Mathematical Modeling In Low-dose PAEs And Multi-dimensional Environmental Exposures With Male Reproductive Damages

BackgroundPhthalates(PAEs),as one of the most widely distributed pollutants on the earth,have been applied as plasticizers in hundreds of products.PAEs,as endocrine disrupting chemicals,can interfere with the body’s endocrine function.Long-term exposure to PAEs may affect reproductive health,including effects on the semen quality and sex hormones.In view of the harmful effects of PAEs on the health,concerns have been risen around the world.As a result,safety limits for human daily intakes on PAEs have been established.For instance,Environmental Protection Agency(EPA),European Food Safety Authority(EFSA)and our country have established the limits called Reference doses(RfD),Tolerable Daily Intakes(TDI)and Health-based guidance values,respectively.Among all limits,the RfD are relatively strict.However,the existing safety limits of PAEs concern little about the reproductive damages and lack the support of population researches.Therefore,it is necessary to combine population researches with exploring whether the existing safety limits are effective in protecting the male reproductive health.Recent researches have gradually realized that the male reproductive health is regulated by plenty of factors including the environment,the mental and behavioral factors,which is true for local personnel or special groups such as the military.Single factor may have few effects on male reproductive damages.At the same time,the previous research methods might increase the publication biases of the results.Therefore,in recent years,scientists have devoted themselves to exploring the impacts of complex factors on male reproductive damages.However,the available databases showed that methods comprehensively considered multi-dimensional exposure factors to evaluate the male reproductive damages were still lacking.This study based on the Male Reproductive Health in Chongqing College Students(MARHCS)to explore the relationships between low-dose PAEs/ multi-dimensional environmental exposures and male reproductive damages.Firstly,we used the optimal scale regression and Joinpoint regression to explore correlations and threshold doses between PAEs with RfD and male semen parameters/sex hormones.In order to comprehensively analyze the relationships between PAEs and semen parameters/sex hormones,the graph-guided fused lasso(GFLASSO)was used to explore associations between environmental-psychological-behavioral exposure factors(including PAEs)and various male reproductive factors(including semen parameters/sex hormones).Then we constructed a predictive model for each reproductive phenotype through exposure factors.Furthermore,based on the predictive model,the hierarchical clustering was used to identify subgroups of population with reproductive damage characteristics such as endocrine disorders or abnormal semen parameters,and conducted risk assessment in semen parameters/sex hormones caused by PAEs under complex exposure conditions.Contents1.The correlations of low-dose PAEs and semen parameters/sex hormones and the risk assessment on male reproductive damages of PAEs below the RfD.The subjects of this study were 796 male college students recruited in June 2013.Biomarkers including the blood,the urine,and the semen of the subjects were collected.Semen samples were collected in the light of ―the WHO Human Semen Inspection and Processing Laboratory Manual‖(2010 5th Edition)and semen routine parameters such as total sperm numbers,progressive motility and normal morphology were tested.Sex hormones(estrogen,follicle stimulating hormone,luteinizing hormone,prolactin,progesterone,and testosterone)were detected using an automatic immune analyzer.The concentrations of PAEs metabolites in the urine were detected by High Performance Liquid Chromatography-tandem mass Spectrometry(HPLC-MS/MS).For the purpose of assessing the risk of RfD on male reproductive damage,the optimal scaling analysis was used to explore the correlations between the 7 PAEs metabolites(MEP,Mn BP,MBz P,MEHP,MEOHP,MECPP,MEHHP)and semen parameters/sex hormones.Joinpoint regression was used to identify whether an inflection point(threshold dose)appeared between PAEs and reproductive phenotypes.For PAEs with the threshold,the method proposed by Wittassek et al.was used to calculate the intake doses.Finally,we compared the intake doses with RfD to assess the risk of RfD on male reproductive damage.2.Correlations between PAEs and semen parameters/sex hormones under the multiple exposures of environmental/behavioral/psychological factors.The reproductive health status and its potential factors were investigated through the social-psychological-behavior questionnaire,biomarkers and physical measurements among the men.A total of 138 environmental-psychological-behavioral factors including PAEs were collected,and 32 reproductive phenotypes such as semen parameters,sex hormones and sperm chromatin were collected.The GFLASSO was applied to comprehensively analyze the correlations between each exposure factor and each reproductive biomarker.A total of 220 correlations which had the largest standardized regression coefficients(negative coefficients below the 2.5th percentile and positive coefficients over the 97.5th percentile)were selected.The predictive score for each reproductive biomarker was calculated.Hierarchical clustering and T-distributed stochastic neighbor embedding(t-SNE)were used to divide the population into subgroups by the predictive scores.Then we compared the levels of semen parameters and sex hormones among subgroups,and found out the PAEs exposure levels in subgroups with abnormal semen quality or sex hormones.Results1.The correlations of low-dose PAEs and semen parameters/sex hormones and the risk assessment on male reproductive damages of PAEs below the RfD.The detection rate of BBz P in the MARHCS was so low that it couldn’t fit the standards for the further study.As for the other 3 of PAEs(DEP,Dn BP,DEHP),the daily intakes were generally lower than corresponding RfD(Proportions of the population above the limit were DEP: 0%、Dn BP:0.8% and DEHP: 1.6%),so as for TDIs(percentage above limit were Dn BP2.3% and DEHP 0.6%)and Health-based guidance values(percentage above limit were DEP:0.2%、Dn BP: 2.3% and DEHP: 0.6%),respectively.There were 12 correlations between the PAEs metabolites and semen parameters/sex hormones: the metabolite MEP of DEP was related to progressive motility,normal morphology and testosterone(P<0.01);Dn BP metabolite Mn BP was related to normal morphology(P<0.05);Among the four metabolites of DEHP,MEHHP and MECPP were correlated with estrogen(P<0.05),MEOHP and MEHP were correlated with estrogen and progesterone(P<0.01),sum of DEHP metabolites was related to the levels of progesterone and estrogen(P<0.01).Inflection points were found in the correlations between DEHP total metabolites and few parameters: threshold doses were lower than corresponding RfD.2.Correlations between PAEs and semen parameters/sex hormones under the multiple exposures of environmental/behavioral/psychological factors.The results of GFLASSO suggested that each reproductive biomarker was related to a variety of exposure factors,and the results in the population which exposures level of PAEs was lower than RfD were consistent with the whole population.Among the total of 220 associations selected form GFLASSO regression coefficients,there were 32 associations between PAEs and reproductive biomarkers.All the predictive values were correlated with the reproductive biomarkers.Then we clustered the men using the predictive scores into five subgroups with the comprehensive reproductive features representing disrupted spermatogenesis and abnormal sperm morphology;low sperm motility and DNA fragmentation;chromatin immaturity;aberrant endocrine;DNA strand breakage.Subgroup 4mainly showed abnormalities in estrogen and progesterone and total PAEs metabolites was the highest concentration of PAEs metabolites in subgroup 4.ConclusionsThe present study concerned on association and safety study between the low-dose PAEs and semen parameters/sex hormones,showed that(1)there were 12 correlations between PAEs and sex hormones/semen parameters.Several inflection points were found in the correlation between DEHP and progesterone/estrogen and the corresponding daily intakes of the men were much lower than RfD(20 μg/(kg·d)).Moreover,other PAEs did not find the inflection point,suggesting that PAEs might have an impact on male semen parameters/sex hormones.All this study represented that the existing safety limits might be insufficient in protecting male reproductive health.(2)We used GFLASSO to explore the correlations between PAEs and other multi-exposures with semen parameters/sex hormones.We established the predictive models for multiple biomarkers based on the exposures and the models could be applied to identify the damages features among the men.This study proposed a new approach to identify the male reproductive damages among the population based on the multiple factors which had an importance for preventing the male reproductive damages regardless in the community or in other special groups.