| Tris-(2,3-dibromopropyl)isocyanurate(TBC)is a bromine compound containing a hexabromo heterocyclic structure.Its chemical structure is relatively stable and volatile low flame retardant,widely used in China for the synthesis of chemical products such as flame retardants,plastics,textiles and rubber.Since the end of the 20 th century,China's TBC production has been no less than500 tons per year,and it has been increasing year by year.At the same time,the use of various countries in the world is also increasing.People are unavoidably exposed directly or indirectly,so it is necessary to study the toxicity of TBC.According to literature reports,TBC is bioaccumulative and can persist in the environment for a long time,causing unavoidable and persistent pollution to the environment.TBC inhibits algae growth and is hepatotoxic to zebrafish and rodents.In this study,after studying the subchronic toxicity of TBC to Wistar rats,the target organs were identified,and its hepatotoxicity to Wistar rats and its mitochondrial-induced apoptosis mechanism were mainly explained.The acute toxicity classification of TBC was determined to be non-toxic through an acute toxicity experiment.Eighty Wistar rats were fed on 28 days,and the females were halved.They were randomly divided into four groups,the control group and the three poisoned groups,the doses were 1.250 g /(kg · bw),0.625 g /(kg · bw),0.313 g /(kg · bw).The growth situation of Wistar rats were recorded everyday.Animals were fasted 12 hours before dissected.Weighed and allocated anesthesia early the next morning,took blood after anesthesia,and weighed heart,liver,spleen,lung,kidney,ovary and testicular organs.After weighing,3 to 4 samples were taken from the male and female control group,low-dose group,medium-dose group and high-dose group,and each organ was fixed with formaldehyde,paraffin sections were HE stained,and histopathological examination was performed to determine the target of TBC toxicity.Other organs were frozen at-80 ? after being fixed with liquid nitrogen.After centrifugation of blood,serum was taken for biochemical index detection and blood analysis.ANOVA was used to compare the mean difference between each exposure group and the control group.Real-time quantitative PCR was used to detect genes that induce apoptosis in rat hepatocytes.At the same time,Western blot was used to detect protein expression of relative proteins with altered gene expression.The correlation between genes and proteins was analyzed,and TBC was explored from the mitochondrial pathway.Mechanism of inducing apoptosis of hepatocytes in Wistar rats via mitochondria pathway.During the 28-day subchronic toxicity process,Wistar rats showed signs of mental decline,loss of appetite,growth retardation,weight loss,and dull hair as TBC exposure increased.Pathological examinations of the heart,liver,spleen,lungs and kidneys of rats were performed.It was found that myocardial hypertrophy,abnormal intercellular spaces,and inflammation and infiltration were appeared.Air bubbles appeared in the lungs and expanded unevenly,and cell swelling and bleeding were seen;kidneys occurred Edema,part of the interstitial structure of the cell is fuzzy and scattered.Liver damage was the most serious,and the liver was inflamed,swollen,and degenerate to different degrees.The tissue structure of the exposed group had obvious lesions compared with the control group.It was determined that it had different degrees of toxicity to rat organs.Among them,liver damage was the most serious.The results of pathological examination were consistent with those of blood.In the detection of genes that induce apoptosis of liver cells,the expression of Bcl-2 gene decreased with increasing dose,and the expression levels of Bax,Cytochrome C,C3,and C9 increased with increasing dose.The expression levels of Bax,Bcl-2,Cytochrome C,C3,and C9 genes were dependent on the dose of TBC to Wistar rats.Protein expression showed that the expression ratio of Bcl-2 / Bax protein in the exposed group(bcl-xl)decreased,and the expression of Cytochrome C and Caspase-9 protein in the cells increased.Caspase-3 in the exposed group gradually increased with the increase of TBC concentration.Endoplasmic reticulum stress response,the expression of Caspase-12 protein gradually decreased with increasing TBC concentration,and the expression of protein and TBC concentration were dose-dependent.The female mice BCL2L1 gene and Bax protein were statistically significant,P <0.05.The difference between BCL2L1 gene and Caspase3 protein was very significant,P <0.01.BCL2L11 gene and Bax protein were statistically significant,P <0.05.The difference between BCL2L11 gene and Caspase3 protein was very significant,P <0.001 BCL2L2 gene and Bcl-xl protein were statistically significant,P <0.05.CASP9 gene and Bax protein were statistically significant,P <0.05.CASP9 gene and Caspase9 protein were statistically significant,P <0.05.The BAX,BCL2,BCL2A1,and CASP3 genes were not significantly related to Bax,Bcl-2,Bcl-xl,Cytochrome C,Caspase-9,and Caspase-3 proteins,P> 0.05.The male mouse BAX gene and Bcl-2 protein were statistically significant,P <0.05.The BCL2 gene and Cytochrome C protein were statistically significant,P <0.05.BCL2L1 gene and Bcl-2 protein were statistically significant,P <0.05.BCL2L2 and Bcl-2 proteins were statistically significant,P <0.05.CASP3 gene and Bcl-2 protein were statistically significant,P <0.05.The CASP3 gene and Cytochrome C protein were statistically significant,P <0.05.CASP9 gene and Bcl-2 protein were statistically significant,P <0.05.There was no correlation between BCL2A1,BCL2L11,and CYCS genes and Bax,Bcl-2,bcl-xl,Cytochrome C,Caspase3,and Caspase9 proteins,P> 0.05.The target organ was mainly the liver.In the mitochondrial signaling pathway that induces hepatocyte apoptosis,TBC reduces the(bcl-xl)Bcl-2 / Bax protein expression ratio and opens the MPTP hole.Membrane potential decreases,Cytochrome C is released outside the mitochondria,activates members of the Caspases family,Caspase-9 forms apoptotic bodies,and awakens Caspase-3 to undergo proteolysis to induce apoptosis;we try to explore,With the increase of TBC concentration,the expression of Caspase-12 increased,and proteolysis occurred,resulting in apoptosis.Analysis of gene-protein binding and similar trends.TBC can induce Wistar rat liver cell apoptosis through the mitochondrial pathway,and may also induce Wistar rat liver cell apoptosis in the endoplasmic reticulum stress response. |
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