Paternal aging impacts expression and epigenetic markers as early as the first embryonic tissue lineage differentiation.

BACKGROUND: Advanced paternal age (APA) is associated with adverse outcomes to offspring health, including increased risk for neurodevelopmental disorders. The aim of this study was to investigate the methylome and transcriptome of the first two early embryonic tissue lineages, the inner cell mass (ICM) and the trophectoderm (TE), from human blastocysts in association with paternal age and disease risk. High quality human blastocysts were donated with patient consent from donor oocyte IVF cycles from either APA (≥ 50 years) or young fathers. Blastocysts were mechanically separated into ICM and TE lineage samples for both methylome and transcriptome analyses. RESULTS: Significant differential methylation and transcription was observed concurrently in ICM and TE lineages of APA-derived blastocysts compared to those from young fathers. The methylome revealed significant enrichment for neuronal signaling pathways, as well as an association with neurodevelopmental disorders and imprinted genes, largely overlapping within both the ICM and TE lineages. Significant enrichment of neurodevelopmental signaling pathways was also observed for differentially expressed genes, but only in the ICM. In stark contrast, no significant signaling pathways or gene ontology terms were identified in the trophectoderm. Despite normal semen parameters in aged fathers, these significant molecular alterations can adversely contribute to downstream impacts on offspring health, in particular neurodevelopmental disorders like autism spectrum disorder and schizophrenia. CONCLUSIONS: An increased risk for neurodevelopmental disorders is well described in children conceived by aged fathers. Using blastocysts derived from donor oocyte IVF cycles to strategically control for maternal age, our data reveals evidence of methylation dysregulation in both tissue lineages, as well as transcription dysregulation in neurodevelopmental signaling pathways associated with APA fathers. This data also reveals that embryos derived from APA fathers do not appear to be compromised for initial implantation potential with no significant pathway signaling disruption in trophectoderm transcription. Collectively, our work provides insights into the complex molecular mechanisms that occur upon paternal aging during the first lineage differentiation in the preimplantation embryo. Early expression and epigenetic markers of APA-derived preimplantation embryos highlight the susceptibility of the future fetus to adverse health outcomes.

Effects of resistance exercise alone or with caffeine on hemodynamics, autonomic modulation and arterial stiffness in resistance-trained women.

AIM: Both an acute bout of resistance exercise (RE) and caffeine consumption can significantly alter hemodynamics, autonomic modulation, and arterial stiffness, which may correlate with adverse cardiovascular events. However, effects of an acute bout of RE and caffeine are unclear in resistance-trained women. PURPOSE: The purpose of this study was to compare the effects of an acute bout of RE with repetitions to failure on squat and bench press, with or without caffeine, on performance, resting and recovery measures of hemodynamics, autonomic modulation, as well as arterial stiffness in resistance-trained women. METHODS: Eleven women participated in a double-blind, placebo controlled cross-over design in which they consumed caffeine (4 mg/kg) or placebo at least 72 h apart. Sixty minutes following ingestion, participants performed two sets of 10 repetitions followed by a third set to failure on squat and bench press. Hemodynamics, autonomic modulation, and arterial stiffness were measured at rest, 60 min post-ingestion, and three minutes and 10 min following RE. RESULTS: Data demonstrated caffeine has no additive effects on performance, hemodynamics, autonomic modulation, or arterial stiffness (p > 0.05) before or following an acute bout of RE in resistance-trained women compared to a placebo. CONCLUSIONS: Resistance-trained women may not observe any alteration to RE performance on the squat and bench press in terms of repetitions to failure following caffeine ingestion. In addition, the data from the present study suggests that there may also not be any further negative effects on the cardiovasculature if caffeine is consumed prior to the RE bout.

Effects of a Cool-Down after Supramaximal Interval Exercise on Autonomic Modulation.

Supramaximal interval exercise alters measures of autonomic modulation, while a cool-down may speed the recovery of vagal modulation. The purpose of this study was to compare the effects of a cool-down (pedaling a cycle ergometer at 50 rpm against a resistance of 45 W) versus passive recovery (no pedaling) after supramaximal interval exercise on autonomic modulation. Sixteen moderately active individuals (Mean ± SD: 23 ± 3 years (men: n = 10; women: n = 6) were assessed for autonomic modulation at Rest, and 15 (R15), 30 (R30), 45 (R45) and 60 (R60) min following supramaximal interval exercise. Linear measures of autonomic modulation included natural log (ln) total power (lnTP), high-frequency power (lnHF), the ratio of low frequency (LF) to HF ln(LF/HF) ratio, root mean square of successive differences between normal heartbeats (lnRMSSD), while non-linear measures included sample entropy (SampEn) and Lempel-Ziv entropy (LZEn). Two-way repeated ANOVAs were used to evaluate the main effects of condition (cool-down, passive recovery) across time (Rest, and R15, R30, R45 and R60). There were significant (p ≤ 0.05) condition by time interactions for SampEn and LZEn, such that they decreased at 15, 30, 45 and 60 min during passive recovery compared to Rest, with the recovery of SampEn and LZEn by 60 and 45 min, respectively, during cool-down. There were significant (p ≤ 0.05) main effects of time for lnTP, lnHF and lnRMSSD, such that lnTP, lnHF and lnRMSSD were attenuated, and lnLF/HF ratio was augmented, at all recovery times compared to Rest. These data demonstrate that a cool-down increases the recovery of nonlinear measures of vagal modulation within 45-60 min after supramaximal interval exercise, compared to passive recovery in moderately active individuals.

Cardiac Autonomic Function Following Bilateral and Unilateral Upper Body Acute Resistance Exercise.

The purpose of this study was to compare cardiac autonomic responses following bilateral and unilateral upper-body (UB) acute resistance exercise (ARE). In total, 14 individuals were assessed for markers of cardiac autonomic responses via heart rate variability (HRV) and baroreflex sensitivity (BRS) at rest and at 10- and 30-min following ARE. Logarithmically transformed (ln) HRV measures included: total power (ln TP), high-frequency power (ln HF power), low-frequency power (ln LF power), sympathovagal balance (ln LF: HF), and the square root of the mean squared differences of successive R-R intervals (ln RMSSD). BRS was assessed using the sequence method. Two-way repeated measures ANOVAs were used to analyze effects of UB ARE (bilateral, unilateral) across time (Rest, 10, and 30 min). There were no significant (p > 0.05) interactions. However, there were significant (p ≤ 0.05) main effects of time such that ln TP, ln HF power, ln RMSSD, and BRS decreased and did not recover within 30 min compared to Rest for both conditions. Collectively, this study suggests that bilateral and unilateral UB ARE yielded similar reductions, for at least 30 min, in respect to vagal measures of HRV and BRS.

Hemodynamic response and pulse wave analysis after upper- and lower-body resistance exercise with and without blood flow restriction.

Resistance exercise (RE) has been shown to elevate hemodynamics and pulse wave reflection. However, the effects of acute RE with blood flow restriction (BFR) on hemodynamics and pulse wave reflection are unclear. The purpose of this study was to evaluate the differences between upper- and lower-body RE with and without BFR on hemodynamics and pulse wave reflection. Twenty-three young resistance-trained individuals volunteered for the study. Hemodynamics and pulse wave reflection were assessed at rest, 10, 25, 40, and 55 min after either upper- or lower-body with or without BFR. The upper-body RE (URE) consisted of the latissimus dorsi pulldown and chest press; the lower-body RE (LRE) consisted of knee extension and knee flexion. The BFR condition consisted of four sets of 30, 15, 15, and 15 repetitions at 30% 1-repetition maximum (1RM) while the without BFR condition consisted of four sets of 8 repetitions at 70% 1RM. Heart rate, rate pressure product, and subendocardial viability ratio significantly (p < 0.05) increased after all exercises. Brachial and aortic systolic blood pressure (BP) significantly (p < 0.05) elevated after LRE while brachial and aortic diastolic BP significantly (p < 0.05) reduced after URE. Augmentation pressure, augmentation index (AIx), AIx normalized at 75 bpm, and wasted left ventricular pressure energy significantly (p < 0.05) increased after URE while transit time of reflected wave significantly (p < 0.05) decreased after LRE. URE places greater stress on pulse wave reflection while LRE results in greater responses in BP. Regardless of URE or LRE, the cardiovascular responses between BFR and without BFR are similar.HIGHLIGHTS High-load resistance exercise and low-load resistance exercise with blood flow restriction may produce similar cardiovascular responses.Upper-body resistance exercise generates greater changes on pulse wave reflections while lower-body resistance exercise induces greater elevations in systolic blood pressure.

Increased Systemic Antioxidant Power Ameliorates the Aging-Related Reduction in Oocyte Competence in Mice.

Ovarian aging is associated with elevated oxidative stress and diminished oocyte developmental competence. We aimed to determine the impact of systemic antioxidant treatment in aged mice. Female outbred CF-1 mice were aged for 9 months prior to an 8-week 45 mg Euterpe oleracea (açaí) daily supplement. The açaí treatment induced a threefold increase in serum antioxidant power (FRAP) compared to both young and aged mice (p < 0.0001). Compared to young mice, aged mice had fewer oocytes and reduced blastocyst development (p < 0.0001); açaí did not affect the oocyte numbers, but improved blastocyst formation (p < 0.05). Additionally, açaí alleviated the aging-related decrease in implantation potential (p < 0.01). The aged mice showed evidence of elevated ovarian ER stress (increased whole-ovary PDIA4 expression, granulosa cell and oocyte GRP78 expression, and oocyte PDIA4 protein), reduced oocyte mitochondrial quality (higher PRKN activation and mitochondrial DNA oxidative damage), and dysregulated uterine glandular epithelium. Antioxidant intervention was sufficient to lessen these effects of ovarian aging, likely in part by the upregulation of NRF2. We conclude that açaí treatment is a promising strategy to improve ER and mitochondrial function in the ovaries, thereby ameliorating the decreased oocyte competence that occurs with ovarian aging.

Vascular Responses to High-Intensity Battling Rope Exercise between the Sexes.

The purpose of the study was to assess high-intensity battling rope exercise (HI-BRE) on hemodynamics, pulse wave reflection and arterial stiffness during recovery and between sexes. Twenty-three young, healthy resistance-trained individuals (men: n = 13; women: n = 10) were assessed for all measures at Rest, as well as 10-, 30-, and 60-minutes following HI-BRE. A one-way repeated measures ANOVA was used to analyze the effects of HI-BRE across time (Rest, 10, 30, and 60-minutes) on all dependent variables. Significant main effects were analyzed using paired t-tests with a Sidak correction factor. Significance was accepted a priori at p 0.05. There were significant reductions in hemodynamic measures of diastolic blood pressure (BP) in women, but not men following HI-BRE at 30 minutes. Further, measures of pulse wave reflection, specifically those of the augmentation index (AIx) and wasted left ventricular energy (ΔEw), were significantly increased in both men and women for 60 minutes, but changes were significantly attenuated in women suggesting less ventricular work. There were also significant increases in arterial stiffness in regard to the aorta and common carotid artery that were fully recovered by 30 and 60 minutes, respectively with no differences between men and women. Thus, the primary findings of this study suggest that measures of hemodynamics and pulse wave reflection are collectively altered for at least 60 minutes following HI-BRE, with women having attenuated responses compared to men.

Forecasting early onset diminished ovarian reserve for young reproductive age women.

PURPOSE: To investigate the biological networks associated with DOR in young women and the subsequent molecular impact on preimplantation embryos. METHODS: Whole peripheral blood was collected from patients: young women presenting with diminished ovarian reserve (DOR) and age-matched young women with normal ovarian reserve. Maternal exome sequencing was performed on the NovaSEQ 6000 and sequencing validation was completed using Taqman® SNP Genotyping Assays. Blastocyst global methylome and transcriptome sequencing were also analyzed. RESULTS: Exome sequencing revealed 730 significant DNA variants observed exclusively in the young DOR patients. Bioinformatic analysis revealed a significant impact to the Glucocorticoid receptor (GR) signaling pathway and each young DOR female had an average of 6.2 deleterious DNA variants within this pathway. Additional stratification based on patient age resulted in a cut-off at 31 years for young DOR discrimination. Embryonic global methylome sequencing resulted in only a very small number of total CpG sites with methylation alterations (1,775; 0.015% of total) in the DOR group. Additionally, there was no co-localization between these limited number of altered CpG sites and significant variants, genes, or pathways. RNA sequencing also resulted in no biologically significant transcription changes between DOR blastocysts and controls. CONCLUSION: GR signaling DNA variants were observed in women with early-onset DOR potentially compromising oocyte production and quality. However, no significant downstream effects on biological processes appear to impact the resulting blastocyst. The ability to forecast premature DOR for young women may allow for earlier identification and clinical intervention for this patient population.

Antioxidant Intervention Attenuates Aging-Related Changes in the Murine Ovary and Oocyte.

Advanced maternal age (AMA) is associated with reduced fertility due in part to diminished ovarian follicle quantity, inferior oocyte quality, chromosome aneuploidy, and lower implantation rates. Ovarian aging is accompanied by increased oxidative stress and blunted antioxidant signaling, such that antioxidant intervention could improve reproductive potential. The first aim of this study was to determine the molecular effects of antioxidant intervention in the ovaries and oocytes of aged mice, utilizing a supplement containing only naturally occurring açaí (Euterpe oleracea) with an oxygen radical absorbance capacity of 208,628 µmol Trolox equivalent (TE)/100 g indicating high antioxidant activity. Nine month old female CF-1 mice were administered 80 mg/day antioxidants (n = 12) or standard diet (n = 12) for 12 weeks. In the ovary, antioxidant treatment upregulated ß-adrenergic signaling, downregulated apoptosis and proinflammatory signaling, and variably affected cell growth and antioxidant pathways (p < 0.05). Exogenous antioxidants also increased the oocyte expression of antioxidant genes GPX1, SOD2, and GSR (p < 0.05). A feasibility analysis was then conducted on female AMA infertility patients as a proof-of-principle investigation. Patients (n = 121; <45 years old) consented to receiving 600 mg antioxidants three times daily for ≥8 weeks preceding infertility treatment. Preliminary results indicate promising outcomes for AMA patients, warranting further investigation.

The inherited methylome landscape is directly altered with paternal aging and associated with offspring neurodevelopmental disorders.

Paternal aging and the prevalence of neurodevelopmental disorders in offspring are well documented. Yet, the underlying mechanism and the mode of inheritance have not been conclusively established. Advancing paternal age is a subtle and varying phenotype. As such, it is likely that a threshold for cumulative risk may exist that, if surpassed, culminates in a predisposition to disease and ultimately an observed phenotype in offspring. Epigenetic regulation provides a plausible explanation for the nongenetic paternal transmission of disease susceptibility. With the use of whole-genome methylation sequencing, the data described herein substantiate an increasingly compromised DNA methylation profile as sperm ages and, for the first time, also demonstrate a generational correlation in sperm and blastocyst of an altered methylome associated with advanced paternal age. Methylation alterations are not randomly distributed across the genome, but appear clustered at certain chromosomal locations, and significantly colocalize with regions of nucleosome retention. Genes associated with autism spectrum disorder, schizophrenia, and bipolar disorder are significantly enriched with causative methylation aberrations in both sperm and embryos from aged fathers. The long-term health burden and societal economic impact of these conditions are substantial and will continue with increasingly prevalent diagnosis. This work provides a mechanistic link between the paternal age effect and offspring neurodevelopmental disorders leading to a better understanding of causation and investigation into potential future therapy.

The Effects of Machine-Weight and Free-Weight Resistance Exercise on Hemodynamics and Vascular Function.

The purpose of this study was to examine hemodynamic and vascular responses between machine-weight and free-weight exercise. Resistance-trained individuals were assigned to a machine-weight (n = 13) or free-weight (n = 15) group. Groups completed two visits consisting of their assigned exercise condition and a control (CON). A 2 × 2 × 3 repeated measures ANOVA was used to test the effects of group across condition and time on the hemodynamic parameters [cardiac output (CO), heart rate (HR), total peripheral resistance (TPR), mean arterial pressure (MAP), and stroke volume (SV)]. A 2 × 2 × 2 repeated measures ANOVA was used to test the effects of group across condition and time on the hemodynamic variable, forearm vascular conductance (FVC), as well as on vascular measures [forearm blood flow (FBF), blood flow peak, and total reactive hyperemia (RH)]. Main effects were analyzed using pairwise comparisons. The results of the present study demonstrate that both machine-weight and free-weight exercise produce similar (p > 0.05) alterations in hemodynamics and vascular function. Specifically, during recovery both groups demonstrated significant (p ≤ 0.05) increases in measures of hemodynamics such as CO, HR and FVC, as well as significant (p ≤ 0.05) decreases in TPR, MAP, and SV. Measures of vascular function such as FBF, blood flow peak, and total RH were also significantly (p ≤ 0.05) increased during recovery. Therefore, this study suggests that either machine weight or free-weight exercise may induce acute hemodynamic and vascular benefits, which may reduce the risk of cardiovascular disease (CVD) and CVD events.

Advanced paternal age directly impacts mouse embryonic placental imprinting.

The placental epigenome plays a critical role in regulating mammalian growth and development. Alterations to placental methylation, often observed at imprinted genes, can lead to adverse pregnancy complications such as intrauterine growth restriction and preterm birth. Similar associations have been observed in offspring derived from advanced paternal age fathers. As parental age at time of conception continues to rise, the impact of advanced paternal age on these reproductive outcomes is a growing concern, but limited information is available on the molecular mechanisms affected in utero. This longitudinal murine research study thus investigated the impact of paternal aging on genomic imprinting in viable F1 embryonic portions of the placentas derived from the same paternal males when they were young (4-6 months) and when they aged (11-15 months). The use of a controlled outbred mouse model enabled analysis of offspring throughout the natural lifetime of the same paternal males and excluded confounding factors like female age or infertility. Firstly, paternal age significantly impacted embryonic placental weight, fetal weight and length. Targeted bisulfite sequencing was utilized to examine imprinted methylation at the Kcnq1ot1 imprinting control region, with significant hypermethylation observed upon natural paternal aging. Quantitative real-time PCR assessed imprinted gene expression levels at various imprinting clusters, resulting in transcript level alterations attributable to advanced paternal age. In summary, our results demonstrate a paternal age effect with dysregulation at numerous imprinted loci, providing a mechanism for future adverse placental and offspring health conditions.

Free-weight versus weight machine resistance exercise on pulse wave reflection and aortic stiffness in resistance-trained individuals.

The purpose of this study was to compare the vascular responses to acute free-weight (FW) resistance exercise (RE) versus weight machines (WM). Thirty-two resistance-trained individuals participated in this study. Both modalities involved performing acute RE and a control. Blood pressure and measures of pulse wave reflection were assessed using pulse wave analysis. Aortic stiffness was assessed using carotid-femoral pulse wave velocity (cf-PWV). A repeated measures analysis of variance was used to determine the effects of modality (FW and WM) and condition (acute RE and control) across time (rest and 10-20 min after exercise) on measures of pulse wave reflection and aortic stiffness. Significance was set a priori at p ≤ 0.05. There were no modality by condition by time interactions for any variable, such that the FW and WM modalities responded similarly across time after acute RE (p > 0.05). There were significant (p ≤ 0.05) increases in heart rate, aortic systolic blood pressure, aortic pulse pressure, augmentation index normalized at 75bpm, and decreases in subendocardial viability ratio (SEVR) after acute RE, compared to rest. There was also a significant (p ≤ 0.05) increase in cf-PWV after acute RE, compared to rest. In conclusion, this study demonstrates that acute free-weight and weight-machine RE are associated with transient increases in measures of pulse wave reflection and aortic stiffness, with reductions in myocardial perfusion. These data demonstrate that both modalities result in significant stress on the myocardium during recovery, while simultaneously increasing pressure on the aorta for at least 10-20 min.

Autonomic modulation following an acute bout of bench press with and without blood flow restriction.

PURPOSE: Traditional resistance exercise decreases vagal tone up to 30 min after an acute bout of resistance exercise, which may increase the risk of cardiovascular events. However, the effects of resistance exercise with blood flow restriction (BFR) on autonomic modulation are unclear. To evaluate autonomic modulation after resistance exercise with and without BFR in resistance-trained men. METHODS: Eleven young men volunteered for the study. Autonomic modulation was assessed at rest, 15 (Rec 1), and 25 (Rec 2) minutes after low-load bench press with BFR (LL-BFR), traditional high-load bench press (HL), and a control (CON). Autonomic modulation assessments were expressed as natural logarithm (Ln), and included total power (LnTP), low-frequency power (LnLF), high-frequency power (LnHF), sympathovagal balance (LnLF/LnHF ratio), root mean square of the successive differences (LnRMSSD), and the proportion of intervals differing by > 50 ms from the preceding intervals (LnPNN50). A repeated measures ANOVA was used to evaluate conditions (LL-BFR, HL and CON) across time (Rest, Rec1, and Rec2) on autonomic modulation. RESULTS: There were significant condition by time interactions for LnTP, LnHF, and LnRMSSD such that they were reduced during recovery after LL-BFR and HL compared to Rest and CON. There were no interactions in the LnLF, LnLF/LnHF ratio, and LnPNN50. CONCLUSIONS: These data suggest that LL-BFR and HL significantly alter autonomic modulation up to 30 min after exercise with significant reduction after HL compared to LL-BFR when exercise volume is equated.

Compromised global embryonic transcriptome associated with advanced maternal age.

PURPOSE: To investigate the global transcriptome and associated embryonic molecular networks impacted with advanced maternal age (AMA). METHODS: Blastocysts derived from donor oocyte IVF cycles with no male factor infertility (< 30 years of age) and AMA blastocysts (≥ 42 years) with no other significant female factor infertility or male factor infertility were collected with informed patient consent. RNA sequencing libraries were prepared using the SMARTer® Ultra® Low Kit (Clontech Laboratories) and sequenced on the Illumina HiSEQ 4000. Bioinformatics included Ingenuity® Pathway Analysis (Qiagen) with ViiA™ 7 qPCR utilized for gene expression validation (Applied Biosystems). RESULTS: A total of 2688 significant differentially expressed transcripts were identified to distinguish the AMA blastocysts from young, donor controls. 2551 (95%) of these displayed decreased transcription in the blastocysts from older women. Pathway analysis revealed three altered molecular signaling networks known to be critical for embryo and fetal development: CREBBP, ESR1, and SP1. Validation of genes within these networks confirmed the global decreased transcription observed in AMA blastocysts (P < 0.05). CONCLUSIONS: A significant, overall decreased global transcriptome was observed in blastocysts from AMA women. The ESR1/SP1/CREBBP pathway, in particular, was found to be a highly significant upstream regulator impacting biological processes that are vital during embryonic patterning and pre-implantation development. These results provide evidence that AMA embryos are compromised on a cell signaling level which can repress the embryo's ability to proliferate and implant, contributing to a deterioration of reproductive outcomes.

Autonomic modulation and baroreflex sensitivity after acute resistance exercise: responses between sexes.

BACKGROUND: The aim of this study was to evaluate autonomic modulation, blood pressure variability and baroreflex sensitivity (BRS) responses to an acute bout of free-weight resistance exercise in resistance-trained men (N.=14) and women (N.=13). METHODS: Participants underwent both an acute bout of resistance exercise (RE) consisting of 3 sets of 10 repetitions at 75% 1-repetition maximum on the squat, bench press, and deadlift, and a quiet control. Autonomic modulation, blood pressure variability (LFSAP), and cardiovagal BRS were assessed at rest, 15-20 minutes (Rec1) and 25-30 minutes (Rec2) postexercise. Log transformed measures of autonomic modulation included root square of the mean ssquared differences of successive RR intervals (LnRMSSD), high-frequency power (LnHFRR) and low-frequency power (LnLFRR) and sympathovagal balance (LnLFRR/HFRR). LFSAP was used as a measurement of vasomotor tone. Cardiovagal BRS was assessed using the sequence method. RESULTS: There were no significant sex differences at rest and no significant sex by time by condition interactions for any variable. Compared with rest there were augmentations in LnLFRR/HFRR (P=0.002) and LFSAP (P=0.001) at Rec1 and Rec2. RMSSD and cardiovagal BRS were significantly (P=0.0001) decreased at Rec1 and Rec2 compared to rest after the acute RE. CONCLUSIONS: Both sexes demonstrated that acute resistance exercise using free weights has a profound impact on autonomic modulation, blood pressure variability and cardiovagal BRS.

Pulse wave reflection responses to bench press with and without practical blood flow restriction.

Resistance exercise is recommended to increase muscular strength but may also increase pulse wave reflection. The effect of resistance exercise combined with practical blood flow restriction (pBFR) on pulse wave reflection is unknown. The purpose of this study was to evaluate the differences in pulse wave reflection characteristics between bench press with pBFR and traditional high-load bench press in resistance-trained men. Sixteen resistance-trained men participated in the study. Pulse wave reflection characteristics were assessed before and after low-load bench press with pBFR (LL-pBFR), traditional high-load bench press (HL), and a control (CON). A repeated-measures ANOVA was used to evaluate differences in pulse wave reflection characteristics among the conditions across time. There were significant (p ≤ 0.05) interactions for heart rate, augmentation index, augmentation index normalized at 75 bpm, augmentation pressure, time-tension index, and wasted left ventricular energy such that they were increased after LL-pBFR and HL compared with rest and CON, with no differences between LL-pBFR and HL. Aortic pulse pressure (p < 0.001) was elevated only after LL-pBFR compared with rest. In addition, there was a significant (p ≤ 0.05) interaction for aortic diastolic blood pressure (BP) such that it was decreased after LL-pBFR compared with rest and CON but not HL. The subendocardial viability ratio and diastolic pressure-time index were significantly different between LL-pBFR and HL compared with rest and CON. There were no significant interactions for brachial systolic or diastolic BP, aortic systolic BP, or time of the reflected wave. In conclusion, acute bench press resistance exercise significantly altered pulse wave reflection characteristics without differences between LL-pBFR and HL.

Inheritance of epigenetic dysregulation from male factor infertility has a direct impact on reproductive potential.

OBJECTIVE: To evaluate the epigenetic consequence on the methylome and subsequent transcriptome in euploid blastocysts of male-factor (MF) infertility patients. DESIGN: Methylome and transcriptome analysis on individual oligoasthenoteratozoospermia (OAT [MF]) blastocysts. SETTING: Infertility clinic. PATIENT(S): Clinical data from 128 couples presenting with OAT (MF) and 118 maternal age-matched control (no MF) subjects undergoing infertility treatment from 2010 to 2014, along with 72 surplus cryopreserved blastocysts donated from 33 couples with their informed consents. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Methyl Maxi-Seq (Zymo Research) was used to determine genome-wide DNA methylation, and small cell number RNA-Seq was used to examine the global transcriptome. Validation experiments were performed with the use of pyrosequencing or quantitative real-time polymerase chain reaction. Statistical analysis used Student t test, analysis of variance in R, Fisher exact test, and pairwise fixed reallocation randomization test where appropriate, with significance at P<.05. RESULT(S): Clinical pregnancy rates were similar between OAT (MF) patients and control (no MF) subjects after euploid embryo transfer. However, the miscarriage rate for OAT (MF) patients was significantly higher (14.7% vs. 2.2%; P<.05). Methylome and transcriptome analyses of individual blastocysts revealed significant alterations in 1,111 CpG sites and 469 transcripts, respectively (P<.05). Pathway analysis elucidated genes involved in "regulation of cellular metabolic process" as universally affected. Validation of the genome-wide approaches was performed for SBF1 and SLC6A9 (P<.05). CONCLUSION(S): Methylation and transcription aberrations in individual OAT (MF) blastocysts illustrate an epigenetic consequence of MF infertility on embryogenesis, significantly altering key developmental genes and affecting embryonic competence. This epigenetic dysregulation provides an explanation for the reduced reproductive potential in OAT (MF) patients despite euploid blastocyst transfers.

The impact of infertility diagnosis on embryo-endometrial dialogue.

Initial stages of implantation involve bi-directional molecular crosstalk between the blastocyst and endometrium. This study investigated an association between infertility etiologies, specifically advanced maternal age (AMA) and endometriosis, on the embryo-endometrial molecular dialogue prior to implantation. Co-culture experiments were performed with endometrial epithelial cells (EEC) and cryopreserved day 5 blastocysts (n = 41 ≥ Grade 3BB) donated from patients presenting with AMA or endometriosis, compared to fertile donor oocyte controls. Extracellular vesicles isolated from co-culture supernatant were analyzed for miRNA expression and revealed significant alterations correlating to AMA or endometriosis. Specifically, AMA resulted in 16 miRNAs with increased expression (P ≤ 0.05) and strong evidence for negative regulation toward 206 target genes. VEGFA, a known activator of cell adhesion, displayed decreased expression (P ≤ 0.05), validating negative regulation by 4 of these increased miRNAs: miR-126; 150; 29a; 29b (P ≤ 0.05). In endometriosis patients, a total of 10 significantly altered miRNAs displayed increased expression compared to controls (miR-7b; 9; 24; 34b; 106a; 191; 200b; 200c; 342-3p; 484) (P ≤ 0.05), targeting 1014 strong evidence-based genes. Three target genes of miR-106a (CDKN1A, E2F1 and RUNX1) were independently validated. Functional annotation analysis of miRNA-target genes revealed enriched pathways for both infertility etiologies, including disrupted cell cycle regulation and proliferation (P ≤ 0.05). These extracellular vesicle-bound secreted miRNAs are key transcriptional regulators in embryo-endometrial dialogue and may be prospective biomarkers of implantation success. One of the limitations of this study is that it was a stimulated, in vitro model and therefore may not accurately reflect the in-vivo environment.

Alterations in the sperm histone-retained epigenome are associated with unexplained male factor infertility and poor blastocyst development in donor oocyte IVF cycles.

STUDY QUESTION: Is there a distinct sperm histone-retained epigenetic signature in unexplained male factor infertility patients resulting in compromised blastocyst development? SUMMARY ANSWER: Using only donor oocyte IVF cycles, sperm DNA methylation patterns and miRNA profiles were significantly altered in normozoospermic patients resulting in poor blastocyst development, reflecting a subset of unexplained male factor infertility. WHAT IS KNOWN ALREADY: Aberrant sperm DNA methylation has been associated with known male factor infertility, particularly noted in oligozoospermic patients. Unexplained male factor infertility remains a significant proportion of in vitro fertilization failures having unknown underlying physiology. STUDY DESIGN, SIZE, DURATION: Sperm samples (n = 40) and blastocysts (n = 48) were obtained during fertile donor oocyte IVF cycles with normozoospermic parameters, thereby excluding known female and male infertility factors. Samples were divided into two groups based on blastocyst development (Good Group = ≥20% embryos with D5 grade 'AA' blastocysts, and ≥60% embryos of transferable quality on D5 and D6; Poor Group = ≤10% embryos with D5 grade 'AA' blastocysts, and ≤40% embryos of transferable quality on D5 and D6). PARTICIPANTS/MATERIALS, SETTING, METHODS: Samples were obtained from patients undergoing IVF treatments with informed consent and institutional review board approval. The Infinium HumanMethylation450 BeadChip microarray was used to identify histone-retained CpG island genes and genomic regions showing differences in sperm DNA methylation between the Good Group and the Poor Group. Pathway and gene network analysis for the significantly altered genes was performed, and targeted DNA methylation validation was completed for 23 genes and two imprinting control regions. Sperm miRNA profiles were assessed using the TaqMan® Human MicroRNA Array Card, with corresponding blastocyst mRNA gene expression examined by qRT-PCR. MAIN RESULTS AND THE ROLE OF CHANCE: Our study is the first to investigate unexplained male factor infertility while significantly eliminating confounding female factors from our sample population by using only young fertile donor oocytes. We identified 1634 CpG sites located at retained histone CpG island regions that had significant sperm DNA methylation differentials between the two embryogenesis groups (P < 0.05). A largely hypermethylated profile was evident in the Good Group, with a small but distinct and statistically significant shift (P < 0.05) observed in the Poor Group. Genes involved in embryonic development were highly represented among histone-retained CpG sites with decreased methylation in the Poor Group (P < 0.05). Ten significantly altered sperm miRNAs (P < 0.05), correlated with altered target gene mRNA expression in the blastocysts from the Poor Group (P < 0.05). Taken together, significantly impacted sperm miRNA and target transcript levels in blastocysts from the Poor Group may contribute alongside aberrant sperm DNA methylation to the compromised blastocyst development observed. LIMITATIONS, REASONS FOR CAUTION: Our examination of CpG island regions restricted to retained histones represents only a small part of the sperm epigenome. The results observed are descriptive and further studies are needed to elucidate the functional effects of differential sperm DNA methylation on unexplained male factor infertility and blastocyst development. WIDER IMPLICATIONS OF THE FINDINGS: Slight epigenetic changes in sperm may have a cumulative effect on fertility and embryonic developmental competence. Knowledge of sperm epigenetics and inheritance has important implications for future generations, while providing evidence for potential causes of unexplained male factor infertility. STUDY FUNDING/COMPETING INTEREST(S): No external funding was used for this study. None of the authors have any competing interest.