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Expression analysis of small noncoding RNA (sRNA), including microRNA, piwi-interacting RNA, small rRNA-derived RNA, and tRNA-derived small RNA, is a novel and quickly developing field. Despite a range of proposed approaches, selecting and adapting a particular pipeline for transcriptomic analysis of sRNA remains a challenge. This paper focuses on the identification of the optimal pipeline configurations for each step of human sRNA analysis, including reads trimming, filtering, mapping, transcript abundance quantification and differential expression analysis. Based on our study, we suggest the following parameters for the analysis of human sRNA in relation to categorical analyses with two groups of biosamples: (1) trimming with the lower length bound = 15 and the upper length bound = Read length - 40% Adapter length; (2) mapping on a reference genome with bowtie aligner with one mismatch allowed (-v 1 parameter); (3) filtering by mean threshold > 5; (4) analyzing differential expression with DESeq2 with adjusted p-value < 0.05 or limma with p-value < 0.05 if there is very little signal and few transcripts.
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ARN Pequeño no Traducido , Humanos , Benchmarking , Secuenciación de Nucleótidos de Alto Rendimiento , ARN Pequeño no Traducido/genética , RNA-Seq , Análisis de Secuencia de ARNRESUMEN
STUDY QUESTION: Are peripubertal blood lead levels (BLLs) associated with semen parameters and serum reproductive hormones among young Russian men? SUMMARY ANSWER: We observed a suggestion of lower ejaculate volume with higher peripubertal BLL but no associations of BLLs with reproductive hormones measured throughout adolescence or with other sperm parameters measured at adulthood. WHAT IS KNOWN ALREADY: Lead is a known reproductive toxicant and endocrine disruptor. Previous literature has shown associations between high lead exposure and poorer semen quality both in occupationally and environmentally exposed men. However, to our knowledge, no longitudinal studies have explored the association of childhood lead exposure with semen parameters and reproductive hormones in young men. STUDY DESIGN, SIZE, DURATION: The Russian Children's Study is a prospective cohort study that enrolled 516 boys at age 8-9 years in 2003-2005 and followed them annually for 10 years. BLLs were measured at entry and lifestyle and health questionnaires were completed. Reproductive hormones were measured in blood samples collected every 2 years. PARTICIPANTS/MATERIALS, SETTING, METHODS: Among the 516 boys enrolled, 481 had BLLs measured at entry. Of these, 453 had at least one measurement of serum testosterone, follicle stimulating hormone (FSH) or luteinizing hormone (LH) (median = 5 samples per boy) and 223 had semen samples collected â¼10 years after enrolment. Semen assessment included ejaculated volume, sperm concentration, progressive motility and total sperm count, and parameters were categorized using published andrology standards for low semen quality based on sperm count and motility. Linear mixed models were used to examine the associations of log-transformed BLLs (and BLL categories) with reproductive hormones and semen parameters, adjusting for potential confounders. MAIN RESULTS AND THE ROLE OF CHANCE: Among the 223 young men with peripubertal BLLs and at least one semen sample (total samples = 438), the median (interquartile range) BLL was 3 (2, 5) µg/dl and 27% had BLL ≥5 µg/dl. Overall, 49% of the semen samples fell below reference levels for sperm count and/or motility. Men with peripubertal BLL ≥5 µg/dl had significantly lower ejaculated volume than those with BLL <5 µg/dl (mean = 2.42 vs 2.89 ml, P = 0.02), but this difference was attenuated in adjusted models (mean = 2.60 vs 2.83 ml, P = 0.25). No associations were observed between BLL measured at age 8-9 years and reproductive hormone levels or sperm parameters, including sperm concentration, total count, progressive motility and total progressive motile sperm count, or with the probability of having low semen quality based on sperm count/motility. LIMITATIONS, REASONS FOR CAUTION: Only a subset of the original cohort participated in the semen quality portion of the study, although inverse probability weighting was used to account for possible selection bias. BLLs were only measured at a single time in peripuberty, and other exposure time periods, including later or longer-term childhood exposure, may be more predictive of semen quality. The young men were also exposed to other chemical contaminants before and during pubertal development. WIDER IMPLICATIONS OF THE FINDINGS: While semen volume often receives less attention than other sperm parameters, it is an important component of male fertility. Additional prospective studies covering different exposure windows and including other seminal plasma biomarkers are warranted to explore our finding of potentially lower ejaculated volume with higher BLLs and to confirm the lack of associations for other semen parameters among youth exposed to environmental BLLs. STUDY FUNDING/COMPETING INTEREST(S): Funding was provided through grants R01ES0014370 and P30ES000002 from the National Institute of Environmental Health Sciences, grant R82943701 from the U.S. Environmental Protection Agency, and grant 18-15-00202 from the Russian Science Foundation (O.S and Y.D.). All authors report no competing interests. TRIAL REGISTRATION NUMBER: N/A.
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Plomo , Semen , Adolescente , Adulto , Niño , Estudios de Cohortes , Humanos , Hormona Luteinizante , Masculino , Estudios Prospectivos , Análisis de Semen , Recuento de Espermatozoides , Motilidad EspermáticaRESUMEN
BACKGROUND: Although phthalate exposures have been associated with adverse effects on male reproductive health, few studies have explored longitudinal associations with male pubertal development. OBJECTIVES: We examined the association of prepubertal urinary concentrations of phthalate metabolites with age at pubertal onset in a prospective cohort of Russian boys. METHODS: At enrollment at ages 8-9 years, medical history, dietary, and demographic information was collected. At entry and annually, physical examinations and pubertal staging [Genitalia (G), Pubarche (P), and testicular volume (TV, in ml)] were conducted and spot urines were collected. Prepubertal urine samples (defined as either TV = 1, 2 and G = 1, 2 or TV = 3 and G = 1) were pooled for each boy and phthalate metabolite concentrations were quantified using isotope dilution LC-MS/MS at Moscow State University. We measured 15 metabolites including those from anti-androgenic parent phthalates (AAPs) such as di (2-ethylhexyl) (DEHP) and di-isononyl (DiNP) phthalates as well as monobenzyl (MBzP), mono-n-butyl (MnBP), and mono-isobutyl (MiBP) metabolites. We calculated the molar sums of DEHP (∑DEHP), DiNP (∑DiNP), and AAP (∑AAP) metabolites. Separate interval-censored models were used to assess associations of quartiles of prepubertal phthalate metabolites with each pubertal onset indicator, G2+, P2+ and TV > 3 mL, adjusted for covariates and urine specific gravity. RESULTS: 304 boys had 752 prepubertal urine samples (median 2, range: 1-6) for pooling. In adjusted models, higher urinary AAPs were consistently associated with later pubertal onset (P2) with mean shifts ranging from 8.4 to 14.2 months for the highest versus lowest quartiles. Significantly later onset for G2 and TV > 3 mL was observed for higher versus lower quartiles of MiBP, MBzP, ∑DEHP and ∑DiNP. CONCLUSIONS: On average, boys with higher concentrations of prepubertal urinary AAPs had later pubertal onset by six months to over a year. The impact of AAPs on timing of male puberty may be attributable to disruption of androgen-dependent biological pathways.
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Dietilhexil Ftalato , Contaminantes Ambientales , Ácidos Ftálicos , Antagonistas de Andrógenos , Niño , Cromatografía Liquida , Exposición a Riesgos Ambientales/análisis , Contaminantes Ambientales/orina , Humanos , Masculino , Ácidos Ftálicos/orina , Estudios Prospectivos , Espectrometría de Masas en TándemRESUMEN
Polybrominated diphenyl ethers (PBDE) are a group of flame retardants used in a variety of artificial materials. Despite being phased out in most industrial countries, they remain in the environment and human tissues due to their persistence, lipophilicity, and bioaccumulation. Populational and experimental studies demonstrate the male reproductive toxicity of PBDEs including increased incidence of genital malformations (hypospadias and cryptorchidism), altered weight of testes and other reproductive tissues, altered testes histology and transcriptome, decreased sperm production and sperm quality, altered epigenetic regulation of developmental genes in spermatozoa, and altered secretion of reproductive hormones. A broad range of mechanistic hypotheses of PBDE reproductive toxicity has been suggested. Among these hypotheses, oxidative stress, the disruption of estrogenic signaling, and mitochondria disruption are affected by PBDE concentrations much higher than concentrations found in human tissues, making them unlikely links between exposures and adverse reproductive outcomes in the general population. Robust evidence suggests that at environmentally relevant doses, PBDEs and their metabolites may affect male reproductive health via mechanisms including AR antagonism and the disruption of a complex network of metabolic signaling.
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Retardadores de Llama , Éteres Difenilos Halogenados , Humanos , Masculino , Éteres Difenilos Halogenados/toxicidad , Epigénesis Genética , Semen/metabolismo , Retardadores de Llama/toxicidadRESUMEN
In the modern world, metabolic syndrome is one of the major health problems. Heredity, overeating, and a sedentary lifestyle are believed to be the main predisposing factors for its development. However, recent data indicate that gut microbiota plays a significant role in metabolic profile formation. In 2004, Derrien et al. isolated and characterized the bacterium Akkermansia muciniphila, which lives mainly in the human intestine and has the ability to utilize intestinal mucin. It proved to be a good candidate for the role of a new-generation probiotic due to its ability to improve the laboratory and physical indicators associated with metabolic syndrome and type 2 diabetes in mice and humans. In this review, we describe the basic microbiological characteristics of this bacterium, its main habitats, clinical effects after oral administration, and different ways of influencing the digestive tract. All these data allow us to understand the mechanism of its beneficial effects, which is important for its future introduction into the treatment of the metabolic syndrome.
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Microbioma Gastrointestinal , Interacciones Microbiota-Huesped , Akkermansia/efectos de los fármacos , Akkermansia/fisiología , Akkermansia/ultraestructura , Antibacterianos/farmacología , Permeabilidad de la Membrana Celular , Susceptibilidad a Enfermedades , Farmacorresistencia Bacteriana , Matriz Extracelular , Vesículas Extracelulares/metabolismo , Regulación Bacteriana de la Expresión Génica , Humanos , Síndrome Metabólico/etiología , Síndrome Metabólico/metabolismo , Mucinas/metabolismo , Especificidad de Órganos , Consumo de Oxígeno , Probióticos , SimbiosisRESUMEN
Probiotics are widely used for treatment of various human diseases, and their spectrum is not limited by intestinal diseases only. That is why there is a growing interest in the development of new probiotics that can modify intestinal microbiome in accordance with the needs for treatment. In 2004, Akkermansia muciniphila was discovered, and later it was shown to prevent the development of metabolic syndrome and diabetes mellitus in mice. Nevertheless, before using it for treatment, conditions necessary for its growth need to be identified. In particular, certain carbohydrates, including amino sugars, such as N-acetylglucosamine and N-acetylgalactosamine, were discovered to be necessary for successful cultivation of A. muciniphila in vitro. This is not surprising, since the natural habitat of A. muciniphila is intestinal mucin, which contains different amino sugars. Besides, ways of A. muciniphila protection from harmful factors on the way to the intestine have been developed. In addition, prebiotics such as oligosaccharides, polyphenols, as well as metformin used for diabetes mellitus treatment can promote its growth in the intestine. Finally, there is the first evidence of A. muciniphila administration to humans, which confirms the safety of its use and describes positive metabolic effects. Overall, these data suggest the possibility of an early introduction of this next-generation probiotic into clinical practice.
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Enfermedades Intestinales/terapia , Probióticos/uso terapéutico , Akkermansia/crecimiento & desarrollo , Akkermansia/fisiología , Animales , Microbioma Gastrointestinal , Humanos , Ratones , Mucinas/metabolismo , Oligosacáridos/metabolismoRESUMEN
Advanced paternal age at fertilization is a risk factor for multiple disorders in offspring and may be linked to age-related epigenetic changes in the father's sperm. An understanding of aging-related epigenetic changes in sperm and environmental factors that modify such changes is needed. Here, we characterize changes in sperm small non-coding RNA (sncRNA) between young pubertal and mature rats. We also analyze the modification of these changes by exposure to environmental xenobiotic 2,2',4,4'-tetrabromodiphenyl ether (BDE-47). sncRNA libraries prepared from epididymal spermatozoa were sequenced and analyzed using DESeq 2. The distribution of small RNA fractions changed with age, with fractions mapping to rRNA and lncRNA decreasing and fractions mapping to tRNA and miRNA increasing. In total, 249 miRNA, 908 piRNA and 227 tRNA-derived RNA were differentially expressed (twofold change, false discovery rate (FDR) p ≤ 0.05) between age groups in control animals. Differentially expressed miRNA and piRNA were enriched for protein-coding targets involved in development and metabolism, while piRNA were enriched for long terminal repeat (LTR) targets. BDE-47 accelerated age-dependent changes in sncRNA in younger animals, decelerated these changes in older animals and increased the variance in expression of all sncRNA. Our results indicate that the natural aging process has profound effects on sperm sncRNA profiles and this effect may be modified by environmental exposure.
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Envejecimiento/fisiología , Exposición a Riesgos Ambientales , Retardadores de Llama/toxicidad , ARN Pequeño no Traducido/genética , Espermatozoides/metabolismo , Animales , Animales Recién Nacidos , Exposición a Riesgos Ambientales/efectos adversos , Exposición a Riesgos Ambientales/análisis , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Masculino , Parto/efectos de los fármacos , Parto/genética , Parto/metabolismo , Edad Paterna , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Efectos Tardíos de la Exposición Prenatal/genética , Efectos Tardíos de la Exposición Prenatal/metabolismo , ARN Pequeño no Traducido/metabolismo , Ratas , Ratas Wistar , Espermatozoides/efectos de los fármacos , Factores de TiempoRESUMEN
Organochlorine pesticides (OCPs) have been linked to adult metabolic disorders; however, few studies have examined these associations in childhood. We prospectively evaluated the associations of baseline serum OCPs (hexachlorobenzene, ß-hexachlorocyclohexane, and p,p'-dichlorodiphenyldichloroethylene) in Russian boys with subsequent repeated measurements of serum glucose, insulin, lipids, leptin, and calculated homeostatic model assessment of insulin resistance (IR). During 2003-2005, we enrolled 499 boys aged 8-9 years in a prospective cohort; 318 had baseline serum OCPs and serum biomarkers measured at ages 10-13 years. Multivariable generalized estimating equation and mediation regression models were used to examine associations and direct and indirect (via body mass index (BMI) (weight (kg)/height (m)(2))) effects of prepubertal OCP tertiles and quintiles with biomarkers. In multivariable models, higher p,p'-dichlorodiphenyldichloroethylene (quintile 5 vs. quintile 1) was associated with lower leptin, with relative mean decreases of 61.8% (95% confidence interval: 48.4%, 71.7%) in models unadjusted for BMI and 22.2% (95% confidence interval: 7.1%, 34.9%) in models adjusted for BMI; the direct effect of p,p'-dichlorodiphenyldichloroethylene on leptin accounted for 27% of the total effect. IR prevalence was 6.6% at ages 12-13 years. Higher hexachlorobenzene (tertile 3 vs. tertile 1) was associated with higher odds of IR in models adjusted for BMI (odds ratio = 4.37, 95% confidence interval: 1.44, 13.28). These results suggest that childhood OCPs may be associated with IR and lower leptin.
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Hidrocarburos Clorados/sangre , Resistencia a la Insulina , Síndrome Metabólico/sangre , Plaguicidas/sangre , Adolescente , Biomarcadores/sangre , Glucemia/análisis , Niño , Colesterol/sangre , Humanos , Insulina/sangre , Leptina/sangre , Masculino , Estudios Prospectivos , Análisis de Regresión , Triglicéridos/sangreRESUMEN
PURPOSE: Varicoceles can be a source of elevated seminal oxidative stress (OS) and sperm DNA fragmentation (SDF). However, it remains unclear whether varicocele repair (VR) could reduce these parameters. This systematic review and meta-analysis (SRMA) aims to investigate the impact of VR on SDF and seminal malondialdehyde (MDA). MATERIALS AND METHODS: A literature search was performed in Scopus, PubMed, Ovid, Embase, and Cochrane databases. This SRMA included randomized controlled trials and observational studies reporting the pre- and postoperative levels of SDF and seminal OS in infertile men with clinical varicocele that underwent VR. Subgroup analyses included techniques of VR and SDF testing. The effect size was expressed as standardized mean difference (SMD). RESULTS: Out of 1,632 abstracts assessed for eligibility, 29 studies with 1,491 infertile men were included. The analysis showed a significant reduction in SDF after VR, compared to preoperative values (SMD -1.125, 95% confidence interval [CI] -1.410, -0.840; p<0.0001) with high inter-study heterogeneity (I²=90.965%). Reduction in SDF was evident with microsurgical technique and non-microsurgical inguinal approaches (SMD -1.014, 95% CI -1.263, -0.765; p<0.0001, and SMD -1.495, 95% CI -2.116, -0.873; p<0.0001), respectively. Reduction in SDF was significant irrespective of testing was done by sperm chromatin dispersion (SMD -2.197, 95% CI -3.187, -1.207; p<0.0001), sperm chromatin structure assay (SMD -0.857, 95% CI -1.156, -0.559; p<0.0001) or TUNEL (SMD -1.599, 95% CI -2.478, -0.719; p<0.0001). A significant decrease in seminal MDA levels was observed following VR (SMD -2.450, 95% CI -3.903 to -0.997, p=0.001) with high inter-study heterogeneity (I²=93.7%). CONCLUSIONS: Using pre- and post-intervention data, this SRMA indicates a significant reduction in SDF and seminal MDA levels in infertile men with clinical varicocele treated with VR. These findings may have important implications for the future management of this selected group of infertile patients.
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BACKGROUND: Modern reproductive behavior in most developed countries is characterized by delayed parenthood. Older gametes are generally less fertile, accumulating and compounding the effects of varied environmental exposures that are modified by lifestyle factors. Clinicians are primarily concerned with advanced maternal age, while the influence of paternal age on fertility, early development and offspring health remains underappreciated. There is a growing trend to use assisted reproductive technologies for couples of advanced reproductive age. Thus, the number of children born from older gametes is increasing. OBJECTIVE AND RATIONALE: We review studies reporting age-associated epigenetic changes in mammals and humans in sperm, including DNA methylation, histone modifications and non-coding RNAs. The interplay between environment, fertility, ART and age-related epigenetic signatures is explored. We focus on the association of sperm epigenetics on epigenetic and phenotype events in embryos and offspring. SEARCH METHODS: Peer-reviewed original and review articles over the last two decades were selected using PubMed and the Web of Science for this narrative review. Searches were performed by adopting the two groups of main terms. The first group included 'advanced paternal age', 'paternal age', 'postponed fatherhood', 'late fatherhood', 'old fatherhood' and the second group included 'sperm epigenetics', 'sperm', 'semen', 'epigenetic', 'inheritance', 'DNA methylation', 'chromatin', 'non-coding RNA', 'assisted reproduction', 'epigenetic clock'. OUTCOMES: Age is a powerful factor in humans and rodent models associated with increased de novo mutations and a modified sperm epigenome. Age affects all known epigenetic mechanisms, including DNA methylation, histone modifications and profiles of small non-coding (snc)RNA. While DNA methylation is the most investigated, there is a controversy about the direction of age-dependent changes in differentially hypo- or hypermethylated regions with advanced age. Successful development of the human sperm epigenetic clock based on cross-sectional data and four different methods for DNA methylation analysis indicates that at least some CpG exhibit a linear relationship between methylation levels and age. Rodent studies show a significant overlap between genes regulated through age-dependent differentially methylated regions and genes targeted by age-dependent sncRNA. Both age-dependent epigenetic mechanisms target gene networks enriched for embryo developmental, neurodevelopmental, growth and metabolic pathways. Thus, age-dependent changes in the sperm epigenome cannot be described as a stochastic accumulation of random epimutations and may be linked with autism spectrum disorders. Chemical and lifestyle exposures and ART techniques may affect the epigenetic aging of sperm. Although most epigenetic modifications are erased in the early mammalian embryo, there is growing evidence that an altered offspring epigenome and phenotype is linked with advanced paternal age due to the father's sperm accumulating epigenetic changes with time. It has been hypothesized that age-induced changes in the sperm epigenome are profound, physiological and dynamic over years, yet stable over days and months, and likely irreversible. WIDER IMPLICATIONS: This review raises a concern about delayed fatherhood and age-associated changes in the sperm epigenome that may compromise reproductive health of fathers and transfer altered epigenetic information to subsequent generations. Prospective studies using healthy males that consider confounders are recommended. We suggest a broader discussion focused on regulation of the father's age in natural and ART conceptions is needed. The professional community should be informed and should raise awareness in the population and when counseling older men.
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Epigénesis Genética , Espermatozoides , Masculino , Animales , Niño , Humanos , Anciano , Estudios Prospectivos , Estudios Transversales , Espermatozoides/metabolismo , Mamíferos/genética , ARN no Traducido , ADNRESUMEN
Background: Peripubertal concentrations of serum dioxins and polychlorinated biphenyls (PCBs) have demonstrated associations with altered age of pubertal onset and sexual maturity in boys, but associations with pubertal progression have received less attention. Methods: The Russian Children's Study is a prospective cohort of 516 boys enrolled in 2003-2005 at age 8 or 9 and followed annually up to 19 years of age. Serum concentrations of dioxin-like toxic equivalents (TEQs), polychlorinated dibenzodioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and non-dioxin-like PCBs (NDL-PCBs) and whole blood lead levels (BLLs) were quantified from blood samples collected at study entry (age 8-9). Testicular volume (TV) was assessed annually using a Prader orchidometer. Pubertal trajectories were identified by applying Group-Based Trajectory Models (GBTMs) to TV measured from ages 8-19. Associations of peripubertal serum TEQs, PCDDs, PCDFs, and NDL-PCBs with specific progression trajectories were modeled using multinomial logistic regression, adjusting for each boy's birthweight, and for BLL, body mass index and nutritional factors at study entry. Results: Among 489 eligible boys with available exposure measures, we identified three pubertal trajectories using GBTMs: slower (34% of boys), moderate (48%) and faster (18%). Boys with higher peripubertal serum TEQs had higher adjusted odds of being in the moderate versus faster trajectory (adjusted odds ratio (aOR) 1.79, 95% CI 1.01, 3.13) and the slower versus faster trajectory (aOR 1.52, 95% CI 0.82, 2.78) per 1 log unit increase in serum TEQs. Boys with higher peripubertal serum PCDFs had higher adjusted odds of being in the moderate compared to the faster trajectory (aOR 1.92, 95% CI 1.20, 3.03) and of being in the slower versus the faster trajectory (aOR 1.42, 95% CI 0.91, 2.33) per 1 log unit increase. Boys with higher NDL-PCBs had higher adjusted odds of being in the faster trajectory versus the moderate (aOR 2.56, 95% CI 0.91-7.20) or slower (aOR 3.31, 95% CI 1.07, 10.25) trajectory. Boys with higher blood lead levels also had higher adjusted odds of being in the slower trajectory of pubertal progression, compared to either the faster (aOR 1.47, 95% CI 0.89, 2.44) or moderate (aOR 1.20, 95% CI 0.83, 1.75) trajectories, per 1 log unit increase in BLL, although these associations did not attain statistical significance. Conclusion: Boys' peripubertal exposure to dioxins and certain PCBs may alter pubertal progression.
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Dioxinas , Bifenilos Policlorados , Dibenzodioxinas Policloradas , Masculino , Niño , Humanos , Plomo , Estudios Prospectivos , Dibenzofuranos Policlorados , Procarbazina , Federación de RusiaRESUMEN
Transcriptomics analysis of various small RNA (sRNA) biotypes is a new and rapidly developing field. Annotations for microRNAs, tRNAs, piRNAs and rRNAs contain information on transcript sequences and loci that is vital for downstream analyses. Several databases have been established to provide this type of data for specific RNA biotypes. However, these sources often contain data in different formats, which makes the bulk analysis of several sRNA biotypes in a single pipeline challenging. Information on some transcripts may be incomplete or conflicting with other entries. To overcome these challenges, we introduce ITAS, or Integrated Transcript Annotation for Small RNA, a filtered, corrected and integrated transcript annotation containing information on several types of small RNAs, including tRNA-derived small RNA, for several species (Homo sapiens, Rattus norvegicus, Mus musculus, Drosophila melanogaster, Caenorhabditis elegans). ITAS is presented in a format applicable for the vast majority of bioinformatic transcriptomics analysis, and it was tested in several case studies for human-derived data against existing alternative databases.
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Background: Women of reproductive age are recommended to consume folic acid and other supplements before conception and during pregnancy. We aimed to investigate the association of the serum folate and total magnesium (Mg) and calcium (Ca) levels before ovarian stimulation with the outcomes of assisted reproductive technology (ART) in normogonadotropic women. Methods: We used a subanalysis of data obtained from a multicentre, randomized prospective study (NCT03088137). A total of 110 normogonadotropic, non-advanced aged, non-obese women with tubal and/or male infertility factors were enrolled for the single fresh ovarian stimulation GnRH antagonist cycle. The main outcome measures were the total oocyte yield, mature oocytes, fertilization rate, biochemical, clinical pregnancy, and live birth. Multivariable generalized linear models adjusted for covariates were used with a Poisson distribution and the log link function for adjusted oocyte counts, and a binomial distribution and the log link function were used for adjusted clinical ART outcomes. Results: The medians (interquartile range (IQR)) were as follows: baseline serum folate, 20.55 ng/ml (10.8, 32.9); Mg, 19.4 mg/L (18.7, 20.7); Ca, 94 mg/L (91.2, 96.4); and Ca/Mg ratio, 4.78 (4.55, 5.02). Women with higher serum folate concentrations (Q4≥33.0 ng/ml) had significantly lower total numbers of oocytes retrieved (adjusted mean (95% CI) 9.2 (7.6-11.3) vs 12.9 (10.9-15.4, p-trend=0.006)) and lower odds ratios (ORs) (95% CI) of 0.12 (0.02, 0.79) for clinical pregnancy and 0.10 (0.01, 0.70) for live birth compared with women in the lowest quartile (<10.8 ng/ml), all p-trend<0.001. Women in the highest Ca/Mg ratio quartile (≥5.02) had ORs (95% CI) of 6.58 (1.31, 33.04) for biochemical pregnancy, 4.85 (1.02, 23.08) for clinical pregnancy and 4.07 (0.83, 19.9) for the live birth rate compared with women in the lowest quartile (<4.55), all p-trend<0.001. Conclusions: Using multivariable models, we suggested that a baseline elevated serum folate level (≥33.0 ng/ml) and a lower Ca/Mg ratio were associated with worse ART outcomes in normogonadotropic women. Our findings might be useful for choosing safe dosages of folate, calcium, magnesium and complex supplementation for both fertile women and women undergoing infertility treatment. Further preconception large-scale studies with known micro- and macronutrient statuses of both parents and serum folate, Ca, Mg, and hormone levels, are needed.
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Calcio , Magnesio , Anciano , Femenino , Fertilización In Vitro , Ácido Fólico , Humanos , Masculino , Inducción de la Ovulación , Embarazo , Índice de Embarazo , Estudios ProspectivosRESUMEN
AIM: To prospectively investigate the associations of urinary phthalate metabolite concentrations measured at four time points spanning pubertal development with semen parameters in Russian men. DESIGN: 516 boys were enrolled at ages 8-9 years (2003-2005) and followed annually. METHODS: Urine samples were collected annually and pooled into four exposure windows [prepuberty, early puberty, late puberty and sexual maturity] based on physician assessed Tanner genitalia stages and testicular volume. Fifteen phthalate metabolites were quantified using isotope dilution HPLC-MS/MS at Moscow State University. We calculated molar sums (∑) of di-2-ethylhexyl phthalate (DEHP), di-isononyl phthalate (DiNP), di-isodecyl phthalate (DiDP) and anti-androgenic phthalate (AAP) metabolites. At sexual maturity (ages 18-19 years), the men provided 1-2 semen samples for analysis. We estimated the associations of quintiles of urinary ∑phthalate metabolites as well as mono-butyl phthalate (MnBP), mono-isobutyl phthalate (MiBP), and mono-benzyl phthalate (MBzP) at each pubertal window, with semen parameters by fitting generalized linear mixed models with random intercepts and adjusting for confounders. RESULTS: A total of 223 men who provided semen samples had phthalates measured at one or more pubertal windows. Higher urinary concentrations of ∑DiNP metabolites during late puberty were related to poorer semen quality (men with the highest quintile of urinary ∑DiNP had 30% lower sperm concentration, 32% lower count and 30% lower progressive motile count, compared to men in the lowest quintile). Also, young men with higher urinary concentrations of MiBP metabolites in early puberty tended to have poorer semen quality. No associations were observed for ∑DEHP metabolites, ∑DiDP metabolites, ∑AAP, MBzP or MnBP metabolites with semen quality parameters. CONCLUSIONS: ∑DiNP metabolites measured during late puberty and MiBP metabolites at early puberty were related to poorer semen quality, highlighting the importance of considering specific windows of exposure when investigating chemical exposures in relation to measures of reproductive health in men.
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Contaminantes Ambientales , Ácidos Ftálicos , Adolescente , Antagonistas de Andrógenos , Niño , Dibutil Ftalato , Exposición a Riesgos Ambientales/análisis , Contaminantes Ambientales/orina , Humanos , Masculino , Ácidos Ftálicos/orina , Pubertad , Análisis de Semen , Espectrometría de Masas en Tándem , Adulto JovenRESUMEN
BACKGROUND: Animal studies have demonstrated that timing of pubertal onset can be altered by prenatal exposure to dioxins or polychlorinated biphenyls (PCBs), but studies of human populations have been quite limited. METHODS: We assessed the association between maternal serum concentrations of dioxins and PCBs and the sons' age of pubertal onset in a prospective cohort of 489 mother-son pairs from Chapaevsk, Russia, a town contaminated with these chemicals during past industrial activity. The boys were recruited at ages 8 to 9 years, and 4 years of annual follow-up data were included in the analysis. Serum samples were collected at enrollment from both mothers and sons for measurement of dioxin and PCB concentrations using high-resolution mass spectrometry. The sons' pubertal onset--defined as pubertal stage 2 or higher for genitalia (G) or pubic hair (P), or testicular volume >3 mL--was assessed annually by the same physician. RESULTS: In multivariate Cox models, elevated maternal serum PCBs were associated with earlier pubertal onset defined by stage G2 or higher (4th quartile hazard ratio = 1.7 [95% confidence interval = 1.1- 2.5]), but not for stage P2 or higher or for testicular volume >3 mL. Maternal serum concentrations of dioxin toxic equivalents were not consistently associated with the sons' pubertal onset, although a dose-related delay in pubertal onset (only for G2 or higher) was seen among boys who breast-fed for 6 months or more. CONCLUSIONS: Maternal PCB serum concentrations measured 8 or 9 years after sons' births--which may reflect sons' prenatal and early-life exposures--were associated with acceleration in some, but not all, measures of pubertal onset.
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Dioxinas/efectos adversos , Bifenilos Policlorados/efectos adversos , Efectos Tardíos de la Exposición Prenatal/sangre , Pubertad/efectos de los fármacos , Adulto , Factores de Edad , Niño , Dioxinas/sangre , Femenino , Edad Gestacional , Humanos , Plomo/efectos adversos , Plomo/sangre , Masculino , Análisis Multivariante , Bifenilos Policlorados/sangre , Embarazo , Modelos de Riesgos Proporcionales , Estudios Prospectivos , Federación de Rusia/epidemiología , Adulto JovenRESUMEN
BACKGROUND: The present study assessed the temporal trend in serum concentrations of polychlorinated dibenzo-p-dioxins, dibenzofurans, and biphenyls (PCBs) among residents of a Russian town where levels of these chemicals are elevated due to prior industrial activity. METHODS: Two serum samples were collected from eight adult women (in 2000 and 2009), and analyzed with gas chromatography-high-resolution mass spectrometry. RESULTS: The average total toxic equivalency (TEQ) decreased by 30% (from 36 to 25 pg/g lipid), and the average sum of PCB congeners decreased by 19% (from 291 to 211 ng/g lipid). Total TEQs decreased for seven of the eight women, and the sum of PCBs decreased for six of eight women. During this nine year period, larger decreases in serum TEQs and PCBs were found in women with greater increases in body mass index. CONCLUSIONS: This study provides suggestive evidence that average serum concentrations of dioxins, furans, and PCBs are decreasing over time among residents of this town.
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Benzofuranos/sangre , Dioxinas/sangre , Exposición a Riesgos Ambientales , Contaminantes Ambientales/sangre , Bifenilos Policlorados/sangre , Adulto , Femenino , Cromatografía de Gases y Espectrometría de Masas , Humanos , Estudios Longitudinales , Federación de Rusia , Adulto JovenRESUMEN
BACKGROUND: Dioxins, furans and polychlorinated biphenyls (PCBs) are endocrine disruptors with half-lives of months to years in humans. Peripubertal exposure to these chemicals may disrupt pubertal timing. Biomarker exposure metrics like the sum of non-dioxin-like PCBs in serum (∑NDL-PCBs) involve chemicals with different hypothetical effects. Empirical exposure metrics represent an alternative that requires no knowledge of biological mechanisms. METHODS: From 2003 to 2005, 516 Russian boys aged 8-9 residing near a plant that historically produced organochlorine pesticides were enrolled in the Russian Children's Study. At enrollment, blood was collected for measurement of organochlorine chemicals by the Centers for Disease Control and Prevention (CDC). Variable cluster analysis was used to empirically group serum levels of 45 dioxins, furans and PCBs into eight cluster scores. These scores were compared to the sum of dioxin-like toxic equivalents (∑TEQs) and ∑NDL-PCBs using Spearman correlation coefficients. RESULTS: Clustering appeared to reflect number and position of chlorine atoms. Clusters 4, 7 and 1 contained, respectively, tri/tetra-, tetra/penta-, and hexa/hepta-chlorinated PCBs with chlorines at the 4,4' positions. Clusters 2, 8 and 5 contained, respectively, tetra-to hexa-, hexa/hepta-, and hepta/octa-chlorinated PCBs with chlorines at the 2,2' positions. ∑NDL-PCBs were highly correlated with 4,4'-chlorinated clusters 1 and 7 (Spearman's ρ = 0.8) and less so with all other clusters (Spearman's ρ = 0.4-0.5). CONCLUSIONS: In this cohort of Russian boys, baseline serum dioxins, furans and PCBs seemed to cluster by chlorination pattern, possibly reflecting shared persistence, metabolism or source of exposure. The widely used measure ∑NDL-PCB seemed most representative of 4,4'-chlorinated PCBs.
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Dioxinas , Bifenilos Policlorados , Niño , Exposición a Riesgos Ambientales/análisis , Furanos , Humanos , Masculino , Federación de RusiaRESUMEN
Aims: Paternal age is increasing in developed countries. Understanding of aging-related epigenetic changes in sperm is needed as well as factors that modify such changes. Materials & methods: Young pubertal and mature rats were exposed perinatally to vehicle or environmental xenobiotic 2,2',4,4'-tetrabromodiphenyl ether. Epididymal sperm was reduced representation bisulfite sequenced. Differentially methylated regions (DMRs) were identified via MethPipe. Results: In control animals, 5319 age-dependent DMRs were identified. Age-related DMRs were enriched for embryonic development. In exposed rats, DNA methylation was higher in young and lower in mature animals then in controls. Conclusions: Sperm methylome undergoes significant age-dependent changes, which may represent a causal link between paternal age and offspring phenotype. Environmental xenobiotics can interfere with the natural process of epigenetic aging.
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Envejecimiento/fisiología , Metilación de ADN/efectos de los fármacos , Retardadores de Llama/efectos adversos , Espermatozoides/efectos de los fármacos , Animales , Desarrollo Embrionario/efectos de los fármacos , Epigénesis Genética/efectos de los fármacos , Epigenoma/efectos de los fármacos , Epigenómica/métodos , Femenino , Masculino , Parto/efectos de los fármacos , Edad Paterna , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Ratas , Ratas WistarRESUMEN
Recent studies demonstrate that sperm epigenome is a vehicle that conveys paternal experiences to offspring phenotype. That evidence triggers interest of both experimental and epidemiological studies of epigenetic markers in sperm. Since samples are often unique in epidemiological studies, a careful and efficient use of the material is a critical requirement. The goal of this study was to provide optimization of methods for the isolation of small RNAs from spermatozoa and library preparation for sequencing. A total 67 fractionated sperm samples from the Russian Children's Study biobank prospectively collected at 18-20 years of age were used to isolate small RNAs with median (IQR) input total sperm count 17.0 (7.4-35.9) million. Twenty-four pairs of libraries were prepared using the NEBNext and NEXTFlex kits, 19 libraries using NEBNext and 6 using NEXTFlex. All libraries were sequenced on NextSeq 500, and the results were evaluated as a function of the number of small non-coding RNA (sncRNA) detected, quality parameters of sequencing libraries, as well as technical features of sample preparation. Although the same amount of miRNA input was used for NEBNext and NEXTFlex libraries, the concentration of DNA in NEBNext libraries was significantly higher in comparison with NEXTFlex libraries. In high input (sperm count >28 million and more than 25 ng miRNA in library) NEXTFlex Small RNA-Seq kit detected more microRNAs. In low input, the NEBNext proved more effective. The tricks and traps to protocol optimization are presented, including an efficient and effector gel-based system for the removal of sequencing library adaptors.
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Secuenciación de Nucleótidos de Alto Rendimiento , MicroARNs , Biblioteca de Genes , Humanos , Masculino , Análisis de Secuencia de ARN , EspermatozoidesRESUMEN
Perinatal exposures to polybrominated diphenyl ethers permanently reprogram liver metabolism and induce a nonalcoholic fatty liver disease-like phenotype and insulin resistance in rodents. Aim: To test if these changes are associated with altered liver epigenome. Materials & methods: Expression of small RNA and changes in DNA methylation in livers of adult rats were analyzed following perinatal exposure to 2,2',4,4'-tetrabromodiphenyl ether, the polybrominated diphenyl ether congener most prevalent in human tissues. Results: We identified 33 differentially methylated DNA regions and 15 differentially expressed miRNAs. These changes were enriched for terms related to lipid and carbohydrate metabolism, insulin signaling, Type-2 diabetes and nonalcoholic fatty liver disease. Conclusion: Changes in the liver epigenome are a likely candidate mechanism of long-term maintenance of an aberrant metabolic phenotype.