Effects Of The Apoptosisis And Oxidative Damage Of Porcine Vascular Endothelial Cells By Fumonisin B₁

Fumonisin B₁（FB₁）,a water soluble secondary metabolites mainly produced by Fusarium moniliforme,is the main constituents of fuminosin and widely exists in food crops.It is well known that FB₁ exhibits hepatotoxicity and nephrotoxicity,and can cause Leukoencephalomalacia in horses and porcine pulmonary edema and oesophageal cancers in humans.In this study,we take porcine vascular endothelial cells（PIEC）as research object,aiming to study the toxicity on FB₁,including oxidative damage and apoptosis,and provide the basis for the study of animal toxicity of fumonisins.Objective:Taking PIEC cells as the research object,we treated the cells with different concentrations of FB₁ to discuss the toxicity of FB₁.Methods:PIEC in logarithmic growth phase were cultured in vitro and treated with different concentrations of FB₁（10,25,50μg/ml）in the experiment.MTT assay was used to determine the FB₁ test dose and duration of action.After cells were treated with FB₁ for 48 h,Inverted phase contrast microscope was used to observe apoptosis.NO contents were reduced using nitrate reductase method.MDA contents in the cells were tested by thiobarbituric acid method.To detect CAT activity,we used ammonium molybdate method.TNB and p-Nitro-Blue tetrazolium chloride（NBT）method were used to detect SOD and TrxR activities in cells.We also detect GSH contents and GSH-PX activity.Apoptosis was detected by Annexin-FITC/PI double staining method and ROS and mitochondrial membrane potential were tested with fluorescent probe DCFH-DA and JC-1 method.In addition,relative mRNA expression of Bax,Bcl-2,Caspase-3,Caspase-9 were detected by Q-PCR..Results:The optimal action time of FB₁ screened by cell proliferation assay was 48 h,and cell viability was 70%.High dose FB₁ inhibited cell proliferation（P<0.05）,increased the contents of NO and MDA（P<0.05）and decrease the activity of SOD（P<0.05）.Medium and high doses FB₁ significantly reduced the activity of CAT（P<0.05）.FB₁ significantly decreased activities of GSH,GSH-PX and TrxR（P<0.05）in dose-dependent relationship.The research revealed that FB₁ induced apoptosis through endogenous mitochondrial apoptotic pathway.FB₁significantly increased intracellular ROS levels（P<0.05）,activited apoptosis gene Bax and anti apoptosis gene Bcl-2,and decreased mitochondrial membrane potential（P<0.05）.And then the loss of mitochondrial membrane potential caused release of cells cytochrome C（P<0.05）,significantly activate the apoptosis promoter Caspase-9 and apoptotic Caspase-3,resulting in the collapse of cell.The results showed that the FB₁ in the high dose group could cause oxidative damage in PIEC cells and affect the normal apoptosis of cells.The effects of FB₁ in the low and medium dose groups on the oxidative damage and apoptosis of PIEC cells were not obvious.