摘要
目的探究糖皮质激素(GC)致高眼压(OHT)后视网膜神经节细胞(RGCs)损伤进程,以及雌二醇(E2)对OHT大鼠RGCs损伤的保护作用与机制。方法将36只(36眼)12周龄雄性SD大鼠随机分成空白对照组、GC-OHT组、OHT-E2组,每组各12只,其中GC-OHT组和OHT-E2组大鼠在结膜下注射GC,空白对照组大鼠结膜下注射相同体积生理盐水,造模后2周,OHT-E2组大鼠除注射GC外同时给予E2滴眼液滴眼。造模前及造模后1、2、3、4周对各组大鼠进行眼压测量,造模后4周,通过视网膜电图(P-ERG)、闪光视觉诱发电位(F-VEP)检测各组大鼠视力变化情况。摘取眼球,采用免疫荧光染色观察各组大鼠RGCs分布情况及数量。免疫组织化学、Western blot、实时荧光定量聚合酶链反应检测各组大鼠NOD样受体蛋白(NLRP3)、半胱氨酸天冬氨酸蛋白酶-1(Caspase-1)和gasdermin-D(GSDMD)蛋白及mRNA的相对表达情况。结果造模前各组大鼠眼压比较差异无统计学意义(P>0.05)。造模后1、2、3、4周,与空白对照组相比,GC-OHT组大鼠眼压均升高,差异均有统计学意义(均为P<0.01)。造模后3、4周,与GC-OHT组相比,OHT-E2组大鼠眼压均降低,差异均有统计学意义(均为P<0.01)。P-ERG及F-VEP检测结果显示,与空白对照组相比,GC-OHT组大鼠P50波及P1波幅均降低,差异均有统计学意义(均为P<0.05)。与GC-OHT组相比,OHT-E2组大鼠P50及P1波幅均升高,差异均有统计学意义(均为P<0.05)。免疫荧光染色结果显示,与空白对照组相比,GC-OHT组大鼠RGCs数量减少,差异有统计学意义(P<0.001)。与GC-OHT组相比,OHT-E2组大鼠RGCs数量增加,差异有统计学意义(P<0.001)。免疫组织化学、Western blot、实时荧光定量聚合酶链反应结果均显示,与空白对照组相比,GC-OHT组大鼠视网膜中NLRP3、Caspase-1和GSDMD蛋白及mRNA相对表达量均升高,差异均有统计学意义(均为P<0.05)。与GC-OHT组相比,OHT-E2组大鼠视网膜NLRP3、Caspase-1和GSDMD蛋白及mRNA相对表达量均降低,差异均有统计学意义(均为P<0.01)。结论GC诱导的OHT会致使RGCs发生焦亡,而E2可能通过抑制焦亡相关NLRP3/Caspase-1/GSDMD信号通路,减轻OHT大鼠的RGCs损伤。
Objective To explore the injury process of retinal ganglion cells(RGCs)after glucocorticoid(GC)-induced ocular hypertension(OHT),as well as the protective effect and mechanism of estradiol(E2)in RGC injury in rats with OHT.Methods A total of 36(36 eyes)12-week-old male Sprague-Dawley(SD)rats were randomly divided into the blank control group,the GC-OHT group,and the OHT-E2 group,with 12 rats in each group.Rats in the GC-OHT group and the OHT-E2 group were subconjunctivally injected with GC,while those in the blank control group were subconjunctivally injected with an equal volume of normal saline.Two weeks after modeling,in addition to being injected with GC,rats in the OHT-E2 group were also provided with E2 eye drops.Before modeling and 1,2,3,and 4 weeks after modeling,the intraocular pressure of rats in each group was measured.The visual acuity changes of rats in each group were detected by pattern electroretinogram(P-ERG)and flash visual evoked potential(F-VEP)4 weeks after modeling.After the eyeballs were removed,the distribution and number of RGCs in rats of each group were observed by immunofluorescence staining.Immunohistochemistry,Western blot,and real-time fluorescence-based quantitative polymerase chain reaction were used to detect the relative protein and mRNA expression levels of NOD-like receptor protein 3(NLRP3),cysteine aspartate protease-1(Caspase-1),and gasdermin-D(GSDMD)in rats in each group.Results There was no statistically significant difference in the intraocular pressure of rats in each group before modeling(P>0.05).Compared with the blank control group,the intraocular pressure of rats in the GC-OHT group increased 1,2,3,and 4 weeks after modeling,and the differences were all statistically significant(all P<0.01).Compared with the GC-OHT group,the intraocular pressure of rats in the OHT-E2 group decreased 3 and 4 weeks after modeling,and the differences were all statistically significant(all P<0.01).The P-ERG and F-VEP results showed that compared with the blank control group,the amplitudes of P50 and P1 waves of rats in the GC-OHT group decreased,and the differences were both statistically significant(both P<0.05).Compared with the GC-OHT group,the amplitudes of P50 and P1 waves of rats in the OHT-E2 group increased,and the differences were both statistically significant(both P<0.05).The immunofluorescence staining results showed that compared with the blank control group,the number of RGCs of rats in the GC-OHT group decreased,and the difference was statistically significant(P<0.001).Compared with the GC-OHT group,the number of RGCs of rats in the OHT-E2 group increased,and the difference was statistically significant(P<0.001).The results of immunohistochemistry,Western blot,and real-time fluorescence-based quantitative polymerase chain reaction showed that compared with the blank control group,the relative protein and mRNA expression levels of NLRP3,Caspase-1,and GSDMD in the retina of rats in the GC-OHT group all increased,and the differences were all statistically significant(all P<0.05).Compared with the GC-OHT group,the relative protein and mRNA expression levels of NLRP3,Caspase-1,and GSDMD in the retina of rats in the OHT-E2 group all decreased,and the differences were all statistically significant(all P<0.01).Conclusion GC-induced OHT can cause pyroptosis of RGCs,and E2 may alleviate the injury of RGCs in rats with OHT by inhibiting the pyroptosis-related NLRP3/Caspase-1/GSDMD signaling pathway.
作者
闫海波
王松涛
杨华
张欧阳
冀世超
杨瑞
王保君
YAN Haibo;WANG Songtao;YANG Hua;ZHANG Ouyang;JI Shichao;YANG Rui;WANG Baojun(Department of Ophthalmology,the First Affiliated Hospital of Xinxiang Medical University,Xinxiang 453003,Henan Province,China;Basic Medical College,Xinxiang Medical University,Xinxiang 453003,Henan Province,China)
出处
《眼科新进展》
CAS
北大核心
2025年第1期27-32,共6页
Recent Advances in Ophthalmology
基金
新乡市科技攻关计划项目(编号:GG2021030)
新乡医学院博士科研基金资助项目(编号:XYB-SKYZZ2015010-505119)。
