蓝光对豚鼠离焦性近视进展的抑制作用及其视锥细胞密度变化机制

Inhibitory effect of blue light intervention on lens-induced myopia development in guinea pigs and the mechanism of cone density change

目的观察蓝光干预对光学离焦性近视豚鼠屈光发育的影响及其作用机制。方法选取普通级2周龄三色豚鼠48只,采用抛硬币法随机分成蓝光组和白光组,每组各24只。所有豚鼠右眼佩戴-5.00 D镜片建立光学离焦模型,为实验眼;左眼为自身对照,不予遮盖。实验前及实验开始后8周,采用带状光检影镜测量豚鼠屈光度,A型超声测量前房深度、晶状体厚度及眼轴长度,角膜曲率计测量角膜曲率半径。实验开始后8周,采用过量麻醉法处死豚鼠,取右眼眼球并分离视网膜,采用视网膜铺片免疫荧光染色观察豚鼠视网膜S及M视锥细胞密度;采用高效液相色谱分析法检测视网膜视黄酸表达;采用实时荧光定量PCR检测视网膜视黄酸受体(RAR-β)和巩膜中基质金属蛋白酶2(MMP-2)、组织金属蛋白酶抑制剂2(TIMP-2)及Ⅰ型胶原的表达;采用苏木精-伊红染色观察巩膜厚度变化。结果实验开始后8周,蓝光组实验眼较白光组实验眼出现(0.63±0.12)D相对远视,眼轴增长延缓(0.08±0.00)mm;蓝光组对照眼较白光组对照眼出现(0.42±0.09)D相对远视,眼轴增长延缓(0.08±0.00)mm;蓝光组实验眼较蓝光组对照眼近视加深(1.52±0.09)D,眼轴增长(0.06±0.00)mm;白光组实验眼较白光组对照眼近视加深(1.66±0.07)D,眼轴增长(0.13±0.00)mm,差异均有统计学意义(均P<0.05)。蓝光组豚鼠视网膜背侧和腹侧M视锥细胞密度小于白光组,背侧和腹侧S视锥细胞密度大于白光组,差异均有统计学意义(t=32.33、52.23、42.09、25.02,均P<0.05)。蓝光干预后近视延缓与腹侧S视锥细胞密度增加呈强正相关(r=0.95,P<0.01)。蓝光组视黄酸含量、RAR-β和MMP-2相对表达量较白光组减少,TIMP-2和Ⅰ型胶原相对表达量较白光组增加,差异均有统计学意义(t=18.73、7.45、3.72、6.19、9.03,均P<0.05)。蓝光组巩膜厚度为(125.0±7.8)μm,较白光组的(102.0±6.3)μm明显增厚,差异有统计学意义(t=26.93,P<0.05)。结论蓝光可抑制豚鼠离焦性近视进展;豚鼠屈光度的改变可能通过视网膜视锥细胞密度变化影响视网膜视黄酸及巩膜胶原的表达来实现。

Objective To observe the effects of blue light intervention on the development of optical defocus-induced myopia in guinea pigs and investigate its underlying mechanisms.Methods Forty-eight normal-grade two-week-old tricolor guinea pigs were randomly divided into a blue light group and a white light group,with 24 animals in each group.The right eye of guinea pigs was fitted with a-5.00 D lens to establish an optical defocus model as the experimental eye,while the left eye served as the control without any covering.Before the experiment and after 8-week intervention,the refractive power of guinea pigs was measured by streak retinoscopy.The anterior chamber depth,lens thickness,and axial length were measured by A-scan ultrasonography.Corneal curvature radius was determined using a keratometer.After 8-week intervention,the guinea pigs were euthanized through overanesthesia,and the right eyeballs were enucleated and the retinas were isolated.The density of S and M cone cells of the guinea pig retinal sections were observed via immunofluorescence staining.The expression of retinal retinoic acid was assessed by high-performance liquid chromatography.The expressions of retinoic acid receptor(RAR-β)in the retina and matrix metalloproteinase-2(MMP-2),tissue inhibitor of metalloproteinase-2(TIMP-2),and typeⅠcollagen in the sclera were detected by real-time fluorescence quantitative PCR.Changes in scleral thickness were observed through hematoxylin-eosin staining.The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.The study protocol was approved by the Ethics Committee of Eye and ENT Hospital of Fudan University(No.2022ETKLD10032).Results After 8 weeks of intervention,guinea pigs in the blue light group showed(0.63±0.12)D of relative hyperopia and a deceleration of axial elongation by(0.08±0.00)mm compared with the white light group in the right eye.In the left eye,guinea pigs in the blue light group showed(0.42±0.09)D of relative hyperopia and a deceleration of axial elongation by(0.08±0.00)mm compared with the white light group.The guinea pigs in blue light group showed(1.52±0.09)D of myopia in the right eye compared with the left eye,with an increase in axial elongation of(0.06±0.00)mm.The guinea pigs in white light group showed(1.66±0.07)D of myopia in the right eye compared with the left eye,with an increase in axial elongation of(0.13±0.00)mm,and the differences were statistically significant(all at P<0.05).The density of M cone cells was lower and density of S cone cells was higher in the blue light group in the dorsal and ventral sides of the retinal sections compared with the white light group,showing statistically significant differences(t=32.33,52.23,42.09,25.02;all at P<0.05).The deceleration of myopia progression in the blue light group was strongly positively correlated with the increase in S cone cell density on the ventral side(r=0.95,P<0.01).The expression levels of retinoic acid,RAR-β,and MMP-2 were decreased,and expression levels of TIMP-2 and typeⅠcollagen were increased in blue light group compared with the white light group,showing statistically significant differences(t=18.73,7.45,3.72,6.19,9.03;all at P<0.05).The scleral thickness in the blue light group was(125.0±7.8)μm,which was significantly thicker than(102.0±6.3)μm in the white light group(t=26.93,P<0.05).Conclusions Blue light intervention can inhibit the progression of defocus-induced myopia in guinea pigs.Refractive power changes in guinea pigs may be influenced by alterations in retinal cone cell density,retinoic acid expression,and scleral collagen expression.