豚鼠形觉剥夺性近视视网膜早基因c-fos与酪氨酸羟化酶关系的研究

形觉剥夺性近视（FDM）动物模型研究表明，高度近视眼形成主要由于局部视网膜的视觉信息可调控巩膜的生长反应，但目前对其具体发生机制不清。我们通过对豚鼠FDM形成及恢复过程中视网膜早基因c-fos和酪氨酸羟化酶（TH）的动态变化及相关关系进行研究，以探讨c-fos和TH在豚鼠FDM视网膜信号转导的可能分子机制。[第一段]     

形觉剥夺性近视中巩膜变化的生物学机制

近视眼的发病机制不明，形觉剥夺可以诱导典型的近视动物模型，形成形觉剥夺性近视眼(form—deprivationmyopia，FDM)。通过细胞和分子水平的研究发现，FDM的变化主要表现在眼球局部，形觉剥夺可诱导巩膜软骨细胞增殖，细胞外基质(extracellularmaterial，ECM)基因表达异常，ECM增加，巩膜胶原纤维改变等引起巩膜重塑，导致眼轴延长，近视屈光度增加。本文就FDM巩膜变化生物学机制的研究进展作一综述。     

豚鼠形觉剥夺性近视形成的影响因素

近视的发病机制为异常的视觉刺激作用于视网膜引起视网膜信号因子的改变,随后通过视网膜色素上皮层-脉络膜途径传导至巩膜,诱导细胞外基质重构,导致巩膜重塑。豚鼠形觉剥夺性近视（form-deprived myopia,FDM）是研究近视的常用模型之一,对豚鼠FDM的研究主要集中在使用不同的药物如一氧化氮合酶、褪黑素、M受体阻滞剂、多巴胺类物质、降眼压药物、过氧化物酶体增生物激活受体α激动剂和拮抗剂、缺氧诱导因子等进行干预,引起多巴胺、基质金属蛋白酶-2、基质金属蛋白酶抑制剂-2、转化生长因子-β、α-平滑肌肌动蛋白等因子及I型胶原等物质的变化,从而对FDM的过程产生影响。（国际眼科纵览,2021, 45： 256-262）     

视黄酸在形觉剥夺性近视眼巩膜中作用的研究

剥夺新生动物视网膜正常成像可导致轴性近视的发生,这种近视称为形觉剥夺性近视(FDM).高度近视形成过程中的眼轴过度延长及巩膜变薄,可能是受局部视网膜信息调控的巩膜细胞外基质(ECM)主动重塑的结果.巩膜主动重塑在眼球生长发育及正视化形成中起重要作用.视黄酸(RA)及其受体在FDM形成中对巩膜基质、软骨层和成纤维细胞层及基质降解相关蛋白酶的改变具有重要调节作用.它可能是调节FDM眼球伸长的信号分子,在巩膜重塑中扮演重要角色.     

视黄酸在形觉剥夺性近视眼巩膜中作用的研究

剥夺新生动物视网膜正常成像可导致轴性近视的发生,这种近视称为形觉剥夺性近视(FDM).高度近视形成过程中的眼轴过度延长及巩膜变薄,可能是受局部视网膜信息调控的巩膜细胞外基质(ECM)主动重塑的结果.巩膜主动重塑在眼球生长发育及正视化形成中起重要作用.视黄酸(RA)及其受体在FDM形成中对巩膜基质、软骨层和成纤维细胞层及基质降解相关蛋白酶的改变具有重要调节作用.它可能是调节FDM眼球伸长的信号分子,在巩膜重塑中扮演重要角色.     

视黄酸在形觉剥夺性近视眼巩膜中作用的研究

剥夺新生动物视网膜正常成像可导致轴性近视的发生,这种近视称为形觉剥夺性近视(FDM).高度近视形成过程中的眼轴过度延长及巩膜变薄,可能是受局部视网膜信息调控的巩膜细胞外基质(ECM)主动重塑的结果.巩膜主动重塑在眼球生长发育及正视化形成中起重要作用.视黄酸(RA)及其受体在FDM形成中对巩膜基质、软骨层和成纤维细胞层及基质降解相关蛋白酶的改变具有重要调节作用.它可能是调节FDM眼球伸长的信号分子,在巩膜重塑中扮演重要角色.     

视黄酸在形觉剥夺性近视眼巩膜中作用的研究

剥夺新生动物视网膜正常成像可导致轴性近视的发生,这种近视称为形觉剥夺性近视(FDM).高度近视形成过程中的眼轴过度延长及巩膜变薄,可能是受局部视网膜信息调控的巩膜细胞外基质(ECM)主动重塑的结果.巩膜主动重塑在眼球生长发育及正视化形成中起重要作用.视黄酸(RA)及其受体在FDM形成中对巩膜基质、软骨层和成纤维细胞层及基质降解相关蛋白酶的改变具有重要调节作用.它可能是调节FDM眼球伸长的信号分子,在巩膜重塑中扮演重要角色.     

视黄酸在形觉剥夺性近视眼巩膜中作用的研究

剥夺新生动物视网膜正常成像可导致轴性近视的发生,这种近视称为形觉剥夺性近视(FDM).高度近视形成过程中的眼轴过度延长及巩膜变薄,可能是受局部视网膜信息调控的巩膜细胞外基质(ECM)主动重塑的结果.巩膜主动重塑在眼球生长发育及正视化形成中起重要作用.视黄酸(RA)及其受体在FDM形成中对巩膜基质、软骨层和成纤维细胞层及基质降解相关蛋白酶的改变具有重要调节作用.它可能是调节FDM眼球伸长的信号分子,在巩膜重塑中扮演重要角色.     

视黄酸在形觉剥夺性近视眼巩膜中作用的研究

剥夺新生动物视网膜正常成像可导致轴性近视的发生,这种近视称为形觉剥夺性近视(FDM).高度近视形成过程中的眼轴过度延长及巩膜变薄,可能是受局部视网膜信息调控的巩膜细胞外基质(ECM)主动重塑的结果.巩膜主动重塑在眼球生长发育及正视化形成中起重要作用.视黄酸(RA)及其受体在FDM形成中对巩膜基质、软骨层和成纤维细胞层及基质降解相关蛋白酶的改变具有重要调节作用.它可能是调节FDM眼球伸长的信号分子,在巩膜重塑中扮演重要角色.     

视黄酸在形觉剥夺性近视眼巩膜中作用的研究

剥夺新生动物视网膜正常成像可导致轴性近视的发生,这种近视称为形觉剥夺性近视(FDM).高度近视形成过程中的眼轴过度延长及巩膜变薄,可能是受局部视网膜信息调控的巩膜细胞外基质(ECM)主动重塑的结果.巩膜主动重塑在眼球生长发育及正视化形成中起重要作用.视黄酸(RA)及其受体在FDM形成中对巩膜基质、软骨层和成纤维细胞层及基质降解相关蛋白酶的改变具有重要调节作用.它可能是调节FDM眼球伸长的信号分子,在巩膜重塑中扮演重要角色.     

豚鼠形觉剥夺超高度近视模型的形态学研究

目的建立豚鼠形觉剥夺超高度近视模型并观察其后极部各层组织的病理组织学变化。方法 2周龄三色豚鼠分为形觉剥夺组（12只）和非形觉剥夺对照组（8只）,于形觉剥夺前,形觉剥夺后4、6、10、14周分别对各组进行检影和眼轴性参数测量。通过病理组织学光镜检查分析形觉剥夺14周后各组视网膜、脉络膜和巩膜厚度及其形态学的变化。结果豚鼠形觉剥夺后随时间延长近视度数逐渐增高,10周后可达-10.00 D以上的超高度近视,14周后近视度数更高、个别个体可达-20.00 D。眼轴性参数相应延长。形觉剥夺超高度近视眼视网膜、脉络膜和巩膜较对照组均明显变薄并有病理性改变。结论应用遮盖法对豚鼠施行长期单眼形觉剥夺会形成超高度近视,10周后可达-10.00 D以上,超高度近视眼巩膜、脉络膜和视网膜明显变薄,脉络膜和巩膜结构发生紊乱。视网膜结构中感光细胞层变薄最为明显,推测其对超高度近视的发生起重要作用。     

Smad1在C57BL/6小鼠形觉剥夺性近视中的作用机制

目的：：通过建立C57 BL/6小鼠形觉剥夺性近视模型,探究Smad1在近视形成中的作用机制。方法：将60只3周龄C57 BL/6小鼠随机分为实验组和正常对照组(NC),包括形觉剥夺3wk组(FDM 3W,n=20),形觉剥夺4wk组(FDM 4W,n=20),FDM3W正常对照组(FDM 3W-NC,n=10)和FDM4W正常对照组(FDM 4W-NC,n=10),实验组右眼遮盖,左眼自然暴露作为自身对照,正常对照组不予任何处理。在实验3 wk及4 wk时检测所有小鼠屈光状态,HE染色观察巩膜及视网膜组织结构变化,免疫组织化学方法及实时荧光定量PCR观察Smad1在视网膜中的表达情况。结果：(1)自身对照眼和正常对照组呈生理性远视化发展,FDM3W、FDM4W实验眼均呈相对近视化发展,差异具有统计学意义(P<0.05);(2)与自身对照组及正常对照组比较,FDM3W、FDM4W实验眼巩膜及视网膜明显变薄;实验眼视网膜中Smad1表达明显下降,差异具有显著统计学意义(P<0.01)。结论：小鼠形觉剥夺性近视眼视网膜(尤其是内核层及内丛状层)中Smad1的表达呈下调趋势, Smad1极有可能通过转导视网膜信号参与了近视发生发展过程。     

Decreased proteoglycan synthesis associated with form deprivation myopia in mature primate eyes

PURPOSE: The rate of proteoglycan synthesis was measured in the scleras of adolescent marmosets that had undergone monocular form deprivation to characterize the scleral extracellular matrix changes associated with the development of myopia in a mature primate. METHODS: Form deprivation myopia was induced in adolescent marmosets by unilateral lid suture for an average of 108 days. After the lids were reopened, the axial lengths and refractions were measured at intervals for up to 39 weeks. At the end of the study period, sclera were isolated and immediately radiolabeled with 35SO4 in organ culture. Proteoglycan synthesis rates were determined by measurement of 35SO4 incorporation into cetylpyridinium chloride-precipitable glycosaminoglycans after digestion of the scleral samples with proteinase K. Collagen content was determined by measurement of total hydroxyproline in scleral digests. Newly synthesized proteoglycans were separated on a Sepharose CL-4B molecular sieve column and identified by their core proteins by Western blot analyses. RESULTS: Lid suture resulted in myopia due to a significant increase in vitreous chamber depth. After Sepharose CL-4B chromatography, newly synthesized scleral proteoglycans isolated from normal, form-deprived, and contralateral control eyes, resolved into one major peak that eluted in the position of decorin, a small chondroitin-dermatan sulfate proteoglycan. After digestion of the major peak with chondroitinase ABC, an approximately 45-kDa core protein was detected by Western blot analyses, confirming the presence of decorin. Form deprivation resulted in a significant reduction in the rate of proteoglycan synthesis in the posterior sclera (-43.55%, P < or = 0.001). Proteoglycan synthesis was also significantly reduced in the posterior sclera of form-deprived eyes relative to total collagen content (-36.19%, P < or = 0.01) and was negatively correlated with the rate of vitreous chamber elongation in the deprived eye (r2 = 0.779, P < or = 0.05). CONCLUSIONS: Significant extracellular matrix remodeling occurs in the posterior sclera of the adolescent primate eye during vitreous chamber elongation and myopia development. The negative correlation between vitreous chamber elongation rates and the synthesis rates of decorin in form-deprived eyes suggests that proteoglycan synthesis within the posterior sclera plays a role in the regulation of ocular size and refraction in the adolescent marmoset.     

哌仑西平影响豚鼠实验性近视眼的研究

目的 探讨玻璃体腔注射选择性M_1受体拮抗剂哌仑西平对豚鼠形觉剥夺性近视眼视网膜、脉络膜、巩膜和虹膜-睫状体组织中M_1和M_4受体mRNA表达的影响.方法 采用随机分组设计的实验研究.1～2周龄的三色豚鼠24只,随机分为4组:正常对照组(N)6只,单纯形觉剥夺组(FDM)6只,药物对照组(S)6只,哌仑西平组(P)6只,均以左眼为实验眼,P组隔日玻璃体腔注射500μg哌仑西平;S组隔日玻璃体腔注射生理盐水.21 d后结束实验.提取各组眼视网膜、脉络膜、巩膜和虹膜-睫状体组织总RNA,半定量RT-PCR检测M_1和M_4亚型mRNA表达变化.3组间数据的比较采用单因素方差分析和Tukey post hoc检验.结果 21d时,FDM与N组相比较,FDM组眼轴相对延长0.29mm,产生相对近视-4.92 D,差异有统计学意义(P<0.001).P组与S组相比较,眼轴相对减少了0.30 mm,产生+0.88 D的相对远视,差异均有统计学意义(P<0.001).S组与FDM之间屈光度数的差异无统计学意义;而眼轴变化有统计学意义(S组相对延长0.08 mm,而FDM组相对延长0.29/mm).半定量PCR结果显示:P组与S组相比较,其视网膜、脉络膜和虹膜睫状体组织M_1和M_4亚型mRNA的表达差异无统计学意义(P>0.05).而在后极部巩膜组织,M_1和M_4亚型mRNA的表达较药物对照组显著性增加(P<0.05),其中M_1受体表达增加19.16%,M_4受体表达增加64.29%.结论 哌仑西平能够有效抑制豚鼠形觉剥夺近视的发展.巩膜组织M_1和M_4亚型及其胆碱能通路可能参与M受体拮抗剂抑制近视发展.     

Comparison of form-deprived myopia and lens-induced myopia in guinea pigs

AIM: To study the efficacy difference between form-deprived myopia (FDM) and lens-induced myopia (LIM), the degree of myopia, axial length and pathological changes of the posterior sclera from guinea pigs were evaluated.METHODS: Four-week pigmented guinea pigs were randomly assigned into 3 groups, including normal control (n=6), FDM group with monocular cover (n=11) and LIM group with monocular -7D lens treatment (n=11). FDM group was form-deprived while LIM group was lens-induced for 14 d. Refractive error and axial length were measured prior to and post treatment, respectively. Morphological changes of sclera were examined using both light and electronic microscopes.RESULTS: After 14d treatment, refractive errors for FDM group and LIM group were -3.05±0.71D and -2.12±1.29D, respectively, which were significantly more myopic than that of normal controls and fellow control eyes (P<0.01). As for axial length, it was 7.93±0.03 mm for FDM group and 7.89±0.06 mm for LIM group, which were significantly longer than both normal and fellow controls (P<0.01). With respect to both refractory error and axial length, the differences between FDM group and LIM group were not significant (P>0.05). Under light microscope, both FDM group and LIM group showed thinned sclera, disarrangement of fibrosis and enlarged disassociation between fibers. Consistently, ultrastructural examination showed degenerated fibroblasts and thinned fibers in posterior sclera.CONCLUSION:Following two weeks of myopia induction in guinea pigs, with regard to the degree of myopia, axial length and pathological alterations, there was no significant difference between FDM and LIM models. Therefore, FDM and LIM are equally effective and useful as a model of experimental myopia and guinea pigs are ideal animals for induction of experimental myopia because their high sensitivity to both form-deprivation and lens-induction.     

鸡形觉剥夺性近视眼后极部巩膜胰岛素样生长因子-1/胰岛素样生长因子-2 mRNA表达的动态变化

目的观察鸡形觉剥夺性近视眼后极部巩膜胰岛素样生长因子(insulinlikegrowthfactor,IGF)IGF1/IGFmRNA表达水平的变化,探讨IGF1/IGF2在实验性近视眼发病中的作用。方法取孵化1d的白色来亨鸡36只,右眼为遮盖眼,左眼为自身对照眼。测量实验前以及单眼遮盖后第7天、第14天、第21天时实验眼和对照眼的屈光度和眼轴长度,并检测不同遮盖时间时鸡眼后极部巩膜IGF1/IGF2mRNA的表达水平。结果随着遮盖时间的延长,IGFmRNA在试验眼和对照眼后极部巩膜的表达水平均升高,但两者之间差异无显著性(P>0.05);实验眼后极部巩膜的IGF2mRNA表达水平在遮盖7d后即明显高于对照眼(P<0.001),随着遮盖时间的延长,实验眼后极部巩膜IGF2mRNA表达水平明显升高,而对照眼则逐渐下降,两者的差值明显增大。结论形觉剥夺可能通过上调鸡眼后极部巩膜IGF2mRNA表达水平,影响巩膜的发育和重塑,从而导致近视的形成。     

Modulation of scleral DNA synthesis in development of and recovery from induced axial myopia in the tree shrew

Visually modulated scleral extracellular matrix remodelling is associated with the development of, and recovery from, induced axial myopia in the tree shrew, a commonly used mammalian model of refractive error development. The involvement of scleral cell proliferation in this process was investigated using [3H] thymidine. Tree shrews were monocularly deprived of pattern vision, using translucent occluders, or the retinal image was optically defocused, using negative lenses, over a period of 5 days. A further group was monocularly deprived for 5 days, then allowed 3 days of binocular recovery. A control group of binocularly open animals was employed to establish normal parameters. On the final day of the experimental period, [3H] thymidine was administered by intraperitoneal injection, then optical and biometric measures were taken and tissue samples collected for assay. Incorporation of [3H] thymidine into cellular DNA was measured in proteinase K digests, following precipitation with trichloroacetic acid. After 5 days, significant amounts of myopia were present in the treated eyes of both form-deprived [-7. 0+/-0.7 Dioptres (D), group mean+/-s.e.m.; P<0.01] and lens-defocused animals (-6.2+/-0.9 D;P<0.01). After 3 days of recovery, 50% of the refractive error had been compensated for, predominantly through shortening of the vitreous chamber in the treated eye. Reduced levels of [3H] thymidine incorporation were observed in sclera from both groups of myopic animals (form-deprived, -34.3+/-9.9%;P<0.05 and lens-defocus, -32.8+/-4.5%;P<0.005). Increased levels of [3H] thymidine incorporation were found in the sclera of recovering animals (+144.0+/-43.2%;P<0.05). The results show that changes in regulation of cell proliferation, during the development of myopia, are visually mediated and inversely related to the direction of change in ocular size. This implies that alterations in the scleral fibroblast population are involved in the modulation of scleral matrix turnover during myopia development.     

Protective effects of riboflavin-UVA-mediated posterior sclera collagen cross-linking in a guinea pig model of form-deprived myopia

AIM:To evaluate the effect of posterior sclera collagen cross-linking induced by riboflavin-ultraviolet A(UVA)on form-deprived myopia in guinea pigs.METHODES:Twenty-five pigmented guinea pigs of 3-week-old were randomly assigned into 4 groups that included normal control(NOR,n=7),form-deprived(FDM,n=7),normal with riboflavin-UVA cross-linking(NOR+CL,n=5)and form-deprived with cross-linking(FDM+CL,n=6).The NOR+CL group and the FDM+CL group received the riboflavin-UVA induced cross-linking at day 0.FDM was induced by monocularly deprived with facemask in the right eyes.The refraction,axial length and corneal curvature were measured by retinoscopy,A-scan and keratometer respectively in scheduled time points(day 0 and 1,2,3,4 wk after form-deprivation).At the end of 4 weeks’experiment,stress-strain tests of sclera were measured and morphological changes of sclera and retina were examined.RESULTS:After 4 wk,the interocular difference of refractive error were-0.11±0.67,-2.93±0.56,1.10±0.58,and-1.63±0.41 D in the NOR,FDM,NOR+CL,and FDM+CL groups respectively.Mixed-effect linear model revealed significant effect of FDM(P<0.01)and CL(P<0.001).Also,after 4 wk,the interocular difference of axial length were 0.01±0.04,0.29±0.07,-0.13±0.06,and 0.11±0.05 mm in the NOR,FDM,NOR+CL,and FDM+CL group.Mixedeffect linear model revealed significant effect of FDM(P<0.001)and CL(P<0.01).As for corneal curvature,significant interocular difference have not found between any of the two groups.At the end of this experiment,the ultimate stress and elastic modulus were found significantly increased in both CL groups.But no difference was found in the groups without cross-linked.There was no abnormality observed in the retina and RPE cells of the treated eyes.CONCLUSION:The posterior sclera collagen crosslinking induced by riboflavin-UVA can slow down the progress of myopia and increase the sclera biomechanical strength in the guinea pig model of form-deprived myopia.