Optical Clearing Imaging Assisted Evaluation Of Urokinase Thrombolytic Therapy On Cerebral Vessels With Different Sizes

Ischemic stroke is a central nervous system disease caused by clogging of cerebrovascular vessels,which has high disability and fatalities.At present,the main method for the treatment of ischemic stroke is intravenous thrombolysis.Urokinase,as a common clinical thrombolytic drug in china,has the advantages of good thrombolytic effect,long time window,and low price,but there are frequent cases in which treatment does not work.In recent years,a large number of studies have explored the factors that affect the effect of urokinase.Among the factors that affect thrombolytic therapy,the size of stroke blood vessels is the focus of researchers.But no research has systematically evaluated the correlation between vessel size and urokinase thrombolytic effect.To carry out this study,the first thing is to bulid stroke models which are close to the clinic and can cause targeted damages of blood vessels with different sizes;Second,the thrombolytic effects of urokinase on vessels with different sizes can be dynamically monitored.The work of this article is focused on the assessment of the response of blood vessels with different sizes to urokinase thrombolytic therapy,mainly including the following two aspects:(1)Establishing a targeted ischemic stroke mice model with embolus components close to clinical: Firstly,stroke models with embolus composition close to clinical conditions were established respectively on the middle cerebral artery,secondary branches of middle cerebral artery and terminal vessels of middle cerebral artery,by combining optical clearing skull window and photothrombosis with the introduction of thrombin.Secondly,combined with the laser speckle imaging technology,the minimum doses of embolization were selected for vessels with different sizes.The results showed that the doses of embolization were 5 m W×2 min for the terminal vessel of the middle cerebral artery,5 m W×5 min for the secondary branch of the middle cerebral artery and 20 m W×5 min for the middle cerebral artery when rose bengal and thrombin were injected at 25 mg/kg and 80 U/kg,respectively.Next,immunofluorescence staining and TTC staining were used to analyze the emboli components,and observe the locations of the infarct areas 24 hours later.The results showed that the embolus contained both platelet and fibrin,which was closer to the clinical situation,and the ischemic tissues were in good agreement with the areas where the target blood vessels were located.(2)Evaluation of the urokinase thrombolytic effects on vessels with different sizes:After thrombin combined photothrombosis stroke models were established,Urokinase was used for non-intervention/intervention therapy to treat ischemic stroke models with vessel with different sizes.Monitoring the changes in cerebral blood flow/oxygen saturation of stroke vessels with optical clearing skull window and laser speckle/hyperspectral dual-mode imaging system could evaluate the thrombolytic effects of urokinase on vessels with different sizes.The results showed that: for small-sized blood vessel(such as the terminal vessels of the middle cerebral artery),urokinase dissolved thrombosis quickly,but it was prone to secondary blockage.For larger blood vessel(such as the secondary branch of the middle cerebral artery),urokinase could quickly dissolve thrombosis,and the blood vessels could always be recanalized.For blood vessel that was too large(such as the middle cerebral artery),thrombolysis of urokinase was slow and incomplete.In the non-intervention condition,none of the three vessel sizes recovered after laser occlusion,and the infarct areas were further enlarged.In summary,this study combined the optical clearing skull window and thrombin combined photothrombosis stroke model to build a targeted ischemic stroke model with embolic components consistent with clinical thrombus,while preserving the intact skull.And we initially explored the effect of vessel size on urokinase thrombolytic therapy.It provides strong support for the basic research of thrombolytic drugs,and is expected to provide a theoretical basis for the choice of clinical use of urokinase.