溶血卵磷脂酰基转移酶1自发突变小鼠视网膜组织和闪光视网膜电图改变

Effect of lysophosphatidylcholine acyltransferase-1 deficiency on the murine retinal structure and electroretinograms

目的 观察溶血卵磷脂酰基转移酶1（LPCAT1）自发突变小鼠视网膜组织和闪光视网膜电图（F-ERG）改变情况.方法 Lpcat1基因自发突变纯合子的rd11新生小鼠60只（实验组）和与之同龄的野生型C57BL/6J小鼠60只（对照组）纳入实验.采用免疫荧光法检测两组小鼠视网膜组织中LPCAT1的表达情况.出生后3、6、9d和2、4、6、8周,行视网膜石蜡切片观察视网膜显微结构;行全视野F-ERG检测,记录暗视杆体反应和明视锥体反应.结果 实验组小鼠视网膜组织中LPCAT1呈阴性表达;对照组小鼠视网膜组织中LPCAT1呈阳性表达,主要分布于视网膜光感受器内节,神经节细胞层可见少量表达.光学显微镜观察发现,实验组小鼠于出生后9d左右视网膜外核层已基本形成,其光感受器细胞核层数随年龄增长而逐渐减少.对照组小鼠视网膜显微结构正常.F-ERG检测结果显示,实验组小鼠出生后2、4周暗视杆体系统反应存在,呈降低趋势,出生后6～8周暗视杆体系统反应消失;对照组小鼠各时间点暗视杆体系统反应正常.出生后2周,实验组和对照组小鼠b波振幅分别为（72.8±15.6）、（105.2±21.1） μV,实验组较对照组降低,但差异无统计学意义（t=-2.760,P=0.025）.出生后4周,实验组和对照组小鼠b波振幅分别为（20.6±6.4）、（231.8±32.0）μV,实验组较对照组明显降低,差异有统计学意义（t=-14.471,P=0.000）.实验组小鼠出生后2、4、6周明视锥体系统反应存在,呈降低趋势,出生后8周明视锥体系统反应消失;对照组小鼠各时间点明视锥体系统反应正常.出生后2、4周,实验组小鼠b波振幅分别为（46.8±7.2）、（78.0±8.2）μV;对照组小鼠b波振幅分别为（42.8±6.4）、（91.4±9.4） μV.两组比较,差异均无统计学意义（t=0.930、-2.401,P=0.379、0.043）.出生后6周,实验组和对照组小鼠b波振幅分别为（17.2±2.0）、（116.2±12.9） μV,实验组较对照组明显降低,差异有统计学意义（t=-17.008,P=0.000）.结论 LPCAT1自发突变小鼠视网膜外层光感受器细胞核层数随年龄增长而逐渐减少,F-ERG暗视杆体和明视锥体反应异常.

Objective To observe the effect of lysophosphatidylcholine acyltransferase-1 （LPCAT1）deficiency on the structure and electrophysiology of the murine retina.Methods Rd11 mice （Lpcat1 homozygous mutant,n =60） and wild-type C57BL/6J mice （n =60） were used in this study.Immunohistochemistry was performed to determine the expression of LPCAT1 in the mouse retina.Retinas of rd11 mice and age-matched control mice at postnatal 3,6,9-day and 2,4,6,8-week were paraffin embedded,sectioned and hematoxylin eosin stained,and full-field electroretinograms （F-ERG） were also recorded at these time points.Statistics were based on independent samples t-test.Results LPCAT1 was absent in rd11 mice retina.In wild-type C57BL/6J mice retina,LPCAT1 was expressed most strongly in the inner segment of photoreceptor cells,weak in the ganglion cell layer.Rd11 mouse exhibited retinal degeneration and eventual photoreceptor cell loss.Retinas of rd11 mice showed nearly half of the photoreceptor cells missing around postnatal week 4,and by 6-week after birth only two layers of nuclei remained.At postnatal week 8,nearly all photoreceptor cells were lost.Dark-adapted F-ERG showed reduced rod-driven response at 2 and 4 weeks of age,which was flattened by 6 and 8 weeks of age.By 2 weeks of age,no significant difference was found in b-wave amplitude between rd11 eyes [（72.8±15.6） μV]and C57BL/6J eyes [（105.2±21.1） μV] （t=-2.760,P=0.025）.Compared with age-matched control mice [（231.8±32.0）μV],rod response of rd11 mice [（20.6±6.4） μV] decreased obviously at postnatal week4 （t=-14.471,P=0.000）.Cone response was nearly normal at 2 and 4 weeks of age but substantially reduced at 6 weeks of age,which was flattened by 8 weeks of age.At 2 and 4 weeks after birth,no significant difference was found in b-wave amplitude between rd11 eyes [（46.8 ± 7.2）,（78.0 ±8.2） μV] and C57BL/6J eyes [（42.8±6.4）,（91.4±9.4） μV] （t=0.930,-2.401; P=0.379,0.043）.Compared with age-matched control mice [（116.2 ± 12.9） μV],cone response of rd11 mice [（17.2 ± 2.0） μV]decreased obviously at postnatal week 6 （t =-17.008,P =0.000）.Conclusion The layers of photoreceptor cells nuclei in rd11 mice decreases with age,and its F-ERG reflection is unusual.