Effect Of Phenethyl Isothiocyanate On Acute Toxicity Of Acrylonitrile In Diabetes And Its Mechanism

Objective:Diabetes mellitus is a chronic disease with endocrine dysfunction,and it can cause multiple complications.Diabetes is one of the sensitive population that is endangered by environmental toxicants.It has been proved that diabetes enhanced the health risk caused by environmental hazards,physical hazards,chemical drugs and microbial infection.Therefore,protecting the health of diabetic population and making it less harmful to environmental toxicants is the hot topic of current research.Acrylonitrile(acrylonitrile,AN)is an organic synthetic industrial material with high volatility and high toxicity.Cytochrome P4502E1(CYP2E1)is a key enzyme of AN.Diabetes can increase the activity of CYP2E1.Up-regulation of CYP2E1 can enhance the toxicity of most environmental toxicants.Previous studies have found that the up-regulation of CYP2E1 could accelerate the speed of AN metabolism and increase the CN~-content,resulting in stronger acute toxicity of AN.Phenethyl isothiocyanate(PEITC)is an enzymatic hydrolysate of glucosinolates from cruciferous plants.It has many pharmacological effects,such as inhibition of CYP2E1 enzymes,anti-oxidation,anti-cancer and anti-inflammatory.On the basis of of the establishment of STZ-inuduced diabetic rats and the purchase of diabetic db/db mice,naturing products PEITC was pretreated to diabetic animals which were treated with AN to investigate the susceptibility of diabetes mellitus to the acute toxicity of industrial toxicants,which is represented by AN,and the mechanism,and the protective effects of PEITC,so as to provide guidelines for the protection of susceptible people such as diabetes.Methods:The diabetic rat model was established by one-off intraperitoneal injection of65mg/kg STZ.To evaluate the susceptibility of streptozotocin(STZ)-induced diabetic rats to the acute toxicity of AN,and biochemical mechanism,as well as the antagonistic effect of natural product PEITC on the acue toxicity of AN in STZ-induced diabetic rats.The specific methods are as follows:diabetic rat model was induced by STZ 14 d after the successful establishment of the model,the rats were randomly divided into normal blank control group,diabetic control group,AN group,20,40,80 mg/kg PEITC+AN groups,diabetes+AN group,diabetes+20,40,80 mg/kg PEITC+AN groups.PEITCwere orally given to rats for 3 days,1 h after the last gavage of the PEITC,rats were intraperitoneally injected with 90 mg/kg AN.The changes of behavior and time to death in 24 h in rats were observed.Liver cytochrome P450 2E1(CYP2E1)activity and brain and liver cytochrome c oxidase(CcOx)activity were detectd to evaluate the role of toxicant metabolizing enzyme CYP2E1 in the susceptibility of STZ-induced diabetic rats and the protective mechanism of PEITC.Glutathione(GSH)content,glutathione-S-transferase(GST)activity and reactive oxygen species(ROS)levels in brain tissue were detected to investigate the effect of PEITC on the oxidative stress in STZ-induced diabetic rats treated with AN.The susceptibility of db/db mice to the acute toxic of AN and biochemical mechanism,and the antagonistic effect of natural product PEITC on the acue toxicity of AN in db/db mice were also studied.The experimental grouping and treatment methods and the corresponding detection index were the same as the experimental methods of STZ induced diabetic rats.Results:1.STZ-induced diabetic rats:behavioral observation showed that compared with normal rats injected with AN,the time to convulsion,time to loss of righting reflex,and time to death in STZ-induced diabetic rats treated with AN were shorter and the mortality rate was significantly higher.,indicating that STZ-induced diabetic rats are more sensitive to acute toxicity of AN.PEITC pretreatment could signifiacantly decrease the time to convulsion,loss of righting reflex and death and reduce the rate of convulsion,loss of righting reflex and mortality.The acute toxicity of STZ-induced diabetic rats which were ppretreated with PEITC,followed by injection of AN was significantly relieved.Compared with the blank control group,the activity of CYP2E1 was significantly increased in diabetic control group.Exposure to AN for 1h had no significant effect on CYP2E1 protein expression.After pretreated with PEITC,followed by injection of AN,the activity of CYP2E1 in liver and the levels of ROS in brain were significantly decreased.The activity of CcOx in liver and brain,the content of GSH,and the activity of GST in brain were significantly increased in diabetic rats.The results further verified the resuts of the animal behavior observation experiment,indicating that the enhanced acute toxicity of AN in STZ-induced diabetic rats is due to its upregulation of CYP2E1 activity.PEITC can effectively antagonize the acute toxicity of AN in STZ induced diabetic rats through inhibition of CYP2E1 enzyme.2.Diabetic db/db mice model:The results of behavior observation showed that compared to wild type mice of the same nest treated with AN,the time to death was significantly decreased and the mortality rate was significantly increased in diabetic db/db mice treated with AN.After PEITC pretreatment,the time to death of diabetic db/db mice treated with AN was obviously prolonged and the mortality rate was significantly decreased.There was no significant difference in protein expression of CYP2E1 and activity of CYP2E1 between db/db mice and wild type mice of the same nest.Acute exposure to AN did not significantly affect the expression and activity of CYP2E1 protein in db/db mice and wild type mice of the same nest(Fig 3.2 and Fig3.3),indicating that the enhanced acute toxicity of AN in db/db mice may not be mediated by CYP2E1 activity.Compared with the db+AN group,PEITC(40 mg/kg)pretreatment significantly decreased liver CYP2E1 activity,increased brain and liver CcOx activity and decreased brain ROS levels,indicating that PEITC can effectively antagonize the acute toxicity of AN in diabetic db/db mice.Conclusion:After acute exposure to AN,compared with the normal rats or mice treated in the same way,the survival time of STZ-induced diabetic rats and diabetic db/db mice modelwas significantly shortened and the mortality rate was significantly increased,indicating that all the two types of diabetes increased the acute toxicity of AN.STZ-induced diabetic rats accelerate the production of CN~-by up-regulation of CYP2E1 activity,thus inhibiting the activity of CcOx in the tissue,resulting in the increase of the mortality rate and enhanced toxicity of diabetic rats after acute AN exposure.Compared with wild type mice of the same nest,the CYP2E1 activity was not altered in diabetic db/db mice,indicating that the enhanced acute toxicity of AN in db/db mice may not be mediated by CYP2E1 activity.PEITC mainly alleviated the acute toxicity of AN by inhibiting the activity of CYP2E1enzyme and relieving oxidative stress.