Male Reproductive Toxicities Of Exposure To Carbamate And Pyrethroid Pesticides

PartⅠGenotoxic effects on human spermatozoa of carbaryl-exposureCarbaryl, one of the most important carbamate pesticides, is widelyproduced and used. To explore carbaryl-induced genotoxic effects ofspermatozoa, particularly DNA damage and chromosome aberrations (CA),we firstly examined conventional semen parameters, the progression andmotion parameters of the spermatozoa among 16 carbaryl-exposed workersand 30 internal and extemal control individuals. Sperm DNA damagerepresented as positive percentage of DNA fragmentation was detected by amodified terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) assay. Then numerical CA and aneuploidy ofchromosome X, Y and 18 were investigated by multicolor fluorescence in situhybridization (FISH). The results showed significant differences in thepercentage of sperm abnormality between carbaryl-exposed group and theexternal control group (P=0.008). Mean (±SD) percentage of spermatozoawith fragmented DNA in carbaryl-exposed group (21.04±8.88％) wassignificantly higher than those in the internal (13.36±12.17％) and externalcontrol groups (13.92±7.15％), respectively (P=0.035 and P=0.030).Using FISH, we observed the frequency of sperm sex chromosome disomywas 0.661±0.238％in exposed group, which was significantly higher onlythan that in the external control group (0.386±0.140％) (P=0.001), andcarbaryl-exposed group (0.276±0.126％) had an elevated chromosome 18disomy compared with the internal (0.195±0.094％) and external controlgroups (0.124±0.068％), respectively (P＜0.05 and P＜0.01). In addition,carbaryl-exposed donors had significantly higher sperm nullisomicfrequencies of sex chromosomes and chromosome 18 than the externalcontrols (P＜0.01) but not the internal controls. In summary, the frequenciesof aneuploidy and numerical CA showed significant differences betweenexposed group and control groups (P＜0.05 and/or P＜0.01). Moreover,positive correlation were found between sex chromosome disomy, aneuploidyrate and morphologic abnormalities in spermatozoa of all donors (r=0.564and r=0.555, P＜0.01). Our findings suggested that carbaryl might inducemorphologic abnormalities and genotoxic defects of spermatozoa amongexposed workers by causing DNA fragmentation and numerical CA in spermatogenesis as a potential genotoxicant. These evidences also indicatedthat the spermatotoxicity induced by carbaryl-exposure might be related toadverse reproductive outcomes.PartⅡGenotoxic effects on human spermatozoa of fenvalerate-exposureFenvalerate, a synthetic pyrethroid pesticides, is widely produced and usedworldwide. To explore fenvalerate-induced genotoxic effects, particularlynumerical chromosome aberration (CA), we firstly examined conventionalsemen parameters, the progression and motion parameters of the spermatozoaamong 12 fenvalerate-exposed workers and 30 donors of the internal andexternal control groups. Then numerical CA and aneuploidy of chromosomeX, Y and 18 were investigated by multicolor fluorescence in situ hybridization(FISH). The results showed the significant differences in the percentage ofsperm abnormality between fenvalerate-exposed group and the externalcontrol group (P=0.024). In aneuploid parameters, the frequency (mean±SD)of sex chromosome disomy was 0.742±0.131％in fenvalerate-exposed group,which was significantly higher than those in the internal (0.563±0.135％)and external control group (0.386±0.140％) (P＜0.01), and the frequency ofchromosome 18 disomy in fenvalerate-exposed group (0.326±0.069％) was significantly higher than those in the internal and external control groups(0.195±0.094％and 0.124±0.068％), respectively (P＜0.01). We also foundthe nullisomies of sex chromosomes and chromosome 18 were significantlyhigher than those in the external control group and two control groups,respectively (P＜0.01). The frequencies of aneuploidy and numerical CA wedetected also showed significant differences between exposed group andcontrol groups (P＜0.05 and/or P＜0.01). Moreover, we found the positivecorrelation not only between nullisomic frequencies of these chromosomesand numerical CA rate (r＞0.70, P＜0.01) but also between disomicfrequency of sex chromosomes or aneuploidy rate and sperm abnormality inall donors (r=0.530 and r=0.536, P＜0.01). Our findings suggest thatfenvalerate or its metabolites induced morphologic abnormality and genotoxicdefects of spermatozoa among fenvalerate-exposed workers by causingnumerical CA in spermatogenesis as a special and potential genotoxic agent.PartⅢThe relation between urinary metabolite of pyrethroidpesticides and semen quality in humansPyrethroids, one of the most frequently used insecticides, are widelyapplied in agriculture and daily life. Studies of laboratory animals and limitedhuman data from occupational exposure suggested the associations betweenpyrethroids exposure and male reproductive toxicities, particularly alteredsemen quality. We explored whether internal exposure levels of 3-phenoxybenzoic acid (3-PBA), a general urinary metabolite of pyrethroids,are associated with altered semen quality in Chinese males. Subjects (n=376)were recruited through the clinic for reproductive health after strict screening.By using gas chromatography-mass spectrometry (GC-MS), individualexposures were measured as spot urinary concentrations of 3-PBA adjustedby urinary creatinine (CR). Semen quality was assessed by semen volume,sperm concentration, sperm number per ejaculum, sperm motility, along withnine sperm progression and motion parameters. These semen parameters wereanalyzed as both a continuous measure and dichotomized based on WorldHealth Organization reference values. The comparison group for thedichotomized analysis was men with all four semen parameters at or abovethe reference values. We found that median 3-PBA concentration was 0.879μg/g of CR. There was suggestive association between increasedcreatinine-adjusted 3-PBA quartiles and sperm concentration (ORs for the 1st,2nd, 3rd, and 4th quartiles= 1.00, 1.31, 1.73, and 2.04 respectively; P-valuefor trend=0.027), whereas sperm volume, sperm number per ejaculum andsperm motility were weakly or nonsignificantly associated with 3-PBAquartiles, respectively. The sperm progression and motion parametersassociated with CR-adjusted 3-PBA levels were straight line velocity (VSL; P=0.003) and curvilinear velocity (VCL; P=0.039). These observedassociations between 3-PBA levels and some altered semen quality indicatedthe slight reproductive effects on adult men of pyrethroids exposure. PartⅣThe relation between sperm telomere length and spermaneupioidySperm chromosome aberration, particularly sperm aneuploidy, is one of themost important causes of infertility, abortion and congenital diseases, whiletelomere length is the key point of chromosome stability. It was suggestedthat telomere shortening and erosion may lead to chromosome end to endfusions and genomic instability, with consequent aneuploidy. Tosimultaneously detect sperm telomere length and sperm aneuploidy and theirpossible correlation, we performed new methods 18C-Q-FISH and17T-Q-FISH, composed by human peptide nucleic acid (PNA) telomere probeand centromeric probe of chromosome 18 or subtelomere probe ofchromosome 17 respectively, together with conventional 18XY-FISH. Ourresults suggested that relative sperm telomere length showed better correlationwith aneuploidy rate than directly detected telomere length and the accuracyand stability of 18C-Q-FISH was better than 17T-Q-FISH. By 18C-Q-FISH,we found significantly negative correlation (r=-0.711 and P=0.021)between relative telomere length and aneuploidy rate of spermatozoa. Theseresults directly suggested the correlation of telomere length and aneuploidy inhuman spermatozoa. It was also proposed that 18C-Q-FISH is an effectivemethod for sperm chromosome analysis.