Effects Of Ag Nanoparticles On The Growth Of Spodoptera Frugiperda Sf9 Cells And Silkworm And Omics Analysis

Ag nanoparticles(AgNPs)are one of the nanomaterials.It has been widely used in recent years,including biomedicine,food packaging.They are of great benefit to humans and also have the potential harm to human health and the environment.AgNPs have already shown toxic and other potentially harmful effects in mammals.Although studies have evaluated the toxic effects of AgNPs,studies on hazards to insects and environmental risks are still limited.In this study,we used Spodoptera frugiperda Sf9 cells and silkworms to study different concentrations of AgNPs,assessing their toxic effects.The effects of AgNPs on the growth status of Sf9 and the body weight,cocoon shells,and moth rate of silkworm were observed.At the level of transcription,the effect of AgNPs on the transcription of midgut was analyzed by whole transcriptome sequencing.At the level of protein,the proteomic differences of midgut and fat body of silkworm were analyzed by two-dimensional electrophoresis(2-DE).The differentially expressed genes or proteins were verified by realtime fluorescence quantitative PCR(qPCR).The main results of this paper are as follows:(1)Different concentrations of AgNPs were added to Sf9 cells.The results showed that the morphology of Sf9 cells wes changed from round to shuttle and other irregularities after adding 10 mg L-1 AgNPs.At the concentration of 50,100 mg L-1,the cells cells outflow efflux seriously and behave cell death.With the increase of AgNPs,the cell death was further aggravated.(2)The effects of different concentrations of AgNPs on the growth of silkworm were studied.The results showed that low concentrations of AgNPs(<400 mg L-1)promoted the growth and baby weights of silkworm.Although the concentration of 800 mg L-1 also improved B.mori growth,they resulted in silkworm death.With the increase of the concentration of AgNPs,the survival rate and moth rate were significantly reduced.(3)Effects of AgNPs on the midgut transcriptome of silkworm: After adding AgNPs,sequencing analysis showed that 252 genes were differentially expressed: 173 upregulated and 79 downregulated,with 43 genes significantly differential expressed:39 upregulated and 4 downregulated.GO analysis of the differentially expressed genes found that AgNPs were able to induce significant differences in the genes involved in transmembrane transport and carbohydrate metabolic process.At the same time,KEGG analysis showed that the genes involved in galactose metabolism,nitrogen metabolism,amino sugar and nucleotide sugar metabolism showed significant difference expression.(4)Effects of AgNPs on proteome of silkworm: The expression of proteins in silkworm midguts was analyzed by 2-DE.The results found 14 significant differentially expressed protein spots,11 downregulated and 3 upregulated.qPCR results showed that at an AgNPs concentration of 400 mg L–1,the expression levels of nine proteins were similar to the transcription levels of their corresponding genes.An analysis of fat body proteomics differences revealed 13 significant differences in fat body protein spots,and nine of which exhibited significantly downregulated expression,while four showed significantly upregulated expression.qPCR results showed that 7 protein spots were validated.Analysis showed that differentially expressed proteins play an important role in antioxidant stress,carbon metabolism and amino acid metabolism.Studies show that AgNPs can induce Sf9 cells morphological changes and cell death;AgNPs induced oxidative stress in silkworms,leading to increased reactive oxygen species levels and toxic effects on the digestive system of silkworms.AgNPs lowered the resistance to oxidative stress,affected cell apoptosis,and induced cell necrosis by regulating related protein metabolism and metabolic pathways in B.mori.