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AIM: To investigate the effect of the intravitreal injection of vascular endothelial growth factor-A165(VEGF-A165)on the scleral remodeling of guinea pigs with form-deprivation myopia(FDM).METHODS: A total of 120 tricolor guinea pigs, aged three weeks, were randomly divided into 6 groups, with 20 in each group. The blank group did not undergo any intervention. In the FDM group, only the FDM model was established. In the phosphate buffer saline(PBS)group, 2.5 μL of PBS was injected into the vitreous cavity before establishing the FDM model. In the 1ng group, 5ng group, and 10ng group, VEGF-A165 was injected into the vitreous cavity at concentrations of 1, 5 and 10ng, respectively, before the establishment of the FDM model. The FDM model was established by covering the right eyes of guinea pigs with translucent balloons for 14d. The diopter and axial length of the right eyes were measured before and after covering. After 14d, the content of dopamine(DA)in retina was measured by high performance liquid chromatography. Additionally, the mRNA and protein expression levels of matrix metalloproteinase-2(MMP-2), tissue inhibitor of matrix metalloproteinase-2(TIMP-2), transforming growth factor(TGF)-β1, TGF-β2 and α-smooth muscle actin(α-SMA)in sclera were detected by reverse transcription polymerase chain reaction(RT-PCR)and Western blot.RESULTS: Before covering, there were no significant differences in the diopter and axial length of the right eyes of guinea pigs in all groups(P>0.05). After 14d of modeling, when compared with the blank group, FDM group showed an increase in the degree of myopia in the right eye, a prolongation of the axial length, a decrease in the content of DA in the retina, and an increase in the expression of MMP-2, TGF-β2 and α-SMA in the sclera. Conversely, the expression of TIMP-2 and TGF-β1 were decreased(P<0.01). However, in comparison to the FDM group, the degree of myopia in the 1ng, 5ng, and 10ng groups of guinea pigs decreased, the growth trend of axial length slowed, the content of DA in the retina increased, and the expression of MMP-2, TGF-β2 and α-SMA in the sclera decreased. Furthermore, the expression of TIMP-2 and TGF-β1 in the sclera increased(P<0.01). As the concentration of intravitreal injection of VEGF-A165 increased, the degree of myopia in the right eye of guinea pigs gradually increased, and the axial length gradually prolonged. The content of DA in the retina gradually decreased, the expression of MMP-2, TGF-β2, and α-SMA in the sclera gradually increased, while the expression of TIMP-2 and TGF-β1 decreased gradually.CONCLUSION: Intravitreal injection of VEGF-A165 can increase the content of DA in the retina of FDM guinea pigs, affect the expression of MMP-2, TIMP-2, TGF-β1, TGF-β2 and α-SMA in the sclera, and inhibit scleral remodeling of guinea pigs. Notably, the VEGF-A165 at the concentration of 1ng showed the most significant efficacy.
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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.
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@#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.
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Myopia is one of the most common ocular problems,which causes visual dysfunction.The prevalence of myopia is increasing,while the prevention and treatment have made no breakthrough because the pathogenesis and mechanism of myopia remains unclear.Certain kinds of molecules were reported to play a role in myopia,such as dopamine,retinoic acid,glucagon,ZENK (Zif269、EGR-1、NGFI-A or Krox-24) etc.As a derivative of vitamin A,retinoic acid is the final metabolite of retinaldehyde,which combines to opsin in the photoreceptor cells on retina.Previous studies demonstrated that retinoic acid from choroid and retina plays significant roles in the development of myopia,which affects different molecules related to myopia development,including transforming growth factor-β,glycosaminoglycan,matrix metalloproteinase (MMP)-2 and tissue inhibitor of metalloproteinase (TIMP)-2.Based on the synthesis,mechanism and effects of the retinoic acid in the eye,this paper describes the new progress of retinoic acid in experimental myopia,including form deprivation myopia,defocus induced myopia and monochromatic myopia,hence offers new clues for the research on the mechanism of myopia.
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Objective To evaluate the inhibitory effect of highly selective M4 receptor antagonist MT3 on the form deprivation myopia in guinea pigs and its potential mechanism. Methods Thirty?two healthy male guinea pigs were ran?domly divided into three groups:control group, form deprivation group, and form deprivation + MT3 group, 8 animals in each group. Refraction was measured by retinoscopy after cycloplegia before and after the experiment. The ocular biological dimensions were measured by A?scan ultrasound. RT?PCR was used to detect the relative expression of TGF?β2 mRNA in the retina and choroid. Results Compared with the right eyes of control group, the right eyes of form deprivation + MT3 group developed relative myopia of -1?44 ± 0?50 D (right?left eye) (P =0?001). The vitreous chamber depth and axial length of the right eyes were significantly prolonged by 0?10 ± 0?02 mm and 0?14 ± 0?07 mm (P<0?001, P<0?001), respectively, but the increases of myopia and axial length were significantly smaller than that of the form deprivation group (P<0?001, P<0?001, P<0?001). Down?regulation of relative mRNA expression of TGF?β2 in retina and choroid was found in the form deprivation group (P<0?001, P =0?014) compared with the right eyes of the control group, while up?regulation of relative mRNA expression of TGF?β2 in retina and choroid was found in the form deprivation + MT3 group ( P<0?001, P<0?001). Conclusions MT3 can inhibit the development of form deprivation myopia in guinea pigs, which may play an important role by the regulation of TGF?β2 mRNA level in the retina and choroid.
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Objective To observe whether form-deprivation myopia/amblyopia could protect retinal microvascular system and postpone the development of diabetic retinopathy. Methods The form-deprivation myopia/amblyopia was established combined with hyperglycemia mouse model,and the retinal vascular endothelial cells and pericytes were counted. Results Endothelial cells/ pericytes ratio in hyperglyce-mia group is significantly lower than that in normal group and form-deprivation myopia/amblyopia combined with hyperglycemia group. Con-clusion Form-deprivation myopia/amblyopia combined with hyperglycemia mice have more pericytes than hyperglycemia mice.
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Background It is well known that sclera remodeling occurs during axial elongation in myopia under the control of growth hormone or its downstream effectors.The role of transforming growth factor-β (TGF-β) in myopia has been determined in previous studies.Bone morphogenetic protein (BMP) is one of members of the TGF-β superfamily,but if it plays an important role in the genesis and development of myopia is not completely clear.Objective This study was to identify the presence of BMPs in normal guinea pigs sclera and investigate the change of BMPs in the sclera in form-deprivation myopia (FDM) of guinea pigs.Methods Thirty young guinea pigs were randomized into normal control group and experimental group using table of random number.FDM models were established by occluding unilateral eyes of guinea pigs with a translucent lens for 14 days in the experimental group,and the fellow eyes served as the controls.Diopter of all eyes was tested by retinoscopy optometry,and ocular axial length was measured by A-sonography before and after modeling.Posterior sclera tissue of the animals was obtained on 14 days,and the relative expression level of BMPs mRNA and protein were assayed by reverse transcription PCR (RT-PCR) and Western blot.The use and care of the animals complied with ARVO Statement.Results On 14 days after occluding of unilateral eyes,the refraction diopter of the experimental group was (-0.48±0.51) D,and that of the fellow eyes was (3.22 ±0.34) D,showing a significant difference between them (t =-12.814,P =0.000).Also,a significant difference in the diopter was seen between the experimental group and normal control group ([-0.48±0.51]D vs.[2.97±0.70]D,t =-11.878,P=0.000).Axial length was (8.30 ± 0.05) mm in the experimental group,(8.11 ±0.06) mm in the fellow eyes and (8.06±0.06) mm in the normal control group,showing a significant increase in the experimental group compared with the fellow eyes and normal control group (t =7.230,P =0.000 ; t =9.084,P=0.000).The expressions of BMP-2 mRNA,BMP-4 mRNA,BMP-5 mRNA in posterior sclera were detected in the normal guinea pigs.Fourteen days after the induction of myopia,the relative levels of BMP-2 mRNA and BMP-5 mRNA in sclera were 0.41 ± 0.11 and 0.65 ± 0.06 in the experimental eyes,which were significantly lower than 0.62 ± 0.07 and 0.84 ± 0.03 in the fellow eyes with the descent range of 34.48％ and 23.67％ respectively (t=2.838,P=0.017; t=2.524,P=0.028).The relative values of BMP-2 protein and BMP-5 protein were 0.44±0.06 and 0.70±0.05 in the experimental eyes,and those of the fellow eyes were 0.61±0.05 and 0.82±0.03,showing significant decline in the experimental eyes with the lowing range of 23.42％ and 15.21％,respectively (t =2.465,P =0.030;t =2.445,P=0.031).No significant differences were found in the expression of BMP-4 mRNA and protein in posterior sclera between the experimental eyes and the normal control eyes (mRNA:t =0.704,P=0.460;protein:t=0.987,P=0.365).Conclusions The expressions of the BMP-2 and BMP-5 in sclera down-regulate significantly in FDM eyes,which suggest that BMP-2 and BMP-5 participate in sclera remodeling during myopia induction.
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PURPOSE: To evaluate the influence of suppression by intermittent exotropia on axial length progression. METHODS: The medical records of patients with intermittent exotropia who had undergone surgery at the Korea University Medical Center from 2003 to 2010 were reviewed. The age upon visit, age at operation, visual acuity, refractive error, type of strabismus, angle of strabismic deviation, suppression test (Vectographic projector test, Reneau, France), and axial length test were analyzed. Subjects with amblyopia or anisometropia were excluded. RESULTS: A total of 75 patients with intermittent exotropia who had definite suppression in 1 eye were identified. The mean age at visit was 6.87 +/- 2.73 years and mean angle of exodeviation was 25.65 +/- 6.68 / 26.17 +/- 6.59 (prism diopters, distant / near). There was no statistical difference in exotropia patients' interocular axial length value who showed suppression in 1 eye (p = 0.992 in the right-eye suppression group, and p = 0.528 in the left-eye suppression group). CONCLUSIONS: In the present study, there was no statistical difference in interocular axial length value of intermittent exotropia patients with suppression in one eye (p > 0.05).
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Humans , Academic Medical Centers , Amblyopia , Anisometropia , Exotropia , Eye , Korea , Medical Records , Refractive Errors , Strabismus , Visual AcuityABSTRACT
Background The domestic and international researches discovered that many proteins and enzymes of the extracellular matrix (ECM) participate in the sclera remodeling by affecting the collagen typeⅠand fibronectin.Objective This study was to investigate the effect of matrixmetalloproteinase-2 (TIMP-2) on expression of collagen typeⅠand fibronectin of ECM in the posterior sclera by injecting liposomes containing tissue inhibitor of TIMP-2 gene into suprachoroidal space of the form-deprivation myopia in guinea pig.Methods Form-deprivation myopia was induced by translucent goggles in 36 clean guinea pig for 2 weeks.Then the animals were randomly assigned to TIMP-2 group,empty plasmid group,saline group and 12 for each group.Liposomes of 5μl containing TIMP-2 gene,empty plasmid and saline were suprachoroidally injected in the right eye respectively,and the left eyes without any treatment were used as self-control group.Other 12 matched guinea pigs only covered the right eyes through out the experimental duration as model control group.The guinea pigs were sacrificed and the posterior sclera tissue of the eyeballs were collected at 2,7 and 14 days after injection of drug.The expressions of collagen typeⅠmRNA and fibronectin mRNA were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR).This study followed the Regulation for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.Results The expression level of collagen type Ⅰ mRNA in the posterior sclera of guinea pig was lower but that of fibronectin mRNA was higher in TIMP-2 group than self-control group,showing significant differences between them (P＜0.05).The expression level of collagen type Ⅰ mRNA in the posterior scleral tissue began to increase from the 2nd day after drug injection and was obviously elevated at the 7th day and then gradually decreased at the 14th day.However,the expression level of fibronectin mRNA in the posterior scleral tissue showed the opposite pattern.The expression levels of collagen typeⅠmRNA and fibronectin mRNA at the 7th after drug injection were significantly lower than that at the 2nd day or 14th day (P＜0.01).Conclusion Suprachoroidal injection of TIMP-2 in form-deprivation myopia could up-regulate the expression of collagen typeⅠmRNA and down-regulate the expression of fibronectin mRNA in the posterior scleral tissue.It may slow down the sclera remodeling of form-deprivation myopia in guinea pig in the early stage.
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Objective Development of eyeball is visually regulated by messengers that released from the retina, and research demonstrated that retinoic acid (RA) is the only extrinsic biochemical candidate that could act as a growth controller. Present study was designed to evaluate the RA system in retina of guinea pig eye with experimental form deprivation myopia. Methods The form deprivation myopia models were monocularly established in the 24 2-week-old guinea pigs by occluding the lateral eyes using white translucent hemispheres for two weeks, and the fellow eyes were as normal control. Refraction diopter was detected with streak retinoscopy after cycloplegia, and axial length and vitreous depth were calculated with Cinescan A/B ultrasonography before and 2 weeks after experiment. All animals were sacrificed and retina was dissected 2 weeks after experiment. The RA level in retina was delected by High Performance Liquid Chromatography (HPLC). The expressions of RA-binding proteins Ⅰ(CRABP-Ⅰ) and RA receptor-β(RAR-β) protein and mRNA were assayed by Western-blotting and Real-time PCR, respectively. The experiment and use of animals followed the Standard of Association for Research in Vision and Ophthalmology. Results The spherical equivalent refraction was (+ 3. 00 ± 0. 75) D in the model group and (+ 2. 88 ± 0. 67) D in control eyes (t= 0. 672, P ＞ 0. 05), the ocular axial length was (7. 822 ± 0. 083) mm in model group and (7. 791 ±0.073) mm in control eyes before experiment (t = 0. 346, P ＞ 0. 05). In 2 weeks after experiment, the spherical equivalent refraction was (- 3. 82 ± 0. 13) D versus (1. 99 ± 0. 58) D and axial length was (8. 346 ± 0. 047) mm versus (7. 888 ± 0. 042) mm between model eyes and control eyes (t = 8. 376, P ＜ 0. 001; t = 3. 343, P ＜0. 05). No significant difference in the level of RA in retina before and after experiment (1. 394 ±0. 079 μg/g vs 1. 295 ±0. 023 μg/g) (t =0. 897, P ＞0. 05) but obviously elevated after experiment in model eyes compared with control eyes (2. 356 ± 0.098 μg/g vs 1.499 ±0.035 μg/g) (t =4. 934, P ＜0. 01). The expression of CRABP-Ⅰ and RAR-β mirrored these directional changes. Conclusion The RA system in retina is upregulated in the eye of guinea pig with form deprivation myopia. This findings suggests that RA may act as a messenger in the development of experimental myopia.
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Objective Researches showed that retinoic acid may be a signal molecular in the regulation of ocular axial length.Sclera is a target tissue of the pathogenesis of form deprivation myopia.Recently,guinea pigs are used to the study of the pathogenesis of form deprivation myopia.However,seldom studies are found in the change of retinoic acid level in the sclera of guinea pigs.This study was designed to investigate the role of retinoic acid of early form deprivation myopia in guinea pigs sclera.MethodsMonocular form deprivation models were established in 24 3-week-old guinea pigs by occluding the left eyes with facemask and randomly divided into form deprivation 15 days group and 24 days group.Other 6 normal guinea pigs were used as control group.The fellow eyes were used as internal control.Refraction was measured before and after experiment by using retinoscopy.Eyeballs were extracted immediately at the corresponding time points and ocular axial length was measured with outside micrometer.The level of retinoic acid in scleral samples was detected by high performance liquid chromatography.The use of animal followed the Regulations for the Administration of Affairs Concerning Experimental Animals by the State Science and Technology Commission.ResultsRefractive diptor reduced by -3.14±1.50 D in experimental eyes after form deprivation,showing statistically differences among before and various time points after form deprivation(F=23.053,P=0.000),and no significant difference was seen in reduce of refractive diptor in fellow eyes after form deprivation( -1.24±1.99 D)(F=1.506,P=0.240).The axial length was considerably increased in form deprivation eyes after experiment (F=14.551,P=0.000) but no evident change in fellow eys (F=3.612,P=0.060).The diopter and ocular axial length was significantly different between form deprivation eyes and fellow eyes (P＞0.05).The expression of retinoic acid in sclera of form deprivation eyes was higher than fellow eyes in both form deprivation 15 days and 24 days(P＜0.01).There was no significant difference in the level of retinoic acid between form deprivation 15 days and 24 days(P＞0.05).ConclusionForm deprivation myopia can be induced in 15 days after monocular occlusion.Retinoic acid levels in guinea pig sclera increased significantly during early form deprivation.
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Objective: To observe the effect of Ginkgo Biloba extract (EGb) on Form deprivation Myopia (FDM) in the chick. Methods: 30 two day chicks were divided into three groups (Control Group, Drug Group Ⅰ, Drug Group Ⅱ). All of them were monocularly deprived by suturation of eyelids. Axial length was measured by A Scan ultrasonograph, the posterior cartilaginous sclera, the sclera fibrous and retina were observed under the light microscope. Results: Both refraction and axial eye length were changed by form deprivation in control group. refraction change ( P