Toxicity Of Trimethyltin Chloride On Intestinal Porcine Epithelial Cells

Organotin compounds are commonly used organometallic compounds in people’s lives and agricultural production.At the same time,there is a certain amount of toxicity,it is easy to remain in the environment.In particular,the extensive use of agricultural production has led to the accumulation of organotin in the environment to varying degrees.And through the food chain to bring different degrees of harm to animal production,caused widespread concern.In this study,intestinal porcine epithelial cells(IPEC-J2)were exposed to different concentrations of Trimethyltin Chloride(TMT).To study the effects of Trimethyltin Chloride on oxidative damage,cell cycle and apoptosis of cells,to investigate the toxicity of Trimethyltin Chloride on IPEC-J2 cells,and to provide the basis for the study of organotin toxicity to animals.IPEC-J2 cells were cultured in vitro with 0.5 μg/m L,1 μg/m L,5 μg/m L and 10μg/m L TMT.Cell viability was measured by MTT assay and morphological changes were observed by inverted phase contrast microscopy.LDH activity in cell supernatant was detected by 2,4-dinitrophenylhydrazine assay;SOD activity was detected by colorimetry;apoptotic rate and cell cycle were detected by flow cytometry;Cyclin B1,Cyclin D1 and P21,apoptotic genes Bax,Bcl-2 and Caspase-3 were detected by Realtime PCR.The results showed that the cell viability decreased with the increase of TMT concentration.After 24 h of TMT,the survival rate of cells decreased from 100% to85.83% and 73.61%,and the difference was significant.Under the microscope,the cells began to shrink and round,and a large number of floating dead cells appeared.LDH in supernatant of cells treated with 1 μg/m L,5 μg/m L and 10 μg/m L TMT for 48 hours were 10.48,11.83 and 14.41U/m L,respectively.Which significantly increased LDH release and enhanced LDH enzyme activity.SOD activity of 1 μg/m L,5 μg/m L,10μg/m L TMT was 11.11,7.18 and 10.46U/mg prot 24 hours after treatment,which significantly decreased SOD activity compared with the control group.Flow cytometry showed that cell proliferation was blocked in G1 phase after TMT treatment,and the expression of Cyclin B1 and Cyclin D1 genes was significantly decreased in TMT treatment groups of 1 μg/m L,5 μg/m L and 10 μg/m L.The expression of P21 was significantly increased,especially in the presence of 5 μg/m L and 10 μg/m L TMT.The expression of P21 increased from 1.03 to 3.15 and 2.52,respectively,in the control group.24 hours after treatment with 1 μg/m L,5 μg/m L and 10 μg/m L TMT,the apoptotic rate increased,the ratio of Bax/Bcl-2 increased,and the expression of Caspase-3 also increased significantly.This indicates that TMT inhibits proliferation of intestinal porcine epithelial cells in vitro and induces oxidative damage and apoptosis.