1.Protection of NEP1-40 on retinal cells following retinal ischemia reperfusion injury
Suoxin, LIU ; Xuehong, JU ; Shuna, YU ; Hao, WANG
Chinese Journal of Experimental Ophthalmology 2014;32(3):220-225
Background The retina ischemia reperfusion injury (RIRI) can lead to apoptosis,which is associated with many genes.It is very significant to explore the protection of drugs on RIRI-induced apoptosis.Objective This study was to explore the relationship between the expression of Nogo-A and the cell apoptosis as well as the therapeutic effect of NEP1-40 in RIRI retina.Methods The device of raising intraocular pressure (IOP)was used to elevate the IOP for 60 minutes and then restore the IOP to normal for the establishment of RIRI models.Seventy-eight SD rats were randomized to the normal group,RIRI group and NEP1-40 group.PBS of 5 ml/(kg · d)was injected intraperitoneally in the rats of the RIRI group,and NEP1-40 of 8 mg/(L · kg) was used in the same way in the NEP1-40 group.The rats were sacrificed in overdose anesthesia at 6 hours,12 hours and 1 day,2,3,7 days after RIRI to prepare the retinal specimens.The ultrastructure of rat retinas was examined under the transmission electron microscope.Cell apoptosis was assessed by the TUNEL method,and the apoptotic index (AI) was calculated.The expressions of Nogo-A protein and mRNA in rat retinas were detected by immunohistochemistry and semiquantitative reverse transcription PCR (RT-PCR).Results The ultrastructure of retinal cells were normal in the normal group.Retinal cell organelle dissolution,mitochondria swelling,cavitation,chromatin edge heterochromatin and apoptotic body were seen in the rats of the RIRI group from 12 hours through 7 days after injection.However,only the slight loose of outer membranous disk,inner and outer nuclear layer and retinal ganglion cells,the nucleus gap broadening,shortness of some mitochondrial cristae in the NEP1-40 group.TUNEL-positive cells appeared 6 hours after RIRI and reached peak in 1 day,and gradually declined after that in the RIRI group.A similar pattern was seen in the rats of the NEP1-40 group with a more mild manifestation.Significant differences were seen in the AI values among the different groups at various time points (Fgroup =100.850,P =0.000 ; Ftime =34.309,P =0.000),and the AI values were significantly higher in the RIRI group and NEP1-40 group compared with normal group;while the AI values in the NEP1-40 group was lower than those of the RIRI group (all at P<0.05).The expressions of Nogo-A protein and mRNA showed a coincident pattern with apoptosis procedure,with a gradually elevated level from 6 hours through 7 days after RIRI and a peak in 2 days,and those in the NEP1-40 group were weaker in comparison with the RIRI group in various time points.Significant differences were detected in the expression of Nogo-A protein and the expression of Nogo-A mRNA among different groups and various time points(protein:Fgroup =164.139,P =0.000;Ftime =21.772,P =0.000.mRNA:Fgroup =93.889,P =0.000 ; Ftime =6.349,P =0.000).Conclusions Nogo-A probably plays an important role in RIRI.NEP1-40 can protect retinal cells from apoptosis following RIRI through down-regulating the expression of Nogo-A.