RÉSUMÉ
AIM: To investigate the effects of modified Zhujing Pill on the mitochondrion structure and dynamin-related protein of retinal pigment epithelial cells(RPEs)in mice with form deprivation myopia.METHODS: 3-week-old C57BL/6J mice were randomly divided into control group, model group and Chinese medicine group, with 10 mice in each group. Myopia model of the right eye of mice was established by means of form deprivation in model and Chinese medicine groups. After 4wk, the Chinese medicine group were given intragastric administration of modified Zhujing Pill suspension 0.546g/(kg·d)(0.2mL/d)for 4wk, and same amount of saline was given to mice in other groups at the same time of modeling. The axial length and diopter of the right eye of the mouse were measured before and after the experiment by A-ultrasound and a strip retinoscope respectively. At the end of the experiment, the mitochondrial ultrastructure of RPEs was observed by transmission electron microscope. Western blot, and real-time fluorescent quantitative PCR(q-PCR)were used to detect quantitative and gene expression of mitofusin 1(MFN1), optic atrophy 1(OPA1), and dynamin-related protein 1(DRP1)in retinal tissues respectively.RESULTS: At the beginning of the experiment, there was no statistically significant difference in axial length and diopter of the right eye of the mouse in control, model and Chinese medicine groups(P>0.05). At the end of the experiment, compared with the control group, the mice in the model group and the Chinese medicine group had lower diopter and continuously prolonged axial length(all P<0.05), while the mice in the Chinese medicine group had significantly shorter axial length and higher diopter than the model group(all P<0.05). Western blot and q-PCR results showed that the relative expression of MFN1 and OPA1 decreased and DRP1 increased in both the model group and the Chinese medicine group compared with the control group(all P<0.05), and the relative expression of MFN1 and OPA1 increased in the Chinese medicine group compared with the model group(all P<0.05). The electron microscopic results showed that the mitochondria in the right retina of the mice were only mildly swollen in the Chinese medicine group, while the mitochondria in the model group were obviously swollen and disordered and empty.CONCLUSION: Modified Zhujing Pill could protect the retinal mitochondria by regulating the key proteins of mitochondrial dynamics(MFN1, OPA1, and DRP1), and it has a protective effect on the retina of axial myopic mice.
RÉSUMÉ
AIM: To investigate the effect of modified Zhujing pill on retinal autophagy in mice with form deprivation myopia.METHODS: Thirty C57BL/6 mice were randomly divided into a negative control group, a myopia model group and a traditional Chinese medicine intervention group, with 10 mice in each group. Except for the negative control group, all mice in the myopia model group and the traditional Chinese medicine intervention group used translucent EP tubes to cover their right eyes to make a form deprivation myopia(FDM)model; The traditional Chinese medicine intervention group gavage Zhujing pill modified suspension 0.546g/(kg·d)(0.15mL/d), the negative control group and the myopia model group were given an equal amount of normal saline(0.15mL/d)for 4wk. At the beginning and the end of the experiment respectively, the right eye diopter of the mouse was measured with a strip retinoscope, measurement of the axial length of the right eye of mouse by A-ultrasound. At the end of the experiment, the right eyes of all mice were taken for detection, and immunofluorescence method was used to locate and detect the activity and migration of the retinal microglia marker(Iba1); Transmission electron microscope observation of autophagosome formation in retinal pigment epithelial cells; Western Blot, real-time fluorescent quantitative PCR(q-PCR)to detect the autophagy marker LC3Ⅱ and p62 protein quantitative and gene expression in retinal tissues.RESULTS: At the end of the experiment, the refractive power of the right eyes of mice showed that the myopia model group and the traditional Chinese medicine intervention group formed relative myopia, the myopia model group and the traditional Chinese medicine intervention group were significantly lower than those of the negative control group(all P<0.01). At the end of the experiment, the axial length of the myopia model group and the Chinese medicine intervention group were significantly increased compared with the negative control group(P<0.01). Immunofluorescence method for locating and detecting Iba1 showed that the average optical density of Iba1 in the retina of the myopia model group increased the most obviously, followed by the increase in the negative control group, and the decrease in the traditional Chinese medicine intervention group. Compared with the negative control group, the myopia model group increased significantly(P<0.05), and the traditional Chinese medicine intervention group was significantly lower than the myopia model group(P<0.05). It was found that Iba1 migrated to the ganglion cell layer in the myopia model group and the traditional Chinese medicine intervention group. Transmission electron microscopy showed that autophagosomes were observed in the retinal pigment epithelial cells of the myopia model group and the Chinese medicine intervention group. The results of Western Blot and q-PCR showed that the expression of LC3Ⅱ and p62 increased most obviously in the traditional Chinese medicine intervention group, followed by the myopia model group, and the negative control group was the lowest.CONCLUSION: The results of the study show that modified Zhujing pill may enhance retinal autophagy in mice with FDM by inhibiting the activation of microglia.
RÉSUMÉ
@#AIM: To investigate the effect of modified Zhujing pill on retinal thickness and apoptosis in form deprivation myopia(FDM)mice.<p>METHODS: Totally 72 C57BL/6J mice aged 3-week-old were randomly divided into control group 1, model group 1, intervention group 1, control group 2, model group 2 and intervention group 2, with 12 mice in each group. The first three groups were tested for 3wk and the last three groups were tested for 6wk, except for the groups of control 1 and control 2, all the mice used translucent goggles to cover their right eyes for form deprivation. The mice of intervention 1 and intervention 2 were respectively given intragastric administration modified Zhujing pill suspension 0.546g/(kg·d)(0.1mL/d)for 3wk at the beginning and after 3wk of the experiment. Same amount of saline was given to mice in other groups at the same time of modeling. The eye axis was measured before and after the experiment. At the end of the experiment, the eye of mice was taken for HE staining to observe the thickness changes of each layer of retina. Immunohistochemistry(IHC)and western blotting(WB)were used to detect Bcl-2 and Caspase3 expression of protein.<p>RESULTS: At the end of the experiment, the axis of model group 1 was significantly higher than that of control group 1(<i>P</i><0.01), the axis of intervention group 1 was significantly lower than that of model group 1(<i>P</i><0.01), and the axis of model group 2 was significantly higher than that of control group 2(<i>P</i><0.01), the eye axis of intervention group 2 was significantly lower than that of model group 2(<i>P</i><0.01); HE staining showed that the thickness of NFL and ONL of model group 1 was significantly thinner than that of control group 1(<i>P</i><0.01). The thickness of INL of model group 1 was significantly thinner than that of control group 1(<i>P</i><0.05), and the thickness of NFL, INL and ONL of intervention group 1 was significantly higher than that of model group 1(<i>P</i><0.05); The thickness of NFL, INL and ONL model group 2 was significantly thinner than that of control group 1(<i>P</i><0.01); IHC testing showed that the average optical density of Bcl-2 protein in model group 1 was significantly lower than that of control group 1(<i>P</i><0.05), which in intervention group 1 was significantly higher than that of model group 1(<i>P</i><0.01), and which in the average optical density of Bcl-2 protein of model group 2 was significantly lower than that of control group 2(<i>P</i><0.01), which in intervention group 2 was significantly higher than that of model group 2(<i>P</i><0.01); Caspase 3 protein average optical density of model group 1 was significantly higher than that of control group 1(<i>P</i><0.01), which in intervention group 1 was significantly lower than that of model group 1(<i>P</i><0.01), which in model group 2 was significantly higher than that of control group 2(<i>P</i><0.05), which in intervention group 2 was significantly lower than that of model group 2(<i>P</i><0.01); WB test proved that the relative expression level of Bcl-2 protein in model group 1 was significantly lower than that of control group 1(<i>P</i><0.01), which in intervention group 1 was significantly higher than that of model group 1(<i>P</i><0.01), and which in model group 2 was significantly lower than that of control group 2(<i>P</i><0.01), which in intervention group 2 was significantly higher than that of model group 2(<i>P</i><0.01); The relative expression level of Caspase3 protein in model group 1 was significantly higher than that of control group 1(<i>P</i><0.01), which in intervention group 1 was significantly lower than that of model group 1(<i>P</i><0.01), the intervention group 2 was significantly lower than that of model group 2. <p>CONCLUSION: The results show that the modified Zhujing pill can interfere with the pathological changes of retinal thickness thinning in the process of myopia and formed myopia mice by regulating the expression of apoptosis-related proteins Bcl-2 and Caspase3, and alleviating the apoptosis of retinal cells in myopia formation and myopia mice.