Effects Of Polystyrene Microplastics And Cadmium On Oxidative Stress And Cell Apoptosis In Mice Liver

Microplastics（MPs）have been widely concerned as pollution substances in the 21st century.Due to their hard degradation,biodeposition and adsorbability,MPs have become potential threats to environmental organisms.MPs can adsorb heavy metals in aquatic and sedimentary environments,and these metals accumulate in microplastics.A variety of heavy metals have been found attached to Marine MPs.Cadmium（Cd）,as a common toxic heavy metal in the environment,is one of the heavy metals carried by MPs.It can cause a series of biological reactions such as oxidative stress,tissue edema,and organ damage,increasing the risk of exposure of environmental organisms to microplastics.At present,a large number of studies have shown that MPs alone or in combination with heavy metals has obvious stress effects on Marine organisms,but its effects on mammalian cells and tissues are still unclear and need further study.Therefore,in this paper,Polystyrene（PS）microplastics and cadmium chloride（Cd Cl₂）were used to simulate the joint effect of MPs and heavy metals,and mice were selected to establish poisoning models.To explore the effects of MPs,Cd and the combined effects of MPs and Cd on oxidative stress and cell apoptosis in mouse liver.180 male mice were divided into six groups（Control;Deionized water）,low dose polystyrene group（PS-L;PS-MPs:0.1 mg/kg/d）,medium dose polystyrene group（PS-M;PS-MPs:1 mg/kg/d）,high-dose polystyrene group（PS-H:PS-MPs:10 mg/kg/d）,cadmium group（Cd;Cd Cl₂:0.3 mg/kg/d）and cadmium+high-dose polystyrene group（Cd+PS-H;Cd Cl₂:0.3mg/kg/d;PS-MPs:10 mg/kg/d）were intragastric treated for 8 weeks.Blood was collected from eyeballs of mice,then neck was broken,and liver tissue was collected for follow-up detection.The activity of ALT and AST in serum was detected.The midline abdominal anatomy of mice was performed to observe the morphology of liver tissue and calculate the liver organ coefficient.The histopathological changes of liver were observed by staining.The levels of MDA,CAT,T-SOD and GSH in liver tissue were detected.The expression of Nrf2 protein in liver was detected by immunohistochemistry.The apoptosis of liver cells was detected by TUNEL staining.The m RNA expressions of Nrf2-Keap1 oxidative stress pathway,p62 and AMPK signaling pathway,Bcl-2/Bax apoptosis pathway mediated by mitochondria,p53 and MAPK signaling pathways in liver were detected by RT-qPCR.The expressions of Nrf2-Keap1oxidative stress pathway,p62,p53 and Bcl-2/Bax apoptosis pathway in liver were detected by Western Blot.The results showed that PS-MPs induced mice liver showed decreased gloss,pale color,increased liver cell swelling and inflammatory cell infiltration,and no significant changes in liver organ coefficient.Serum ALT and AST activities increased,liver tissue MDA levels increased,CAT,T-SOD and GSH levels decreased,and showed a dose correlation.The liver of mice induced by Cd alone showed rough mottling,pale liver edge color,liver congestion,cell swelling,large-scale particle degeneration,and significant reduction of liver organ coefficient.Serum ALT and AST activities increased,liver tissue MDA levels increased,CAT,T-SOD and GSH levels decreased.Compared with the effects of MPs and Cd alone,the liver color was darker and the hepatocyte granular degeneration was reduced.There were no significant differences in liver organ coefficient,serum ALT and AST activities,and liver tissue levels of MDA,CAT,T-SOD and GSH.ALT level increased and GSH level decreased significantly after MPs and Cd combined action compared with MPs alone action.Studies on Nr F2-related signaling pathways showed that MPs and Cd significantly decreased Nrf2 m RNA and protein levels in liver tissues,increased Keap1,HO-1,NQO1,GCLC and p62 m RNA and protein levels,and increased AMPK1,AMPK2 and AKTm m RNA levels.Compared with Cd,Nrf2m RNA level and HO-1 protein level increased,while GCLC,GCLM,NQO1 protein level and Keap1 m RNA level decreased.Compared with MPs,the levels of Keap1 m RNA,HO-1 m RNA and HO-1 protein were increased,while the levels of NQO1 m RNA and Nrf2 protein were decreased.The apoptosis level of hepatocytes detected by TUNEL showed that MPs and Cd both caused increased apoptosis of hepatocytes in liver tissue,and the protein and m RNA levels of Bax,Cyt-c,Caspase-3,p53 and p38,JNK and ERK were increased.The protein and m RNA levels of Bcl-2 and the ratio of Bcl-2/Bax protein expression were decreased.Under the combined action,the m RNA levels of Bax,Caspase-3 and p38 were decreased compared with Cd,the protein levels of Bcl-2 were decreased compared with MPs,and the ratio of Bcl-2/Bax was decreased.In conclusion,MPs and Cd independently cause pathological changes,oxidative stress and increased apoptosis levels in mouse liver through Nrf2-Keap1 pathway and Bcl-2/Bax-mediated mitochondrial apoptosis pathway.The combined effect of MPs with Cd mitigates the toxicity of Cd in Nrf2,Keap1,GCLC,GCLM,NQO1,Bax,Caspase-3,p38,and Bcl-2compared with the effect of Cd alone.The interaction between MPs and heavy metals does not increase the toxicity of heavy metals,so the mechanism of action remains to be further studied.