Effect Of Human Fetal Dermal Mesenchymal Stem Cells On Burn-induced Microvascular Permeability

BackgroundSevere burn patients often experience tissue edema,shock and organ dysfunction,threatening the lives of patients.The pathological basis of increased microvascular permeability,microcirculatory disturbance,tissue edema,and hypovolemic shock after burns is structural damage and functional damage of the vascular endothelial barrier.Mesenchymal stem cells(MSCs)are a type of stem cells with strong proliferative capacity and multi-directional differentiation potential.They come from a wide range of sources,mainly in bone marrow,cord blood,subendothelium of umbilical vein,fat,peripheral blood and muscle.Because of its characteristics of self-renewal,self-reproduction,easy storage,multi-directional differentiation,and strong secretory functions,it has become a seed cell for the treatment of many clinical diseases.Among them,fetal dermal mesenchymal stem cells(FDMSCs)are pluripotent stem cells derived from fetal skin tissues of human fetuses which were accidental abortions without hereditary disease or drug therapy.FDMSCs have all the functional characteristics of MSCs.FDMSCs have excellent exosome secretion activity,cell proliferation,self-renewal ability,low immunogenicity,homing to damaged tissues,and promoting tissue regeneration.Nevertheless,the efficacy of them on burn-induced microvascular hyperpermeability remains poorly understood.ObjectiveTo observe effects of conditioned media of FDMSCs(F-CM)on burn-induced microvascular hyperpermeability,then to explore the feasibility of FDMSCs-related therapies in treating microcirculation disorders after burns.MethodsFDMSCs were extracted from the skin of fetal tissue,and F-CM were prepared using FDMSCs.Human microvascular endothelial cell line 1(HMEC-1)were obtained from the American Type Culture Collection(ATCC).In vivo experiments,the shaved skin was scalded in a circulating water bath(54 ℃)for 25 seconds,causing a deep Ⅱscald wounds in mice.And then 200 μL F-CM(0.5×CM,1×CM,2×CM)or SFM was injected into burn skin of back.After 24 hours,the effect of F-CM on vascular permeability in mice was tested by FITC-Dextran intravascular circulation method.In vitro experiments,HMEC-1 was treated with SFM,burn serum,and different concentrations of F-CM(0.5×CM,1×CM,2×CM),and the following indicators were detected:①The effect of F-CM on vascular permeability was tested in vivo using the Fluorescein isothiocyanate(FITC)-dextran internal circulation method to measure the permeability coefficients of HMEC-1 paracellular and transendothelial cells;②Effect of burn serum on endothelial cell via transwell inserts;③HMEC-1 proliferation by CCK-8;④The protein expression of VE-cadherin and Occludin in HMEC-1 by western blot;⑤The expression of F-actin and VE-cadherin was detected by immunofluorescence.Results①As the concentration of F-CM increases,it is more obvious to reduce vascular permeability in vivo;②With the higher the serum concentration of burns,the longer the action time,it is more obvious to increase endothelial cell vascular permeability;③F-CM could significantly improve HMEC-1 proliferation ability in a dose-dependent manner;④ With the increasing concentration of F-CM,the increasing protein expression of Occludin and VE-cadherin in HMEC-1 by western blot;⑤With the increasing concentration of F-CM,the increasing expression of F-actin and VE-cadherin was detected by immunofluorescence.ConclusionThrough a series of experiments,the results show that F-CM inhibited burn-induced microvascular hyperpermeability in vivo.In vitro,F-CM improved the stabilizing effect of HMEC-1 by increasing protein expression of occludin and VE-cadherin,restoring the remodeling of endothelial F-actin.It can reduce the exudation of intravascular fluid caused by burns,tissue edema,and even prevents the occurrence of hypovolemic shock.