Philip S. Hjelmborg

Relation between serum xenobiotic-induced receptor activities and sperm DNA damage and sperm apoptotic markers in European and Inuit populations

Persistent organic pollutants (POPs) can interfere with hormone activities and are suspected as endocrine disrupters involved in disorders, e.g. reproductive disorders. We investigated the possible relation between the actual integrated serum xenoestrogenic, xenoandrogenic and aryl hydrocarbon receptor (AhR) activities, and the sperm DNA damage and sperm apoptotic markers of 262 adult males (54 Inuits from Greenland, 69 from Warsaw (Poland), 81 from Sweden, and 58 from Kharkiv (Ukraine)) exposed to different levels of POPs. Xenobiotic-induced receptor activities were determined by receptor-mediated luciferase reporter gene expression. Sperm DNA damage was measured using terminal deoxynucleotidyl transferase-driven dUTP nick labeling assay (TUNEL) and pro- (Fas) and anti-apoptotic (Bcl-xL) markers were determined by immune methods. Different features of xenobiotic-induced receptor activity in serum and sperm DNA fragmentation and apoptotic markers existed between the Inuits and the European Caucasians. Negative correlations between xenobiotic-induced receptor activities and DNA damage were found for Inuits having relatively lower xenoestrogenic, lower dioxin-like activity, and lower sperm DNA damage, but higher xenoandrogenic activity. In contrast, in the European groups, xenobiotic-induced receptor activities were found to be positively correlated with the DNA damage. Further research must elucidate whether altered receptor activities in concerted action with genetic and/or nutrient factors may have protecting effect on sperm DNA damage of the Inuit population.

Xenoandrogenic Activity in Serum Differs across European and Inuit Populations

Background Animal and in vitro studies have indicated that human male reproductive disorders can arise as a result of disrupted androgen receptor (AR) signalling by persistent organic pollutants (POPs). Our aim in the present study was to compare serum xenoandrogenic activity between study groups with different POP exposures and to evaluate correlations to the POP proxy markers 2,2′,4,4′,5,5′-hexachlorobiphenyl (CB-153) and 1,1-dichloro-2,2-bis(p-chlorophenyl)-ethylene (p,p′-DDE). Methods We determined xenoandrogenic activity in the serum fraction containing the lipophilic POPs but free of endogenous hormones. Adult male serum (n = 261) from Greenland, Sweden, Warsaw (Poland), and Kharkiv (Ukraine) was analyzed. Xenoandrogenic activity was determined as the effect of serum extract alone (XAR) and in the presence of the synthetic AR agonist R1881 (XARcomp) on AR transactivated luciferase activity. Results The study groups differed significantly with respect to XARcomp activity, which was increased in the Inuits and decreased in the European study groups; we observed no difference for XAR activity. We found the highest level of the AR antagonist p,p′-DDE in Kharkiv, and accordingly, this study group showed the highest percent of serum samples with decreased XARcomp activities. Furthermore, the percentage of serum samples with decreased XARcomp activities followed the p,p′-DDE serum level for the European study groups. No correlations between serum XAR or XARcomp activities and the two POP markers were revealed. Conclusions The differences in XARcomp serum activity between the study groups suggest differences in chemical exposure profiles, genetics, and/or lifestyle factors.

Semen quality in relation to xenohormone and dioxin-like serum activity among Inuits and three European populations.

Background Semen quality in humans may be influenced by exposure to endocrine-disrupting compounds. Objectives We analyzed associations between semen characteristics and serum xenoestrogen receptor (XER), xenoandrogen receptor (XAR), and aryl hydrocarbon receptor (AhR) transactivity. XER and XAR activity were measured in serum samples cleared for endogenous steroid hormones and AhR activity in raw lipophilic serum extracts free of proteins. Results All together, 319 men from Warsaw (Poland), Greenland, Kharkiv (Ukraine), and Sweden provided semen and blood samples. No strong and consistent associations between xenobiotic activity and semen quality measures were observed in the four populations. However, when the data were combined across populations sperm concentration increased 40% per unit increase in XER activity [95% confidence interval (CI), 1–79%] in the subgroup with XER activity below the reference level. Among subjects with XER activity above the reference level an increase of 14% (95% CI, 2–28%) was found. Furthermore, an increase of 10% motile sperm per unit increase in XER activity below reference level (95% CI, 0.2–20) was found. We are unable to exclude that the associations are chance findings. Conclusion Alteration of XER, XAR, or AhR transactivity within the range found in serum from the general European and Inuit population seems not to markedly deteriorate sperm cell concentration, motility, or morphology in adult men.

Xenohormone transactivities are inversely associated to serum POPs in Inuit.

AbstractBackgroundThe persistent organic pollutants (POPs) are highly lipophilic and resistant to biodegradation and found in e.g. seafood and marine mammals. Greenlandic Inuit have high intake of marine food and thus high POP burden that varies according to local conditions and dietary preference. We do for the very first time report the serum POP related non-steroidal xenohormone activity of Inuit across Greenland. The aims were 1) to determine the integrated xenohormone bioactivities as an exposure biomarker of the actual lipophilic serum POP mixture measuring the effect on estrogen (ER) and androgen receptor (AR) transactivity in citizens from different Greenlandic districts and 2) to evaluate associations to serum POP markers (14 PCBs and 10 pesticides) and lifestyle characteristics.MethodsSerum samples from 121 men and 119 women from Nuuk, Sisimiut and Qaanaaq were extracted using SPE-HPLC fractionation to obtain the serum POP fraction free of endogenous hormones. The serum POP fraction was used for determination of xenohormone transactivity using ER and AR reporter gene assays.ResultsIn overall, the xenohormone transactivities differed between districts as well as between the genders. Associations between the transactivities and age, n-3/n-6 and smoker years were observed. The xenoestrogenic and xenoandrogenic transactivities correlated negatively to the POPs for the combined female and male data, respectively.ConclusionThe non-steroidal xenohormone transactivities can be used as an integrated biomarker of POP exposure and lifestyle characteristics. The actual serum POP mixtures antagonized the age adjusted sex hormone receptor functions. Comparison of different study populations requires in addition to age inclusion of diet and lifestyle factors.

Isolation of Lipophilic Persistent Organic Pollutants From Human Breast Milk

Background: Lipid removal from biological samples can be achieved by addition of concentrated sulfuric acid. However, certain persistent organic pollutants (POPs) such as chlorophenols are decomposed by sulfuric acid treatment and, thus, a more gentle lipid reduction method is needed for extraction of many environmental contaminants from biological samples. Membrane dialysis extraction (MDE) is a non-disruptive method to extract POPs from biological matrices. Methods: Human breast milk samples were spiked with radiolabelled p,p′-dichlorodiphenyl trichloroethane ([C-14]-DDT) as a POP proxy and extracted using solid phase extraction (SPE). The extracts obtained were dialyzed by MDE in low-density polyethylene tubings containing a mixture of n-hexane and dichloromethane for 24 h, 48 h, or 72 h. Results: The lipid content was reduced by 86.2% after one dialysis cycle of 24 h using MDE, and 87.1% recovery of the [C-14]-DDT standard was obtained. The DDT recovery could be further increased up to 96.3% and 98.1% by repeating the dialyses for one or two more cycles, respectively. However, the increased [C-14]-DDT recovery includes a concomitant increase in lipid carryover from 13.8% with one dialysis cycle to 22.1% with three cycles. Conclusion: An SPE procedure for extracting POPs from breast milk and dialytic conditions for isolation of the extracted POP with minimal lipid carryover was established. The method is nondestructive and acceptable recoveries can be obtained within a single solvent shift as demonstrated by spiking standards. The lipid carryover was minimized, and the method may be considered for lipid removal before HPLC or GC analysis of environmental contaminants.