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The purpose of traumatic optic neuropathy (Traumatic optic neuropathy, TON) is a serious complication of brain, eyes and facial trauma, severe visual impairment after injury, and often left permanent visual impairment. About TON of treatment have long been controversial. The study found that the differences in the degree of injury is an important factor to determine the therapeutic effect. Establish a standardized, reliable, convenient and easy, and can be repeated in varying degrees to quantify damage model, can provide an important prerequisite for further research TON of treatment mechanism. Traumatic optic nerve injury model is only qualitative injury or semi-quantitative damage, but the lack of a unified standard quantitative models. The constant pressure of the experiment using three vascular clips intended to cause the rabbit light, moderate and severe optic nerve injury model animal models, evaluate the therapeutic effect as the optic nerve damage. Materials and Methods animals and grouping healthy adult 48 white rabbits of either sex, weighing 2.0-2.5kg, eye examination was normal. 12 randomly selected, the left and right eyes were randomly divided into normal control group and the sham control group, and the remaining 36 were randomly divided into the damage the group Ⅰ damage Ⅱ group, injury group Ⅲ. 2 clamping force and injury intensity was measured accurately measured and calculated vascular clips clamping force and injury intensity (average impulse). The three vascular folder (small, medium and large) clamping force: 32g, 98g, 148g, average impulse were: 397.52g. s / mm 2 , 1209.88g. s / mm 2 , 1549.74g. s / mm 2 . 3 animal model using small, in the the queen three kinds of microvascular clip gripping three animal side of the eye injury group optic nerve 20s, respectively, resulting in varying degrees of injury group Ⅰ, the injury group Ⅱ injury group Ⅲ injury model; false injury group were exposed only to the side of the eye optic without applying clamping; other eye served as normal controls. 4 observation project and the point in time after injury, 3d, 1w 2w observe local histopathological changes of the optic nerve damage, Optic nerve Glee silver staining or Ponceau G-bright green stained optic nerve fibers and myelin integrated optical density was measured; retina Morphological observation of retinal ganglion cells (Retinal Ganglion Cell, RGC) counting RGC apoptosis detection and RGC apoptosis rate calculation. H after injury, 6h, 1d, 3d, the 1w line F-VEP examination, observed 5 groups P2 wave latency and amplitude changes. Statistical analysis data using one-way ANOVA and t-test for statistical analysis. Results 1 optic nerve damage local pathology observed results 1.1 optic nerve damage local HE staining and tissue morphology observed normal group and sham group, the optic nerve fibers arranged in dense rules, even dyeing, a small amount of glial cells. Damage Ⅰ group compared with the normal group change slightly. When the injury the Ⅱ group of 3d optic nerve edema, the axis zone infarction, glial cells arranged in disorder, with the passage of time, changing the heavier. Injury group Ⅲ 3d optic nerve edema, multiple necrosis, over time, the rapid development of lesions 2w nerve bundle structure disappeared. Each time point damage Ⅲ group change compared to the serious damage Ⅱ group. 1.2 optic nerve fibers Glee dip silver staining and integrated optical density measurement results at each time point sham group, the optic nerve morphology consistent with the normal group; optic nerve fibers integrated optical density compared with normal group, there was no significant difference (P> 0.05). At each time point the injury group Ⅰ optic nerve morphology similar to the normal group; integrated optical density of the optic nerve fibers than normal group decreased, but there was no significant difference (P> 0.05). When optic nerve injuries II group and injury group Ⅲ 3d fiber sparse distortions, over time, change is becoming apparent, at each time point injury group Ⅲ change significantly compared to the damage Ⅱ group than in the normal group; two groups at each time point optic fiber integrated optical density lower, the difference was significant (P <0.05), with the passage of time, the integrated optical density of the optic nerve fibers were reduced. Optic fiber integrated optical density at the same point in time, between the injury group compared the difference was statistically significant (P <0.01). 1.3 optic nerve slice Ponceau G-brilliant green stain and myelin integral optical density measurement results at each time point sham group, the optic nerve morphology is consistent with the normal group; myelin integrated optical density in comparison with the normal group, no significant difference ( P> 0.05). At each time point the injury group Ⅰ optic nerve morphology similar to the normal group; lower the myelin integral optical density than normal group, but there was no significant difference (P> 0.05). Injuries II group and injury group Ⅲ 3d demyelination, over time, change is becoming apparent, a large number of group Ⅲ myelin damage 2w disintegration nerve bundle structure disappeared each time point injury III group than the injury group Ⅱ change significantly; two groups at each time point myelin integrated optical density than those in the normal group decreased, and the difference was statistically significant (P <0.05), with the passage of time, the myelin integral optical density decreased. At the same point in time, the myelin integrated optical density among the injured group, the difference was very significant (P <0.01). 2 retinal pathology observed results clear level of 2.1 retinal the HE staining tissue morphology observed normal group and sham group, the retina ganglion cells in a monolayer arranged neatly intensive clearly nucleus, nuclear membrane smooth and complete. Damage Ⅰ group compared with the normal group change slightly. The injury group Ⅱ 3d RGC nuclear condensation, stain deepened reduce the number; subsequent retinal layers thinner lesions progressive increase over time. Large number of RGC injury group Ⅲ 3d karyopyknosis coloring deepened, RGC arrangement obviously sparse lesions rapidly worsened over time. Each time point injury III group than in the group Ⅱ lesions. 2.2 RGC counting the sham group RGC number each time point compared with normal group, there was no significant difference (P> 0.05). Each time point damage I RGC number decreased compared to the normal group, but there was no significant difference (P> 0.05). Each time point injury group Ⅱ and group Ⅲ RGC number of normal group, were significantly reduced, the difference was statistically significant (P <0.01), progressive reduction of RGC number over time. At the same point in time, RGC number among the injured group, the difference was statistically significant (P <0.05). 2.3 RGC apoptosis detection and average apoptosis rate of normal group and sham group, the retinal slice no apoptotic cells. The injury group apoptotic cells in the GCL. Damage to a small number of apoptotic cells in group Ⅰ was no significant difference (P> 0.05) between the RGC apoptosis rate at each time point. The injury group Ⅱ apoptotic cells increased resistance over time, the difference was statistically significant (P <0.05) in the RGC apoptosis rate between each time point. A large number of apoptotic cells in the injury group Ⅲ 3d, RGC apoptosis rate progressively increased over time, the differences between the various time points RGC apoptosis rate was significant (P <0.05). RGC apoptosis rate between each injury group at the same point in time, there are very significant differences (P <0.01). 3 F-VEP test results normal white rabbit P 2 -wave latency and amplitude (71.72 ± 3.66) ms (20.53 ± 4.15), respectively μv. Sham group and normal group at all time points P 2 wave latency and amplitude comparison, the difference was no statistically significant (P> 0.05). H after injury, the injured group showed P 2 wave latency delay and amplitude reduction, compared with the normal group, the differences were statistically significant (P <0.05). Injury the Ⅰ group of 1d when P 2 wave latency and amplitude returned to normal (P> 0.05). Injury in the II group and injury group Ⅲ, over time, P 2 wave latency delay and amplitude decreased. The Conclusion 1 application pressure 32g, 98g, 148g microvascular clip holder rabbit optic nerve 20s, can be made stable, repeatable light, medium and severe optic nerve injury animal model. Mild injury group after injury optic nerve morphology change slightly, no significant pathological changes, optic nerve conduction; moderate injury group after injury optic nerve morphology change significantly progressive increase over time damage the optic nerve conduction; severe injury group injured optic nerve rapid emergence of irreversibility degeneration, optic nerve function after injury completely lost. 3 moderate optic nerve injury model could be used as a treatment of traumatic optic neuropathy and effects of animal models.
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