MP firstly 30mg/kg 1 h post-crush, and 6 h later, received 5.4 mg/kg per hour, total dosage 246.8 mg/kg within 48 h. Crush control group received equal volume normal saline. All animals were sacrificed respectively at 4, 7 and 14 d after optic nerve crush, and the eyes were enucleated. The cell apoptosis of RGC from the injured optic nerve was examined by the terminal-deoxynucleotidyl transferase mediated DUTP-biotin nick end labeling (TUNEL) reaction, and the expressions of bcl-2 and bax genes were measured with immunohistochemical method at 4 d, 7 d, 14 d post-crush . All the data were analyzed statistically by ANOVA.Result1. The location and calculation of apoptosis cells: Cells with DNA fragmentations were detected using TUNEL staining. Apoptosis cells were only seen in the ganglion cell layer. Few TUNEL-positive cells were observed at normal retina. A large number of labeled cells were detected at 4 d after optic nerve crush, TUNEL-positive cells were 14.5 /mm2, 22.78 /mm2 and 5.36 /mm2 at 4 d, 7 d and 14 d post-crush, respectively. In the MP treated groups, the number of TUNEL-positive cells were 11.44 /mm2, 17.84 /mm2 and 1.88 /mm2 at 4 d, 7 d and 14 d after crush, respectively. High dose MP significantly reduced the number of apoptotic cells(/7<0.05) at various time points.2. The location and calculation of Bcl-2 and Bax positive cells:Bcl-2 positive cells were only seen in the ganglion cell layer at all kind of timepoints. Normal retina expressed bcl-2 gene at low level, the number of the positive cell were 6.37 /mm2, 6.63 /mm2 and 5.77 /mm2 4 d, 7 d and 14 d, respectively. The expression of bcl-2 gene gradually increased after optic nerve crush, the number of Bcl-2 positive cells were 17.38 /mm2, 21.54 /mm2 and 27.64 /mm2 at 4d, 7d and 14d post-crush, respectively. In the MP treated groups, the number...
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