NMNAT1缺失对小鼠视网膜双极细胞和无长突细胞的影响  

Effect of nicotinamide mononucleotide adenylyltransferase 1 deletion on retinal bipolar cells and amacrine cells in mice

作　　者：曹瑾 吴彩霞 毛建华 CAO Jin;WU Caixia;MAO Jianhua(Department of Ophthalmology,Xianning Central Hospital,Xianning 437100,Hubei Province,China)

机构地区：[1]咸宁市中心医院眼科,湖北省咸宁市437100

出　　处：《眼科新进展》2022年第5期359-363,368,共6页Recent Advances in Ophthalmology

摘　　要：目的探索烟酰胺单核苷酸腺苷转移酶1(NMNAT1)缺失对视网膜神经元细胞结构及功能的损伤作用。方法利用NMNAT1^(fl/fl)小鼠与Six3启动子下表达Cre重组酶的转基因(Six3-Cre)小鼠杂交,获得NMNAT1条件基因敲除小鼠及其同窝对照小鼠,实时荧光定量PCR和Western blot进行检测。在基因敲除小鼠出生后第0、4、10、30天(P0、P4、P10、P30)取材,制作冰冻切片,分别进行HE染色测量视网膜厚度,免疫荧光染色鉴定受损细胞类型以及检测恢复蛋白和视紫质表达水平。结果与同窝对照小鼠相比,基因敲除小鼠在P0时视网膜并没有明显的形态差异,视网膜厚度比较差异无统计学意义(P>0.05);到P4时基因敲除小鼠表现出视网膜内层和外层中的分层破坏和大规模细胞死亡,基因敲除小鼠P4时距视神经500μm、1000μm、1250μm的视网膜厚度均明显低于同窝对照小鼠(均为P<0.05),而距视神经1750μm的视网膜厚度尚未受影响(P>0.05);基因敲除小鼠P10时距视神经500μm、1000μm、1250μm、1750μm的视网膜厚度均低于同窝对照小鼠(均为P<0.05)。基因敲除小鼠全部视网膜内外层的退化在P30时几乎完成。此外,与同窝对照小鼠相比,P4时基因敲除小鼠视网膜神经节细胞、双极细胞和无长突细胞的数量差异均没有统计学意义(均为P>0.05),P10时视网膜神经节细胞没有显著差异(P>0.05),基因敲除小鼠双极细胞、无长突细胞分别减少了约75%、51%(均为P<0.05)。与同窝对照小鼠相比,基因敲除小鼠在P4时视网膜几乎不表达恢复蛋白和视紫质(均为P<0.05)。结论NMNAT1缺失主要导致视网膜双极细胞和无长突细胞损害以及恢复蛋白和视紫质缺失。Objective To investigate the damage effect of nicotinamide mononucleotide adenylyltransferase 1(NMNAT1)deletion on the structure and function of retinal neurons in mice.Methods NMNAT1^(fl/fl) mice were hybridized with Six3-Cre transgenic mice expressing Cre recombinase under the Six3 promoter to obtain NMNAT1 conditional knockout mice,and mice in the same litter were selected as controls.They were detected by real-time quantitative polymerase chain reaction and Western blot.Frozen sections of knockout mice were made at 0,4,10,and 30 days after birth(P0,P4,P10,and P30).HE staining was performed to measure retinal thickness,and immunofluorescence staining was performed to identify damaged cell types and detect the expression levels of recoverin and rhodopsin.Results Compared with the control mice,the retina of the knockout mice had no obvious morphological variation at P0,and the retinal thickness also had no significant change(P>0.05).However,at P4,knockout mice exhibited layered damage and massive cell death in inner and outer retinal layers,the retinal thicknesses at 500μm,1000μm and 1250μm away from the optic nerve in the knockout mice were significantly lower than those in the control mice(all P<0.05),while the retinal thickness at 1750μm away from the optic nerve did not change(P>0.05).The retinal thicknesses at 500μm,1000μm,1250μm,and 1750μm from the optic nerve in knockout mice were significantly lower than those in the control mice at P10(all P<0.05).The degeneration of both the inner and outer layers of the retina in knockout mice was almost finished at P30.Compared with the control mice,there was no significant difference in the numbers of retinal ganglion cells,bipolar cells,and amacrine cells in knockout mice at P4(all P>0.05),while at P10,there was no significant difference in the number of retinal ganglion cells in knockout mice(P>0.05),but the numbers of bipolar cells and amacrine cells decreased by about 75%and 51%,respectively(both P<0.05).Knockout mice almost completely lack the expressio

关 键 词：烟酰胺单核苷酸腺苷转移酶1 双极细胞 无长突细胞 恢复蛋白 视紫质 

分 类 号：R774.1[医药卫生—眼科]