Protective effect of neuroglobin on retinal ganglion cell in glaucoma mice model / 中华实验眼科杂志

ABSTRACT

Background Neuroglobin (Ngb) is a newly discovered member of globin superfamily.It is thought to regulate cell survival under hypoxia or oxidative stress condition.Ngb is expressed at a high level in retinal neuron,suggesting that retina may be one of important functional sites of Ngb.Objective The aim of this study was to investigate the protective role of endogenous Ngb on retina ganglion cells (RGCs) following chronic high intraocular pressure(IOP)in mice and the underlying mechanisms.Methods This study included the in vitro and in vivo experiment.RGCs derived from adult C57BL/6J wild type(WT) mice and Ngb-transgenic(Ngb-Tg) mice which cultivated by our laboratory were incubated with 5.0,7.5,10.0 mmol/L glutamic acid for 3 days.RGCs survival rate was calculated for the ration of dead and survival cells using a double labeling kit to evaluate the influence of Ngb on RGCs survival rate in the addition of glutamic acid.Chronic ocular hypertension models were established by injection of fluorescent microballon(MB) (10 μm)into the anterior chamber of WT mice and Ngb-Tg mice,The mice were divided into WT control group(n=18),Ngb-Tg control group(n=30),WT+MB single injection group(n=38),Ngb-Tg+MB single injection group (n =38),WT+MB twice injection group (n =6) and Ngb-Tg + twice injection group (n=6).In addition,WT+PBS injection group (n =6) and Ngb-Tg+ PBS injection group (n =6) were designed as negative controls to identify if it can affect IOP or not.The mice were sacrificed on 0 day(control group),3 days and 1,4,8 weeks followed the MB anterior chamber injection.Real-time PCR,Western blot and immunoytochemistry were used respectively for the analysis of the expressions of Ngb mRNA and protein in mouse retina,and the survival rates of RGCs were compared between the two types mice.Dihydroethidium (DHE) in retina was detected after cardiac perfusion and ATP level in mouse retina homogenate was analyzed.Results The RGCs survival rate was significantly higher in Ngb mice compared with WT mice in 5.0,7.5 and 10.0 mmol/L glutamic acid treatcd groups (t =2.810,3.020,3.110,P＜ 0.01).IOP of WT+ MB single injection group and Ngb-Tg+ MB single injection group were elevated in comparison with the WT control group and WT+PBS injection group and the high IOP remained for 4 weeks.Ngb level was raised in the WT+MB single injection group on the third day following injection,but the Ngb concentration remained a high level in the Ngb-Tg mice during the period of the experiment duration.The RGCs apoptosis rate was elevated both in the WT mice and Ngb-Tg mice 1,4,8 weeks after injection of MB,however,the cell apoptosis rate was higher in WT mice than that of Ngb-Tg mice(P＜0.05).DHE content in retina in the Ngb-Tg+MB single injection group was significantly lower than that in the WT+MB single injection group (t =3.212,P=0.008),and ATP content in retina was elevated in the Ngb-Tg+MB single injection group compared with WT+MB single injection group(t =2.864,P＜0.01).Conclusions It is suggested that Ngb might be a neuroprotective molecules against RGCs death by decreasing oxidative stress and improving mitochondria function.