豚鼠形觉剥夺性近视眼视网膜蛋白激酶C的变化

Change in retinal protein kinase C in the development of form-deprivation myopia in the guinea pig

摘 要：目的 研究豚鼠形觉剥夺性近视眼形成过程中视网膜蛋白激酶C（PKC）的变化.方法 选取3周龄雄性三色豚鼠48只,用半透明眼罩遮盖豚鼠右眼建立近视眼模型,分为遮盖3、7、14 d组,每组16只豚鼠.每组中8只豚鼠遮盖右眼作为实验眼.以对侧未遮盖眼作为自身对照,剩余8只豚鼠作为正常对照.按预定时间观察豚鼠角膜曲率半径、屈光度和眼轴长度的变化,然后处死豚鼠,取视网膜,采用非放射性方法测定视网膜PKC活性,免疫组化染色定位分析PKC蛋白在视网膜的表达情况.各组间角膜曲率半径、屈光度、眼轴长度、视网膜PKC活性及免疫反应强度比较,采用单因素方差分析;组内左、右眼之间的比较采用t检验：视网膜PKC活性与PKC蛋白免疫反应强度的关系采用相关分析.结果 眼罩遮盖能诱导豚鼠遮盖跟眼轴延长、近视形成,且近视程度随遮盖时间的延长而加深.与对照组比较,各组遮盖眼视网膜PKC活性均升高（P〈0.05）.眼罩遮盖3 d后,遮盖眼视网膜PKC活性升高至（0.28±0.08）units/ml,遮盖7 d时PKC活性进一步升高至（0.43±0.10）units/ml,遮盖14 d时降低为（0.32±0.07）units/ml.免疫组化染色结果显示,PKC蛋白表达于视网膜神经节细胞和内核层细胞,呈棕黄色或棕褐色颗粒.与正常对照眼及自身对照眼比较,遮盖眼视网膜内核层细胞PKC蛋白表达上调（P〈0.05）,神经节细胞PKC蛋白表达无变化.遮盖眼视网膜内核层细胞PKC蛋白表达的变化趋势与PKC活性一致,两者之间有明显的相关关系（r=0.994,P=0.000）.结论 视网膜PKC活性升高参与了豚鼠形觉剥夺性近视的形成.

Objective To investigate the change in retinal protein kinase C （PKC） in form-deprivation myopia in the guinea pig. Methods Guinea pigs were randomly divided into three groups： 3-day deprivation, 7-day deprivation and 14-day deprivation. The right eye of 8 guinea pigs was deprived, and other eyes used as a control in each group. Form-deprivation myopia was induced with translucent eye shields on the right eye. Following a schedule, ocular biometric measurements （comeal curvature radius, refraction and axial length） were performed. The guinea pigs were sacrificed and retinal samples were obtained. Retinal PKC activity was detected by non-radioactive methods, and protein was evaluated by immunohistochemical staining. Results The deprived eyes in the guinea pigs became myopic with an elongation of the ocular axis dependent on eye occlusion. As deprivation time increased, there was an increase in the severity of myopia. Compared to the control eyes, retinal PKC activity was significantly up-regulated in the deprived eyes of each group （P〈0.05）. Retinal PKC activity was elevated after 3 days of occlusion [（0.28 ±0.08）units/ml, P〈0.05], and reached a maximum after 7 days of occlusion [（0.43±0.10）units/ml, P〈0.05]. Subsequently, a decline in retinal PKC activity was observed after 28 days of occlusion [（0.32±0.07）units/ml, P〈0.05]. PKC protein was expressed in the retinal ganglion cells and inner nuclear layer. PKC protein was up-regulated in the inner nuclear layer of the retina in the deprived eyes （P〈0.05）, but there was no change in ganglion cells （P〉0.05）. The tendency for a change in retinal PKC activity coincided with the protein in the inner nuclear layer of the retina in the deprived eyes. There was a significant correlation between the intensity of PKC immunoreactivity in the inner nuclear layer and retinal PKC activity in the deprived eyes （r=0.994, P=0.000）. Conclusion The increase in retinal PKC activity played an important role in the development of form-deprivation myopia in guinea pigs.