| Hexavalent chromium(Cr(Ⅵ))is a heavy metal pollutant.Its accumulation and distribution in animal organs are different.Some studies have shown that the organs with high to low accumulation of hexavalent chromium in poultry are: testis,kidney,liver,heart and brain.At present,the research mainly focuses on the toxicity of Cr(Ⅵ)to broiler liver and kidney,but the toxicity of Cr(Ⅵ)to broiler myocardium is not clear.The purpose of this study was to explore the toxic effect and mechanism of Cr(Ⅵ)on broiler myocardium,and to provide experimental basis and reference for the study of Cr(Ⅵ)poisoning mechanism and the preparation of subsequent antidotes.96 1-day-old AA white-feathered broilers were randomly divided into 4 groups after a 7-day adaptation period: Control group,Low-dose group,Medium-dose group and High-dose group,the intragastric concentrations of potassium dichromate were 0,18.56,37.12 and 74.24 mg/kg/d,respectively.Then,six broilers in each group were selected at14,21,28 and 35 days respectively.The heart was collected after anesthesia,the heart organ index was detected,the heart tissue was stained with HE,and the contents of SOD and MDA in myocardial tissue were detected.In the in vitro experiment,11 d broiler embryo cardiomyocytes were isolated by enzyme digestion,and the optimal experimental concentration of Cr(Ⅵ)was determined by CCK-8.Cardiomyocytes were divided into 3groups: Control group,Low-dose group,High-dose group,the concentrations of potassium dichromate in the cell culture medium were 0,16 and 32 μmol/L respectively.After 24 hours of exposure,the cells were subjected to morphological observation,flow cytometric apoptosis detection,mitochondrial membrane potential detection,ROS detection,RNA-seq,RT-qPCR and Western blot tests.In the in vivo experiment,the body weight of the Cr(Ⅵ)exposed group was lower than that of the Control group,and the heart index was higher than that of the Control group,but there was no significant difference.The pathological observation of myocardium showed that the myocardial structure of the Control group was normal without obvious lesions,and the Cr(Ⅵ)exposed group showed fibrous connective tissue hyperplasia and mild inflammatory cell infiltration.Compared with the Control group,the content of SOD in myocardial tissue of the exposed group decreased significantly and the content of MDA increased significantly.In the in vitro experiment,the results of CCK-8showed that the half inhibitory concentration of Cr(Ⅵ)was 64 μmol/L.Compared with the Control group,the Cr(Ⅵ)exposed group had a large number of cell death and fewer adherent cells.The results of flow cytometry showed that the apoptosis rate of cardiomyocytes in Cr(Ⅵ)exposure group was significantly higher than that in Control group;RNA-seq results showed that there were 6246 differential genes between the Control group and the Cr(Ⅵ)exposure group,including 2871 up-regulated genes and3375 down-regulated genes.Pathway enrichment analysis showed that the differential genes were mainly concentrated in p53 signaling pathway,apoptosis pathway and ferroptosis pathway.The m RNA expression levels of apoptosis related genes Bax and p53 in the Cr(Ⅵ)exposure group were significantly higher than the Control group,and the m RNA expression level of Bcl-2 was significantly lower than the Control group.Compared with the Control group,the m RNA and protein levels of autophagy related genes LC3-Ⅰ and LC3-Ⅱ increased significantly,the protein level of Beclin1 increased significantly,and the protein level of p62/SQSTM1 decreased significantly in the Cr(Ⅵ)exposure group.The content of ROS in Cr(Ⅵ)exposure group was higher than Control group.The Cr(Ⅵ)exposed group showed lower mitochondrial membrane potential than the Control group.In addition,compared with the Control group,the m RNA levels of mitochondrial function related genes SIRT1 and Mfn2 in the High-dose group decreased significantly,and the m RNA level of SIRT3 in the Low-dose group and High-dose group decreased significantly.The results showed that Cr(Ⅵ)could cause toxicity and damage to broiler cardiomyocytes,enhance the oxidation level of broiler cardiomyocytes and induce oxidative stress;Cr(Ⅵ)can also induce the decrease of mitochondrial membrane potential,apoptosis and autophagy in broiler cardiomyocytes. |
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