| Background:Retinal neovascularization（RNV）is the main pathologic feature of severe retinopathy.Abnormal growth of blood vessels and associated vascular leakage in diabetic retinopathy,retinopathy of prematurity and vascular occlusions are major causes of vision loss. For suchdiseases, the most widely used anti-VEGF treatment and other methods can not inhibitangiogenesis efficiently. And even vitrectomy is really no choice at all when complicationssuch as retinal detachment occurs. The formation of new blood vessels of retina is a verycomplicated process. In this process, adenosine plays an important role.Researches show thatadenosine or its analogues can increase VEGF, IGF-1, bFGF, IL-8, Ang-2angiogenic factors,etc and induce angiogenesis by inhibit the angiogenesis factor like TNF-a. Taking all theseinto consideration, we conclude that adenosine, as wide spread in vivo physiologicalconditioning, may play an important triiger role in retinal angiogenesis. Therefore, it is ofgreat significance to inhibit RNV from the source.Objective：To verify the role of "adenosine homeostasis " in retinal angiogenesis and provide newresearch directions and therapeutic targets for the pathogenesis of retinal neovascular disease,the present study investigated the relationship between adenosine levels in vitreous of patientswith proliferative diabetic retinopathy (PDR) and angiogenesis and determined the inhibitoryeffect of adenosine on retinal angiogenesis by blocking adenosine in animal experiment.Methods：1. We selected30patients who underwent vitrectomies in the Ophthalmology at DapingHospital. According to the patient whether there were any retinal neovascularization anddivided them into two groups. Sixteen patients (16eyes) were in group diseases of retinalneovascularization (PDR group). Fourteen patients (14eyes) were in control Group, whichwas without revascularization in retinal. In this group, rhegmatogenous retinal detachment,idiopathic macular hole and macular epiretinal membrane were contained. The levels of adenosine in vitreous were measured by HPLC. The expression of CD73in preretinalmembranes was measured by western blot.2. C57BL/6J mice were used in these experiments to build a mouse model of oxygen-induced retinopathy(OIR). Mice are exposed to75%oxygen tension from postnatal day P7toP12. They were then returned to normal air and maintained for another5days (P17).Normoxia control mice of age identical with the hyperoxia group were maintained at room air.Retinal neovascularization was observed by fluorescence angiography and was quantified bycounting the endothelial nuclei protruding into the vitreous cavity after HematoXylin-eosinstaining. High performance liquid chromatograph (HPLC) and westem blot analysis wereused to determine retinal adenosine content and CD73protein levels at Postuatal days(P)13,15, and17.From P12to P17, the pups received daily intraperitoneal injections of thenonselective adenosine receptor antagonist xanthine amine congener (XAC). The antagonistwas administered at dose of30.0ug/kg of body weight after Preliminary experiments. Vehiclecontrol mice were injected with0.1%dimethyl sulfoxide (DMSO) in isotonic saline used todissolve the adenosine receptor antagonists.Results：1. The levels of adenosine in group PDR was higher than that in group control [(63.50±11.60) μmol/L vs (30.34±6.41)μmol/L；P=0.000].2. There is no correlation between the age,course of diabetes, fasting plasma glucoseleves in pre-operation of PDR patients and the concentration of adenosine in vitreous（r=0.255，P=0.340；r=0.318，P=0.230；r=0.341，P=0.197）.3. The expression of CD73in group PDR was higher than that in group control(P=0.002).4. The amount of E-ADO in group PDR was higher than that in group control (P=0.002)at20min after reaction.5. In fluorescence angiograms,irregular neovascularization and fluoresce in leakagewere observed surrounding the unperfused areas in the OIR group. Normal control groupmice did not see obvious retinal vascular anomalies.The OIR group had,on average,41.7±6.1neovascular nuclei protruding into the vitreous body,while similar nuclei was2.7±1.9in the control group.6. In the The adenosine content at P13, P15, P17days of normal control group mice were r（0.048±0.008）ng/mg retinal、（0.069±0.004）ng/mg retinal、（0.52±0.007）ng/mgretinal, respectively；In the The adenosine content at P13, P15, P17days of OIR group micewere（0.839±0.043）ng/mg retinal、（1.066±0.108）ng/mg retinal、（1.369±0.107）ng/mgretinal, respectively. There were significant difference of retinal adenosine at P13,15,and17between the two group (P <0.01).7. OIR model group compared with normal control group,CD73protein in P13, P15,P17days OIR retinal were expression (P=0.003; P=0.001; P=0.000). Compared to thenormoxic group,there was significant increases in retinal CD73protein levels in the OIRgroup at P13,15,and17（P=0.003；P=0.001；P=0.000）.8. Fluorescence imaging results show decreased number of retinal neovascularization inXAC injected mice in comparson with the DMSO injection group. Retinal tissue HE stainingresults show: significantly decreased breakthrough injection retinal membrane of vascularendothelial nuclei number in SAC group compared with DEMOS group [(17.3+3.9) vs.(38.1+5.6); p=0.000].Conclusion：1. The levels of adenosine in group PDR was higher than that in group control (P=0.000).The expression of CD73in group PDR was higher than that in group control (P=0.002). Theexpression of adenosine and CD73may be involved in the progress of revascularization withproliferative diabetic retinopathy.2. Experimentation on animals confirms that adenosine and CD73may be involved inthe pathogenesis of retinal revascularization."Adenosine homeostasis " may be the triggerfactors of the formation of RNV.3. In vivo,adenosine receptor antagonist inhibits retinal revascularization of OIR.Thatsuggests that adenosine receptors may become therapeutic targets for retinal neovasculardisease. Adenosine receptor antagonist inhibits the formation of new blood vessels in retinalof the OIR model, suggesting that adenosine receptors may become a new therapeutic targetsto retinal neovascular disease. |
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