Protective Effect Of IL-33 On Ischemic Injury Of Lower Limbs

The lower limb ischemic disease belongs to a kind of peripheral vascular disease.It mainly refers to a series of diseases in which a series of clinical manifestations such as intermittent claudication,pain,ulcer,gangrene and so on,are caused by stenosis or occlusion of lower extremity arteries due to various reasons,resulting in decreased ischemic injury.In recent years,the incidence rate has gradually increased,and seriously affecting the quality of life of many patients,limb survival and even life-threatening.At present,the clinical treatment of lower limb ischemic diseases mainly includes traditional medicine treatment,such as anticoagulants and thrombolytics,surgical treatment,and interventional treatment.But the therapeutic effects of these treatmentsthe are not stable and most patients have cardiovascular and cerebrovascular diseases,so the treatment of lower limb ischemic diseases is still an important topic.Restoring the blood perfusion of the lower limbs of ischemic injury is the key to the treatment of ischemic diseases of the lower limbs.Biological therapy has gradually become a research hotspot.IL-33 belongs to the IL-1 family of cytokine members and mediates many immune-related diseases by binding to its specific receptor ST2.This study established a mouse model of lower limb ischemia,aiming to explore the protective effect and the regulatory mechanisms of IL-33 in ischemic injury of lower limbs,and provide new treatment methods and ideas for ischemic diseases of lower limbs.Objective:To study the protective effect of IL-33 on the lower limbs of ischemic injury in mice with lower limb ischemia,including IL-33 regulating angiogenesis and promoting the reperfusion of lower limb blood flow in ischemic injury.In vitro experiments were conducted to study the regulation of IL-33 on the biological functions of proliferation and migration of endothelial cells EAhy.926.Primary endothelial progenitor cells were isolated and cultured from mouse bone marrow to study the regulation of IL-33 on the biological functions such as proliferation and migration of bone marrow-derived primary endothelial progenitor cells.To explore the protective effect and regulation mechanism of IL-33 in ischemic injury of lower limbs.Method:The mouse model of lower limb ischemia was established and the mice were treated by intraperitoneal injection of IL-33.Laser Doppler scanning imaging was used to observe the blood perfusion of lower limbs in mice with ischemic injury at different time points.To determine the protective effect of exogenous IL-33 on ischemic injury of lower limbs.The effect of IL-33 on angiogenesis was detected by Matrigel embolization experiment and mouse aortic ring budding test.The ischemic injury of the lower limbs of the gastrocnemius muscle was exfoliated and the angiogenesis and expression of M2 macrophages were detected by immunofluorescence.The expression levels of cytokines VEGFA,VEGFR-2,TNF-α,Arg1,YM1,IL-4,IL-13,CXCL1,CXCL2 were detected by RT-PCR at the molecular level.The expression of M2 macrophages and endothelial progenitor cells in tissues was examined by flow cytometry.In vitro cell experiments,flow cytometry,cell scratch test and Transwell chamber assay were used to detect the regulation of IL-33 on the biological functions of proliferation and migration of endothelial cells EAhy.926 and its mechanism.Bone marrow-derived primary endothelial progenitor cells were cultured and the regulation of IL-33 on the biological function of endothelial progenitor cells and its mechanism were examined.Results:1.IL-33 improves blood flow reperfusion in ischemic injury tissueThe mouse model of lower limb ischemia was successfully established.And the mice were treated by intraperitoneal injection of IL-33.Laser Doppler scanner was used to detect the blood flow of lower limbs in mice with ischemic injury.The results showed that IL-33 can promote the recovery of blood flow to the lower extremity of ischemic injury and promote tissue repair.2.IL-33 promotes angiogenesis in ischemic tissuesImmunofluorescence assay showed that IL-33 has an effect of promoting angiogenesis.At the molecular level,RT-PCR results showed that IL-33 can up-regulate the expression of cytokines VEGFA,VEGFR-2 and TNF-α in ischemic injury tissues.Matrigel embolization experiments and mouse aortic ring budding experiments confirmed that IL-33 has an effect of promoting angiogenesis.At the same time,IL-33 regulates the polarization of macrophage to M2 type in ischemic injury,and up-regulates the expression of chemokines CXCL1,CXCL2 and VEGFA to promote angiogenesis.3.Effects of IL-33 on biological functions such as proliferation and migration of EAhy.926 cells in vitroIn vitro,cell experiments showed that IL-33 has the effect of promoting the proliferation of endothelial cells EAhy.926,and this effect is exerted by PI3K/AKT/STAT3 signaling.IL-33 has an effect of promoting the migration of EAhy.926 cells,and this effect is regulated by up-regulating the cell surface chemokine receptor CXCR2.4.Effect of IL-33 on the biological function of endothelial progenitor cellsIL-33 has the function of promoting the migration of endothelial progenitor cells to the ischemic injury tissues and promoting the proliferation of endothelial progenitor cells.IL-33 can up-regulate the expression of the surface chemokine receptor CXCR2 on endothelial progenitor cells.Conclusion:IL-33 has the effect of promoting neovascularization in ischemic injury tissues.And IL-33 can restore tissue blood flow by promoting blood flow reperfusion in ischemic injury tissue.IL-33 plays a protective role in ischemic injury to lower limbs by promoting ischemic injury tissue repair and promoting tissue function recovery.IL-33 can regulate angiogenesis by promoting the biological functions of endothelial cell such as proliferation and migration.At the same time,IL-33 has the effect of promoting the expression of endothelial progenitor cells in ischemic injury tissues.IL-33 can promote the proliferation of endothelial progenitor cells and IL-33 can promote the migration of endothelial progenitor cells to ischemic injury.Endothelial progenitor cells can be further differentiated into mature endothelial cells involved in angiogenesis.