In vitro effects of single and binary mixtures of regulated mycotoxins and persistent organochloride pesticides on steroid hormone production in MA-10 Leydig cell line

Epidemiological studies have shown strong deterioration in male reproductive health globally due to compromised testosterone production leading to altered spermatogenesis and poor sperm quality. However, the effects and mechanisms through which mycotoxins and persistent organochloride pesticides contribute to poor reproductive health in males remain unclear. The effects of single and binary combinations of ochratoxin A, deoxynivalenol, zearalenone, alpha-zearalenol, beta-zearalenol and 1,1,1-trichloro-2,2-bis(p-chlorophenyl) ethane on testicular steroidogenesis were evaluated using the MA-10 Leydig cell line after 48 h of exposure. Zearalenone exposure, especially at 16 μM, had a stimulatory effect on progesterone secretion (4.7 ± 0.48 ng/mL compared to 0.60 ± 0.07 ng/mL in control), but inhibited testosterone production after 48 h compared to the solvent control. Ochratoxin A treatment significantly increased both progesterone and testosterone levels. Combination of alpha-zearalenol with beta-zearalenol showed a synergistic stimulation of progesterone hormone level at 1 and 8 μM. The results presented here show that the MA-10 Leydig cell line is a useful model for assessing the effects of xenoestrogens on testicular steroidogenesis. In addition, the inhibitory effects of zearalenone, alpha-zearalenol and beta-zearalenol on testosterone production was enhanced by co-exposure with 1,1,1-trichloro-2,2-bis(p-chlorophenyl) ethane, further compounding the threat posed by these mycotoxins to male reproductive health

On-chip mouse embryo culture: Evaluation of effects of uterine cells-conditioned media on embryo development and gene expression

Microfluidics has recently been proposed as a method to overcome the limitations of traditional oocyte and embryo culture methods. In this work, we report the use of a microfluidic polydimethylsiloxane device as promising alternative for in vitro embryo culture, and we have evaluated the effects of cells- conditioned media (CM) on embryo development. The microfluidic device was fabricated using traditional soft-lithographic technique. To produce CM, mouse uterine epithelial cells (Creative Bioarray, USA) were cultured in KSOM (Merck Millipore, UK) for 24 h. The CM was used to culture groups of 12, 1 cell murine embryos (B6C3F1xB6D2F1 strain, EmbryoTech, USA) into a microfluidic device. Control embryos were cultured in the device using KSOM. We compared blastocyst rates of embryos cultured in CM with those obtained using KSOM. The effect of treatment on embryo gene expression was assessed in cDNAs generated from individual stage matched, blastocysts using a custom, real time PCR array. Developmental ability of mouse embryos in the presence of CM was significantly higher (p<0.05) in comparison with control media. Blastocyst rates for the CM (n=15 devices, 180 embryos) and control media (n=15 devices, 180 embryos) groups were 68.9% and 45.1%, respectively. qPCR results showed that expression of Makorin Ring Finger Protein (MKRN, p=0.036), DNA Methyltransferase 3β (DNMT3β, p=0.012), DNA (Cytosine -5-)-Methyltransferase 3-Like (DNMT3L, p=0.043), Histone Acetyltransferase 1 (HAT1, p=0.006), Keratin 18 (KRT18, p=0.028), and Ubiquitin Like With PHD And Ring Finger Domains 1 (UHRF1, p=0.043) was significantly different between the treatment groups. Specifically, we observed in the CM group increased expression of DNMT3β and DNMT3L, which play an important role in early embryo development. Those finding revealed that the new microfluidic device supports mouse preimplantation embryo development in vitro. Uterine epithelial cells-conditioned medium has the potential to enhance blastocyst development. Further investigations are required to identify the mechanism of this effect

Human oocyte-derived methylation differences persist in the placenta revealing widespread transient imprinting

Thousands of regions in gametes have opposing methylation profiles that are largely resolved during the post-fertilization epigenetic reprogramming. However some specific sequences associated with imprinted loci survive this demethylation process. Here we present the data describing the fate of germline-derived methylation in humans. With the exception of a few known paternally methylated germline differentially methylated regions (DMRs) associated with known imprinted domains, we demonstrate that sperm-derived methylation is reprogrammed by the blastocyst stage of development. In contrast a large number of oocyte-derived methylation differences survive to the blastocyst stage and uniquely persist as transiently methylated DMRs only in the placenta. Furthermore, we demonstrate that this phenomenon is exclusive to primates, since no placenta-specific maternal methylation was observed in mouse. Utilizing single cell RNA-seq datasets from human preimplantation embryos we show that following embryonic genome activation the maternally methylated transient DMRs can orchestrate imprinted expression. However despite showing widespread imprinted expression of genes in placenta, allele-specific transcriptional profiling revealed that not all placenta-specific DMRs coordinate imprinted expression and that this maternal methylation may be absent in a minority of samples, suggestive of polymorphic imprinted methylation

Imprinted gene expression in hybrids:perturbed mechanisms and evolutionary implications

Diverse mechanisms contribute to the evolution of reproductive barriers, a process that is critical in speciation. Amongst these are alterations in gene products and in gene dosage that affect development and reproductive success in hybrid offspring. Because of its strict parent-of-origin dependence, genomic imprinting is thought to contribute to the aberrant phenotypes observed in interspecies hybrids in mammals and flowering plants, when the abnormalities depend on the directionality of the cross. In different groups of mammals, hybrid incompatibility has indeed been linked to loss of imprinting. Aberrant expression levels have been reported as well, including imprinted genes involved in development and growth. Recent studies in humans emphasize that genetic diversity within a species can readily perturb imprinted gene expression and phenotype as well. Despite novel insights into the underlying mechanisms, the full extent of imprinted gene perturbation still remains to be determined in the different hybrid systems. Here we review imprinted gene expression in intra- and interspecies hybrids and examine the evolutionary scenarios under which imprinting could contribute to hybrid incompatibilities. We discuss effects on development and reproduction and possible evolutionary implications