Study Of Bak,Bcl-xL And Apoptosis In The Development Of Guinea Pigs Form-deprivation Myopia

Background and ObjectiveHigh myopia is also called pathological myopia, which is usually associated with progressive elongation of the eye axis, myopia aggravation, the degeneration of retina and other complications. Because of the hazard of high myopia, the world has focused on it. During the study history of more than 200 years, many hypotheses were proposed, there was still no satisfactory explanation to myopia pathogenesis. Form-deprivation theory proved that some factors influenced the normal image in retina, which can lead to myopia. Form-deprivation myopia (FDM) animal model was established in 1977, which excessively pushed the study of myopia. High myopia eye posterior pole pathological extension, accompanied retinal nerve cell decreased. While nerve cells died by apoptosis in many retinal degeneration and nerve system diseases. Furthermore, Xu Gezhi investigated photoreceptor cells apoptosis in human high myopia.Cell apoptosis is also called programmed cell death which is regulated by many genes. Bcl-2 gene protein family is one of the most important factors, which includes Bcl-2, Bax,Bak,Bcl-xL, and so on. The family members combined each other to control the course of apoptosis. We would investigate whether there are obvious retinal apoptosis in guinea pig FDM and observe the expression of Bak,Bcl-xL in order to know the relationship between Bak,Bcl-xL and retinal apoptosis in high myopia.Materials and MethodsSixty healthy colored guinea pigs were selected. All right eyes underwent form deprivation with translucent goggles, and the left eyes were used as control. They were randomly divided into three groups, included A,B and C groups, which were deprived for 4 weeks,8 weeks and 12 weeks respectively. The eyes refractive state was determined by means of streak retinascopy. The eyes axial length was determined by ophthalmological A ultrasound. Extracted the eyes as quickly as possible, routine fixation, dehydration, paraffin embedding were done and 4μm paraffin sections were taken. Routine hematoxylin-eosin staining were done and morphological changes were detected by light microscope. We would use TUNEL technique to observe the retinal apoptosis. Furthermore measured the expression level of Bak,Bcl-xL on the guinea pigs retina using streptavidin-peroxidase immunohistochemical method.Results1. Guinea pig monocular deprivation could obviously lead to development of myopia , the refractive state and axial length showed significant difference between the occluded eyes and the control eyes (P<0.05) . The longer the eyes were occluded ,the more serious the myopia was, and there was significant difference between the occluded eyes of the three groups(F<0.05) .There was the linear correlation between the refractive state and the axial length in the occluded eyes (r =-0.855).2. TUNEL staining showed apoptosis cells scattered in the retinal inner nuclear layer and outer nuclear layer of B and C groups' occluded eyes, and the nucleolus of them were brown-yellow. The retinal cells apoptotic rate of group B and group C were (0.27 + 0.14)% and (2.33±0.37)%. There was significant difference between the two groups(P<0.05). In the occluded eyes of A group and control eyes no apoptosis cells were found.3. Immunohistochemical staining showed that Bak positive cells were detected in the photoreceptor cells layer inner segment, inner nuclear layer and ganglion cell layer. Bcl-xL positive cells were detected in the photoreceptor cells layer, outer nuclear layer, and ganglion cell layer. Both Bak and Bcl-xL positive cells staining showed cytolymph brown-yellow. Optical density value of Bak indicated: the group C occluded eyes (113.48±7.77) has significant difference between and its control eyes (94.52±7.10) and group B occluded eyes (97.26±7.23) (P<0.05). The expressions of Bcl-xL have no significant difference between these groups (P>0.05 ) .4. The correlation coefficient between apoptotic rate of occluded eyes retinal cells of group B,group C and the expressions of Bak are 0.79 and 0.88; which and the expressions of Bcl-xL are -0.37 and -0.31.Conclusions1. Guinea pig monocular deprivation could obviously develop myopia, and the longer the eyes are occluded, the more significant the difference was, the guinea pig FDM model was successful. High myopia could be formed until 12 weeks.2. The apoptotic cells were detected in the inner nuclear layer,outer nuclear layer of guinea pigs deprived eyes. Apoptosis is one of the FDM retinal cell death mechanisms.3. Retinal cells from FDM expressed Bak and Bcl-xL.4. There was positive correlation between the Bak and the retinal cells apoptosis. Bak creased maybe the reason of FDM retinal cell apoptosis.