The Management And Molecular Genetics Mechanism Of Retinopathy Of Prematurity

PurposeThis study investigated the effect of anti-neovascularization treatment for retinopathy of prematurity(ROP)and the timing and risk factors of recurrent ROP after treatment.Meanwhile,this study studied the profiles of inflammatory cytokines and extracelluar circulating mi RNAs in the plasma and aqueous humor of patients with ROP and explored the molecular genetics mechanism of ROP.Materials and methods(1)Fifty eyes of 50 infants treated with IVR monotherapy for Type 1 ROP were studied retrospectively.The mean follow-up time was 31 weeks after intravitreal injection of ranibizumab(IVR).Recurrent ROP(recurrence of extraretinal fibrovascular proliferation,EFP)was determined by Retcam wide-angle fundus photographs.Risk time of recurrence was estimated by Kaplan–Meier survival analysis with recurrence as the end point.Time-varying recurrence hazard rate was determined using the hazard function of life-table analysis.The risk factors of recurrence were explored by logistic regression analysis.(2)Thirty-nine eyes of 20 patients,who were treated for type 1 or milder ROP(zone 2 stage 3 ROP with preplus)at postmonstrual age(PMA)over 40 weeks,were studied retrospectively.Of these,nineteen eyes were treated primarily with IVR and20 eys were treated with laser ablation.The clinical characteristics of these eyes,their reponse to treatment,and rate of retreatment were analyzed.(3)Fifty samples of aqueous humor from eyes with threshold ROP or pre-threshold ROP were collected before IVR and 15 samples from eyes with congenital cataract were collected during surgery as a control.Concentrations of 27 cytokines including VEGF were determined by Bio-Plex Pro TM magnetic color-bead-based multiplex analysis system and compared among groups.(4)Six samples of plasma in patients with type 1 ROP and four aqueous samples from eyes with type 1 ROP were collected;six samples of plasma in preterm patients without ROP and four aqueous samples from eyes with congenital cataract were collected as controls.Extracellur circulating mi RNAs were exstracted.Profiles of mi RNAs were determined with Agilent Human mi RNA array and then compared between groups.Different mi RNAs and targeted cytokines and pathways were analyzed.Results:(1)Recurrence of ROP occurred in 32(64%)of 50 eyes at 7.9 ± 2.7 weeks after IVR.Most of recurrence(94%)occurred in 2.5 to 12 weeks following IVR treatment.The recurrence hazard rate reached its maximum at 8 weeks.Recurrence affecting the initial site of EFP occurred significantly earlier than recurrence only at the new vascular advancing edge(4.5±1.4 weeks versus 9.1±2.0 weeks after IVR,P < 0.001).The independent risk factors of recurrence included extensive retinal neovascularization(P = 0.005)and oxygen requirement after IVR(P = 0.016).(2)Of the 39 eyes,30 eyes(77%)had prominent fibrotic membranes,14 eyes(36%)had wide stage 3 ridges,and 20 eyes(51%)had hemorrhage on the ridge.Seventeen eyes with type 1 ROP(44%)required further treatment.Of the 17 eyes,there were 11 eyes developing ROP persistence and two eyes having ROP recurrence after primary IVR,and there were two eyes having ROP recurrence and two eyes developing retinal detachment after primary laser ablation.(3)The aqueous humor concentration of VEGF was highest with threshold ROP,middle with prethreshold ROP,and the lowest with the control(986.9±475.7,353.7±190.3,and 53.2±44.5,P<0.001).In pre-threshold ROP group,samples having VEGF>300pg/ml were collected between 34 and 38 PMA weeks;in threshold ROP group,samples with VEGF>850pg/ml were collected between 34 and 37 PMA weeks.Concentration of IFN-g,MIP-1a,IL-12,IL-10 with threshold ROP was higer than that with prethreshold ROP.Higher concentration was also found in the ROP group with cytokines IL-7,eotaxin,IL-8,IL-1ra,while GM-CSF and FGF were lower with ROP group than control.The levels of IL-12,IFN-γ were found positively correlated with the level of VEGF(r = 0.900,and 0.914,P < 0.005).(4)Detection rate of mi RNAs was 6.0±1.4% in plasma and 10.7±3.0% in aqueous.There were 11 mi RNAs which were detected in top 30 mi RNAs both from plasma and aqueous.In plasma,there was no difference between mi RNAs profiles from ROP and control.In aqueous humor,seven mi RNAs were found(fold change>2,P<0.05)higher in ROP group: mi R-184,mi R-4687-3p,mi R-6090,mi R-6510-5p,mi R-6727-5p,mi R-7110-5p,and mi R-762.The differentially expressed mi RNAs mi RNAs may participate in the regulation networks of VEGF,FGF and FGF receptor,IFNγ receptor,and IL12 receptor,and also in multiple targeted genetic pathways.Conclusions:(1)Recurrence of Type 1 ROP should be carefully watched in a long term follow-up after IVR monotherapy,particularly in the first 12 weeks after IVR and for those with extensive retinal neovascularization or prolonged oxygen therapy.(2)Treatment-requiring ROP after 40 weeks PCA has tendency of fibrotic formation,associated with decreased potentials of vascular advancing;while zone II stage 3 ROP with preplus responds well to primary IVR or laser treatment,type 1 ROP is predisposed to ROP persistence and fibrotic proliferation after either treatment pattern.(3)VEGF level in aqueous correlates well with severity of ROP and ranks most during 34 to 38 PMA weeks.Other cytokines including IFN-γ,MIP-1α,MIP-1β,and IL-12 may also increase in more severe ROP.(4)Seven increased mi RNAs in aqueous from eyes with ROP were mi R-184,mi R-4687-3p,mi R-6090,mi R-6510-5p,mi R-6727-5p,mi R-7110-5p,mi R-762.The differentially expressed mi RNAs may underly molecular genetic mechanism of ROP by regulation of cytokines networks and genetic pathways.