Study On Reproductive Toxicity And Lipid Metabolism Interference Of Perfluorohexane Sulfonate(PFHxS)

Perfluorohexanesulfonic acid(PFHx S)is a functional fluorinated surfactant,which is widely used in daily life and industrial production.In view of its environmental persistence,long-distance migration,high Bioaccumulation and various biological toxicity,PFHx S and its salts and related compounds were included in the list of persistent organic matter elimination under the Stockholm Convention on persistent organic pollutants in June 2022.However,compared with Perfluorooctanesulfonic acid(PFOS),there are fewer toxicological studies on PFHx S,which lacks systematic exploration of its biological safety.In view of the hot research of PFOS and the previous research basis of our research group,this paper focuses on the reproductive toxicity and lipid metabolism interference effects of PFHx S by using in vivo and in vitro experimental models.The main results are as follows:(1)Lipid metabolism interference in mice after PFHx S exposureAfter 14 days of PFHx S exposure,BALB/c mice had weight loss,decreased total food intake,increased blood sugar,and decreased muscle specific gravity,indicating that PFHx S had adverse effects on the growth and development of mice.Abnormal weight of organs and tissues in mice,including liver,brown fat,epididymis fat,inguinal fat,and perirenal fat,suggests that changes in lipid distribution may affect energy metabolism in mice.Further utilizing energy metabolism cages to evaluate the energy metabolism of mice exposed to PFHx S,the results showed an increase in oxygen consumption and disruption of energy metabolism balance in the PFHx S treated group.Pathological sections showed varying degrees of morphological changes in adipose tissue.The results of cold stimulation experiments showed that PFHx S treatment increased adaptive heat production and increased expression of the BAT heat production related gene UCP1.Exposure to PFHx S causes liver steatosis and liver function damage,inducing significant lipid droplet accumulation in the liver,and also altering blood lipid levels.Fluorescence quantitative PCR results showed that genes related to lipid metabolism in mice(ATGL,PEPCK,PPARγ,DGAT,HSL)changed,which was consistent with the accumulation of fat in the liver and the changes of blood lipids.After exposure to PFHx S,there was an accumulation of fat vacuoles in the bone cavity and bone marrow of mice’s femur,indicating that PFHx S may lead to adipogenicity differentiation of bone marrow mesenchymal stem cells,leading to bone loss and possibly osteoporosis.In addition,in the PFHx S-treated group,testicular spermatogenic cells fell off,accompanied by crescent shaped cytoplasm margin,supporting cells vacuolized,and the number of spermatocytes was small,which proved that PFHx S had certain reproductive toxicity,but its related molecular mechanism needed further exploration.(2)Study on the reproductive toxicity of parents of zebrafish and generation transfer effect of long terms low dose exposure of PFHx S with environmental related concentrationsAfter 58 days exposure to F0 zebrafish with environmental related concentrations(0,0.01,and 0.1 μmol/L PFHx S),the gonadal index of female Zebrafish decreased with the increase of concentration.In the 0.1 μmol/L PFHx S treatment group,the ovarian tissue was damaged and the number of mature egg cell decreased,which may lead to the decrease of female spawning.The transcription level of genes related to the hypothalamic pituitary gonadal axis(cyp19a1a,fshr,3βHsd,hsd17β3,lhr)decreased,and accordingly the content of female Zebrafish gonadal sex hormone estradiol(E2)decreased,and the content of follicle stimulating hormone(FSH)also decreased.histological and pathological sections showed that male testis development was also inhibited and mature sperm decreased.Sperm dynamics analysis revealed a decrease in various parameters of sperm motility,which is also the reason for the decrease in fertilization rate of F1 embryos.The decrease in transcription levels of steroid synthesis related genes(cyp17a1,lhr,3βHsd,hsd17β3)and the decrease in testosterone(T)content in male fish indicate that PFHx S may interfere with hormone secretion by regulating the expression of steroid synthesis related genes,thereby affecting reproductive function.In addition,both male and female fish exposed to PFHx S exhibited varying degrees of steatosis in the liver,with changes in the levels of triglycerides(TG)and total cholesterol(T-CHO),further demonstrating the lipid interference effect of PFHx S.It is worth noting that the toxic effects of PFHx S have been passed on from generation to generation.Parental exposure has passed on the toxicity to F1 generation through reproductive activities.F1 generation zebrafish larvae have symptoms such as short time to live,accelerated heartbeat,decreased swimming speed and distance,increased levels of triglycerides in the body,and changed the expression levels of TG synthesis related genes acox1,tpilb,fasn,dgat2 and acaca,All the results showed that the development and lipid metabolism of F1 generation zebrafish were affected by parental exposure.(3)Effects of PFHx S exposure on the proliferation and steroid hormone synthesis of H295 R cellsH295R cells can express 10 key genes involved in human steroid synthesis and metabolism,and are often used to analyze the endocrine disrupting effects of compounds.This chapter takes H295 R cells as the research object to further explore the impact of PFHx S on the biological steroid hormone synthesis pathway and verify the reproductive toxicity of PFHx S.The results showed that after 48 hours of exposure to H295 R cells with different concentrations of PFHx S,the cell survival rate decreased and showed a significant dose-dependent effect.The results of cell cycle analysis showed that H295 R cells stagnated in the S and G2 phases after PFHx S exposure,and the regulation of cell cycle was affected.The expression levels of steroid hormone synthesis related genes HMGR,3βHSD2,CYP21a2,CYP19 and CYP17 were significantly reduced compared to the control group,indicating that PFHx S exposure can lead to abnormal cholesterol and steroid hormone synthesis,leading to fertility disorders.In conclusion,this paper studies the reproductive toxicity and lipid interference effects of PFHx S,explores its potential molecular mechanism,and proves that the toxic effects of PFHx S can be transmitted to the next generation.These research results can provide reliable scientific basis for the biosafety and health risk assessment of PFHx S.