Effects And Mechanism Of Ionic Liquid [Bmim]Cl On Energy Metabolism Of Human Hepatocellular Cells HepG2 And Zebrafish(Danio Rerio)

Ionic liquids are a kind of organic solvents widely used in production and research,which makes their ecological safety and toxicological effects on organisms gradually become a research hotspot.In this paper,human hepatocellular cells（HepG2）and Danio rerio were used as experimental subjects to systematically study the effects of 1-butyl-3-methylimidazolium chloride（[Bmim]Cl,a commonly used imidazole ionic liquid）on glucose and lipid metabolism.The main results are as follows:1.In order to verify the effect of[Bmim]Cl on glucose metabolism of HepG2cells,HepG2 cells were exposed to[Bmim]Cl at the concentration of 1,2,4 or 8mmol·L^-1（mM）for 24 h.The results showed that the glucose consumption in the culture medium and the intracellular glycogen content in HepG2 cells decreased significantly after exposure to[Bmim]Cl,and the glucose consumption in the culture medium of 2 mM treatment group was only 19%of that in control group.Through real-time quantitative PCR（RT-qPCR）,it was found that the expression of glycogen phosphorylase（pygl）and pyruvate kinase M（pkm）increased in all treatment groups,and the expression of glucose-6-phosphatase（g6pc）,the key gene of gluconeogenesis,was significantly down-regulated in 2,4 and 8 mM treatment groups.The expression of rate-limiting genes in tricarboxylic acid cycle（TCA cycle）was also affected.The expression of isocitrate dehydrogenase（idh2）was significantly up-regulated in 8 mM treatment group,which was 5.3 times higher than that of control group.In brief,our study found that[Bmim]Cl exposure can affect the extracellular glucose absorption of HepG2 cells,affect the synthesis and decomposition of intracellular glycogen,promote glycolysis,inhibit gluconeogenesis.High concentration of[Bmim]Cl exposure can cause glucose metabolism disorder in HepG2 cells.2.In order to further verify the glucose metabolic toxicity of[Bmim]Cl,adult Danio rerio was used as an experimental model,which was exposed to[Bmim]Cl at the concentration of 50,100,200 or 400 mg·L^-1,and the related indexes were determined 15 d and 30 d later.It was found that the experimental results of 15 d and30 d were consistent.The liver of Danio rerio was damaged to varying degrees by exposure to[Bmim]Cl.Histological lesions such as lymphocyte infiltration,cell vacuolation,loose and light staining of hepatocytes and so on appeared in the liver of Danio rerio in 400mg·L^-11 treatment group.Further studies found that glucagon（an important hormone regulating blood glucose homeostasis）significantly increased after exposure to[Bmim]Cl,thus inhibiting glycolysis,promoting gluconeogenesis and promoting glycogen decomposition in Danio rerio.For detail,the contents of glycogen and pyruvate in liver decreased,pklr（the key gene of glycolysis）decreased,while g6pca（the key gene of gluconeogenesis）and pygl（the key gene of glycogen decomposition）increased significantly.In summary,our study found that exposure to ionic liquid[Bmim]Cl can damage the liver of Danio rerio and affect the liver glucose metabolism of Danio rerio.3.Similar to sugar metabolism,lipid metabolism is also an important part of energy metabolism.In order to further the effect of[Bmim]Cl on energy metabolism of Danio rerio,on the basis of the above experiments,the changes of related indexes of lipid metabolism were detected to explore the effect and mechanism of[Bmim]Cl on liver lipid metabolism of Danio rerio.The results showed that after exposure to ionic liquid[Bmim]Cl for 30 d,the content of triglycerides in Danio rerio（including liver）decreased significantly,indicating that[Bmim]Cl may affect the lipid metabolism of Danio rerio.Further studies showed that lipca（the key gene that catalyzes the hydrolysis of triglycerides）in liver of Danio rerio was significantly up-regulated in several treatment groups,while gpat3（the important gene involved in triglyceride synthesis）was down-regulated,and mcat and loc（two related genes which inhibit fatty acid decomposition and promote triglyceride synthesis）were down-regulated in varying degrees.In summary,our study found that exposure to[Bmim]Cl can accelerate lipid decomposition and inhibit the synthesis of fatty acids and triglycerides in Danio rerio,resulting in lipid metabolism disorder.To sum up,it was found that exposure to ionic liquid[Bmim]Cl can induce glucose metabolism disorder in HepG2 cells;cause liver damage of Danio rerio,and lead to disorder of glucose metabolism and lipid metabolism of Danio rerio.The study of this paper defines the metabolic toxicity of[Bmim]Cl,enriches the toxicity data of ionic liquids,and provides a basis for the establishment of eco-environmental pollutant standards.