Study On The Treatment And Related Mechanism Of Fundus Neovascularization Disease

Ocular fundus neovascularization is one of the important causes of blinding eye disease.At present,the clinical treatment for ocular fundus neovascularization disease mainly include two types:surgery(such as laser photocoagulation)and drug interventions(such as anti-VEGF treatment).These treatments can reduce the occurrence and development of the ocular neovascularization to a certain extent,but there are still a large number of patients with varying degrees of adverse reactions or ineffective treatments.Therefore,it is important to explore the new pathogenesis and treatment of fundus neovascularization.This study focused on the optimization of fundus neovascularization disease mouse model and the exploration of the new pathogenesis,and thus laied a foundation for the exploration of new treatment for ocular fundus neovascularization.The main contents of this dissertation include three parts:1.A hand-held houlder for mouse was designed to optimize the parameters of laser-induced CNV model.Laser-induced mouse CNV model as one of the most widely recognized mouse models of wet AMD mimics the common features of choroidal neovascularization.However,the complicated operation of modeling process requires long time training to ensure the success rate of induction and the uniformity of neovascular lesions,which limits the subsequent evaluation of new drug.We pioneering designed a handheld holder for mice,and invited six ophthalmology residents to study the laser induced CNV mice modeling.By evaluated the learning time of mastering the modeling method,we found that utilization the holder can significantly shorten the average learning time to grasp the method of modeling and can effectively relieve physical and mental fatigue in the process of operation.On the other hand,we invited an operator proficient in laser-induced CNV mouse modeling,comparing by hand,modeling by holder can significantly shorten the time to complete one mouse modeling and improve the success rate of CNV and the uniformity of CNV lesions.These results indicate that handheld holder of mouse can optimize the parameters of laser-induced CNV mouse model,which could provide theoretical and experimental basis for further study of the treatment of ocular fundus neovascularization.2.CD146 as a new therapeutic target for fundus neovascularization was explored.Based on the above experimental,we investigated the role of adhesion molecule CD146in hypoxia-induced fundus neovascularization.In physiological and pathological angiogenesis of tumor,CD146 plays a decisive role.Our results suggested that CD146protein expression was elevated in retina of laser-induced CNV mouse model and OIR mouse model,which mainly concentrated in abnormal neovascularization,indicated that CD146 was abnormally activated in fundus neovascular area.Further studies showed overexpression of CD146 of HRECs increased hypoxia-related tube formation and migration,while knockdown of CD146 significantly weaked these abilities,which suggested that CD146 played a key role in hypoxia-induced fundus angiogenesis.By established an OIR model using endothelial-specific Cd146 conditional knockout mice(Cd146^EC-KO),we further confirmed these results that CD146 deficiency inhibited ocular angiogenesis.Therefore,we regarded CD146 as a therapeutic target of fundus neovascularization.In vitro,by using anti-CD146 m Ab AA98 inhibiting the activity of CD146,the capacity of hypoxia-induced tube formation and cell migration was significantly reduced.In vivo,preventive or therapeutic targeted CD146 by vitreous injecting with AA98 could effectively reduce the occurrence of fundus neovascularization and vascular leakage in CNV and OIR mouse model.In addition,we found that the inhibitory effects of CD146 specific antibody AA98 and VEGF specific antibody Bev on downstream signaling pathways were both overlapping and independent,and the combined use of the two antibodies significantly enhanced the inhibitory effect on neovascularization,which suggested that combined treatment of anti-CD146 and anti-VEGF has a synergistic antiangiogenic effect.These results suggest a new therapeutic target of CD146 for fundus neovascularization,and a new strategy for anti-CD146combined with anti-VEGF therapy.3.A new strategy for fundus neovascularization therapy using redox regulated nanozyme was proposed.Aiming at the oxidative stress caused by hypoxia in retinal neovascularization,we designed a nanozyme with redox regulation ability,called Pt@Mito Lipo nanozyme,and tested its therapeutic value in fundus neovascularization.This nanozyme is mainly composed of liposome shell,Pt nanozyme content and TPP group with mitochondrial targeting function,which has excellent SOD and CAT enzyme activities.In vitro,Pt@Mito Lipo nanozyme eliminated intracellular ROS accumulation,alleviated mitochondrial damage,and rescued cell hypoxia degree by generating O₂,which indicated a potential ability to inhibit neovascularization.And in vivo,we further found that nanozyme could effectively reduce the occourence of retinal neovascularization and accelerate physiological vascular repair,which is achieved by correcting the pathological hypoxic retinal vascular environment.The improvement of hypoxia environment corrected the abnormal activation of VEGF signaling pathway,and rescued the loss of the tip cell numbers of retinal vascular in mice and reduced number of filopodia of tip cell,thus preventing the growth of pathological angiogenesis and making them repaired in normal form.In addition,Pt@Mito Lipo nanozyme had no damage to vascular and photoreceptor of both immture and adult mice,which indicated that the biosafety of Pt@Mito Lipo nanozyme is reliable.These results suggest a nanozyme therapy for fundus neovascularization,and provide a new redox regulation strategy for hypoxia-induced fundus neovascularization.