Effect Of The Reproductive Toxicity Of GONPs Exposure To Bombyx Mori

The silkworm has a history of more than 8,500 years of domestication.It is a completely metamorphic economic insect.It has four developmental stages:eggs,larvae,pupae,and adults(moths).Due to the short generation cycle and small individuals suitable for large-scale breeding,the number of eggs laid by adults is about 500.The silkworm is also a lepidopteran model insect.The number of eggs laid by the silkworm is related to the genetics of the species,and is also affected by nutritional and environmental conditions.After a long period of indoor domestication,silkworm is less resistant to the external environment,and nanomaterials and chemical pesticides will cause adverse effects on them.Graphene oxide Nanoparticulates(GONPs)are widely used due to their unique properties,and the biosafety assessment of their environmental residues received widespread attention.This paper used the silkworm as a model organism to study the effects of GONPs on the reproduction of the silkworm from the individual and cellular levels.The main results are as follows:1.The effect of nano-graphene oxide on oxidative stress of silkwormUsing the 5th-instar silkworm(Jingsong)as experimental material,the oxidative stress level and body weight changes of the silkworm after feeding GONPs were studied,and the transcription level of key genes in the CncC/Keap1 signaling pathway and the activity of downstream antioxidant enzymes were analyzed.The results showed that when the GONPs concentration was greater than 25 mg/L,the body weight of the GONPs treatment group decreased significantly(P≤0.05),and there was a dose effect.The level of oxidative stress after 25 mg/L GONPs fed with silkworm was measured.The peroxide level of hemolymph and ovarian tissues changed significantly,and the reactive oxygen species(ROS)and peroxides in hemolymph tissues Hydrogen(H2O2)and malondialdehyde(MDA)were significantly upregulated by 1.16,1.50,and 1.63 times(P<0.05),respectively,and ROS,H2O2,and MDA in ovarian tissue were significantly upregulated by 3.18,3.03,and 12.07 times,respectively(P≤0.05).Further research found that Keapl was significantly upregulated by 1.43 times(P≤0.05)in hemolymph;CncC was no significantly.The transcription levels of CncC and Keapl in the ovary were significantly up-regulated by 1.53 and 1.21 times(P≤0.05).Its downstream antioxidant enzymes activities(TPX,SOD,CAT)were also significantly up-regulated(P≤0.05).The results showed that after feeding GONPs(25 mg/L),the oxidative stress level in the silkworm increased,the CncC/Keap1 signaling pathway was activated,and the antioxidant enzyme activity level increased.2.The effect of nano-graphene oxide on silkworm ovaryIn order to further study the effects of GONPs on silkworm reproduction,this article studied the effects of GONPs on the gonads of the silkworm.The results showed that after GONPs(25mg/L)fed with silkworm larvae,the female gonad index decreased significantly by 40.84%(P≤0.001),but there was no significant difference in the impact on male gonads;pathological sections showed that the ovarian tissue in the GONPs treatment group The number of oocytes and oocytes decreased,and the ultrastructure of electron microscopy showed that vacuoles and peroxisomes appeared in the follicle cells;qRT-PCR was used to analyze the genes related to the growth and development of the silkworm ovary(Vg,Ovo,Sxl-S,Sxl-L and Otu),the results showed that they were 0.61,0.65,0.75,0.72,and 0.42 times of the control group,respectively,and they were all significantly decreased(P≤0.05);after mating of adults,the egg production of GONPs treatment group decreased by 52.25%,respectively(P≤0.05).The results showed that feeding GONPs has reproductive toxicity to silkworm,and has a greater impact on females.It will cause damage to ovarian tissues,cause developmental obstruction,and ultimately reduce silkworm egg production.3.The effect of nano-graphene oxide on BmN cellsIn order to study the toxicity of GONPs at the cellular level,this paper also studied the damage of GONPs to BmN cells after exposure.The results showed that higher concentrations of GONPs(≥25 mg/L)induced oxidative stress in BmN cells,ROS accumulation and DNA damage,and significantly decreased the survival rate of BmN cells(P≤0.05).The transcription levels of antioxidant enzyme genes(TPX,CAT,SOD)were significantly up-regulated by 1.40,1.42 and 1.33 times(P≤0.05).This study shows that GONPs have a toxic effect on the reproduction of female silkworm.GONPs exposure caused the accumulation of ROS,oxidative stress in germ cells,damage to ovarian tissues,and hindered development,which may lead to the decrease in egg production.The results of the study provide a reference for the evaluation of the reproductive toxicity of GONPs environmental residues.