Early Reproductive Development Research Articles

P-316 Fertility, pregnancy outcomes, and perinatal results related to experimental endometriosis in a murine model

Abstract Study question Does the transplantation of endometrial tissue fragments into the peritoneal wall of female mice reduce natural fertility and lead to compromised obstetric and perinatal outcomes? Summary answer Experimental murine endometriosis did not compromise fertility, but females with more ectopic endometria had litters with reduced birth weight and fewer offspring at weaning. What is known already It has been demonstrated that endometriosis is strongly associated with subfertility and is increasingly linked to late pregnancy complications, such as those caused by placental insufficiency. At the molecular level, the presence of ectopic endometrium may perturb the endometrial hormonal, cellular, and immunological milieu, negatively influencing the decidualization, placentation, and developmental programming of the embryo. However, it remains unclear if and how such early aberrant reproductive development relates to pregnancy outcomes in endometriosis. Thus, the use of animal models may be very useful in deepening our understanding of the pathogenic mechanisms of infertility and compromised pregnancy outcomes related to endometriosis. Study design, size, duration C57BL/6J female mice were divided into three groups: Control (42 animals underwent sham surgery), Endometriosis1 (19 animals underwent surgery for transplantation of one fragment of endometrial tissue on each side of the peritoneal wall), and Endometriosis2 (23 animals received two fragments on each side). The maximum experiment duration was 15 weeks, including a 4-week rest period post-surgery, up to 4 weeks for mating, 3 weeks for delivery, and 4 weeks until weaning of the offspring. Participants/materials, setting, methods Reproductive-age female mice from the three groups had a 4-week rest period post-surgery. Then, they were paired for mating with proven fertile males and checked for the presence of a vaginal plug daily. Pregnant females were euthanized after weaning the litter at 4 weeks. Tissue from the implants was collected for histological analysis. The occurrence of pregnancy, gestation duration, litter size, pup weight at birth and weaning, and mating return were compared among the groups. Main results and the role of chance All females in the experimental groups were confirmed for the presence of ectopic endometrium through histology. No statistically significant differences were observed in pregnancy rates, gestation duration, litter size at birth, and the number of live pups at birth when comparing the three groups (C, E1, and E2). However, the litter weight at birth was significantly lower in group E2 (Medial (Md): 8g; Percentile (P)25: 6; P75: 10) than in groups C (Md: 10g; P25: 8; P75: 10) and E1 (Md: 10g; P25: 10; P75: 12), and significantly higher in E1 compared to C. The number of live pups at weaning was significantly lower in E2 (Md: 5; P25: 3.5; P75: 7) compared to groups C (Md: 7; P25: 5; P75: 7) and E1 (Md: 8; P25: 7; P75: 8), and significantly higher in E1 than in C. The litter weight at weaning was significantly lower in E2 (Md: 96g; P25: 67; P75: 109) than in both E1 (Md: 116g; P25: 110; P75: 128) and C (Md: 104g; P25: 96; P75: 112). The litter weight at weaning was higher in E1 compared to group C. There was no statistically significant difference in mating return rates among the three groups. Limitations, reasons for caution This study is limited by the use of an animal model that does not have spontaneous endometriosis. Wider implications of the findings Litters from the E2 group may have had impaired development during gestation, as indicated by lower birth weight and higher perinatal mortality. Although this is not an ideal translational model for studying endometriosis-related infertility, it may serve as a suitable model for evaluating gestational and perinatal outcomes associated with endometriosis. Trial registration number Not applicable

PaWOX3 and PaWOX3B Regulate Flower Number and the Lip Symmetry of Phalaenopsis.

The standout characteristic of the orchid perianth is the transformation of the upper median petal into a distinctively formed lip, which gives orchid flowers their typically zygomorphic symmetry and makes them the most popular ornamental plants worldwide. To study orchid flower development, two WUSCHEL-related homeobox (WOX) genes, PaWOX3 and PaWOX3B, were identified in Phalaenopsis. PaWOX3 and PaWOX3B mRNAs accumulate abundantly during early reproductive development and perianths of young buds, significantly decreasing in mature flowers and absent in vegetative leaves and roots. PaWOX3 and PaWOX3B virus-induced gene silencing (VIGS) knockdown in Phalaenopsis significantly reduces floral bud numbers, suggesting that PaWOX3/PaWOX3B may be involved in flower initiation. Transgenic Arabidopsis ectopically expressing repressor forms of PaWOX3/PaWOX3B and their Oncidium ortholog, OnPRS, exhibit lateral organ development defects, implicating these genes likely have function in regulating growth and differentiation for lateral organs. Neither PaWOX3, PaWOX3B single nor PaWOX3/PaWOX3B double VIGS Phalaenopsis altered the flower morphology. Interestingly, double silencing of PaWOX3 or PaWOX3B with OAGL6-2, which controlled the identity/formation of lips, altered the symmetry of 'BigLip' produced in OAGL6-2 VIGS. This result indicated that the levels of PaWOX3/PaWOX3B are still sufficient to maintain the symmetry for the OAGL6-2 VIGS 'BigLip'. However, the symmetry of the OAGL6-2 VIGS 'BigLip' cannot be maintained once the expression of PaWOX3 or PaWOX3B is further reduced. Thus, in addition to controlling lip identity, this study further found that OAGL6-2 could cooperate with functionally redundant PaWOX3/PaWOX3B in maintaining the symmetric axis of lip.

RETINOBLASTOMA RELATED 1 switches mitosis to meiosis in rice

The transition from mitosis to meiosis is a critical event in the reproductive development of all sexually reproducing species. However, the mechanisms that regulate this process in plants remain largely unknown. Here, we find that the rice (Oryza sativa L.) protein RETINOBLASTOMA RELATED 1 (RBR1) is essential to the transition from mitosis to meiosis. Loss of RBR1 function results in hyper-proliferative sporogenous-cell-like cells (SCLs) in the anther locules during early stages of reproductive development. These hyper-proliferative SCLs are unable to initiate meiosis, eventually stagnating and degrading at late developmental stages to form pollen-free anthers. These results suggest that RBR1 acts as a gatekeeper of entry into meiosis. Furthermore, cytokinin content is significantly increased in rbr1 mutants, whereas the expression of type-B response factors, particularly LEPTO1, is significantly reduced. Given the known close association of cytokinins with cell proliferation, these findings imply that hyper-proliferative germ cells in the anther locules may be attributed to elevated cytokinin concentrations and disruptions in the cytokinin pathway. Using a genetic strategy, the association between germ cell hyper-proliferation and disturbed cytokinin signaling in rbr1 has been confirmed. In summary, we reveal a unique role of RBR1 in the initiation of meiosis; our results clearly demonstrate that the RBR1 regulatory module is connected to the cytokinin signaling pathway and switches mitosis to meiosis in rice.

Insights into the early gonad development of the protogynous ballan wrasse (Labrus bergylta)

The reproductive biology of ballan wrasse, a popular cleaner fish species now bred in captivity, is not well understood particularly the early stages of development and their accompanying endocrine changes. The protogynous nature of this species has been well documented in the wild, although it is still unclear if it follows a monandric or diandric hermaphroditism. In this study, we investigated the gonad development of hatchery produced ballan wrasse (n = 102) between 3 and 185 g body weight and 5–19 cm total length. About 90 % of all fish examined showed gonads undergoing primary oocyte growth, exhibiting chromatin-nucleolar stage, perinucleolar stage, and early cortical alveolar stage. Mean size of fish that entered primary oocyte growth (i.e., chromatin-nucleolar stage) is 26.2 ± 1.6 g and 10.9 ± 0.2 cm. The remaining 10 % were undifferentiated and constituted the smallest fish examined. The gonadosomatic index and circulating levels of 17β-Estradiol and Testosterone increased as fish progressed through the ovarian development. Expression levels of selected gonadal genes including cyp19a1a, figla, bmp15, dmrt1, amh, fshr, and lhr were also quantified. To the best of our knowledge, this paper is the first to investigate the early reproductive development of ballan wrasse, providing important insights into histological and endocrine changes. Results from this study provide basis for further research on the reproductive physiology of ballan wrasse as well as for the development of broodstock management techniques in this economically important cleaner fish.

2. Gilt progeny dams have higher litter sizes compared to sow progeny dams which may be influenced by differences in early reproductive development

2. Gilt progeny dams have higher litter sizes compared to sow progeny dams which may be influenced by differences in early reproductive development

Effect of sunflower seed husk and rape straw in total mixed ration during peri-puberty on testis development in Hu lambs

Sunflower seed husk (SH) and rape straw (RS) are widely used in mutton sheep husbandry in the Hexi Corridor of China. This study explored whether or not a pre-puberty lamb diet consisting of sunflower SH and RS affects ram testis development. Fifty 60-day-old healthy Hu ram lambs (average body weight of 22.3 ± 0.1 kg) were randomly divided into two groups. In each treatment group, 25 lambs were allocated to five pens (five lambs per pen). The SH group received a diet containing 20 % SH, and the RS group received a diet containing 20 % RS. After a 77-day feeding trial, three lambs, except for the heaviest and lightest in each pen, were humanely slaughtered and investigated. Results revealed that the two groups had no difference in body weight (39.23 ± 0.52 kg vs. 38.53 ± 0.46 kg, P = 0.321). However, the testis weight (288.74 ± 23.45 g vs. 215.18 ± 17.96 g), volume (294.29 ± 25.39 mL vs. 208.67 ± 16.63 mL), epididymis weight (38.38 ± 1.44 g vs. 33.86 ± 1.37 g), testicular index (0.74 ± 0.06 vs 0.56 ± 0.05), and epididymis sperm count (4.88 ± 0.92 ×109/g vs. 2.27 ± 0.69 ×109/g) in the SH group were significantly higher than those in the RS group (P < 0.05). SH reduced the mtDNA relative copy number (P < 0.01) in the testis and increased the total antioxidant capacity (3.41 ± 0.94 U/mg prot vs. 1.54 ± 0.38 U/mg prot, P = 0.05). RNA sequencing results showed 50 differentially expressed genes, with 27 genes upregulated and 23 genes downregulated in the SH group compared with the RS group. KEGG analysis revealed that the pathways related to polyunsaturated fatty acid metabolism, and vitamin digestion and absorption were enriched. The findings suggest that feeding SH in the early reproductive development stage of lambs is more beneficial for testis development and spermatogenesis compared with feeding RS.

Complications and outcomes of pregnancy in patients with antiphospholipid antibodies during various treatment methods

Introduction. Antiphospholipid antibodies (APAs) exert multifaceted effects on the course of pregnancy by disrupting microcirculation, affecting the hemostasis, as well as damaging the endothelial membranes, leading to early reproductive loss and development of placenta-associated complications depending on the affected gestation stage. Planning and management of pregnancy in women in the absence of criteria for complete antiphospholipid syndrome (APS) currently remains unresolved issue. The absence of generally accepted treatment standards for this category of patients and inability to substantiate the diagnosis according to the APS classification criteria complicate selection of therapeutic tactics.Aim: to conduct a comparative analysis of therapy-based complications and outcomes of pregnancy in APA carriers.Materials and Methods. During the period 2019–2021 a prospective study of 150 patients who entered pregnancy with aggravated obstetric and gynecological history, serum APA level was examined. Considering the risks of developing obstetric and thrombotic complications, all patients were prescribed prophylactic doses of low molecular weight heparins (LMWHs) and low doses of acetylsalicylic acid (ASA). The patients were divided into 3 groups using a random number generator. Group 1 (n = 50), in addition to the prescribed LMWH (enoxaparin sodium 40 mg 1 time per day) and ASA (150 mg 1 time per day), also underwent plasmapheresis (PF) 4 sessions per 1 course in 6–8, 12–14 and 22–24 weeks of pregnancy; group 2 (n = 50) received courses of intravenous immunoglobulins (IVIG) at a course dose of 300 ml (15 g) simultaneously; group 3 (n = 50) received no additional therapies. Rate of pregnancy complications was comparatively assessed – development of fetal growth retardation (FGR), low birth weight fetus, gestational arterial hypertension (AH), moderate and severe preeclampsia (PE), anemia and delivery outcomes.Results. It was found that in group 3 there was a higher incidence of gestational hypertension (p2,3 &lt; 0.0001), moderate PE (p 1,3 =0.071; p 2,3 = 0.0019), low weight fetus for gestational age (p2,3 = 0.0002) and FGR (p2,3 = 0.003). In group 1, compared with group 2, there were more often observed small weight for gestational age fetus (p1,2 = 0.018) and FGR (p1,2 = 0.024), gestational hypertension (p1,2 = 0.0008), anemia (p1,2 &lt; 0.0001) and latent iron deficiency (p1,2 &lt; 0.0001). Also, groups 2 and 3 vs. group 1 were more likely to have intrahepatic cholestasis during pregnancy (p1,2 = 0.013; p1,3 = 0.003).Conclusion. In the group of patients receiving complex therapy consisting of LMWHs prophylactic doses, low ASA doses and IVIG courses, the risks of developing placenta-associated complications and iron deficiency were reduced compared to other groups indicating about a higher efficiency of this therapy regimen. However, the development of intrahepatic cholestasis during pregnancy was less common in the group of patients receiving PF courses, in contrast to using IVIG courses, which can be accounted for by additional effect of efferent therapeutic methods and should be taken into account in a differentiated approach for management of patients with liver and gallbladder pathology.

Изменчивость репродуктивного развития полусибов плюсовых деревьев Pinus sibirica Du Tour

Selection of Pinus sibirica Du Tour plus trees by seed productivity, breeding of half-sibs, characterized by an abundance of cones and pollen, serve as the basis for the creation of high-yielding target plantations. The aim of the research was to select families with a large percentage of early reproductive development's trees, and specimens with a lot of cones and shoots with male inflorescences. Also, to select specimens in different families of Pinus sibirica Du Tour plus trees by early reproductive development, taking into account the formation of cones and pollen. The subject of the study was the seed progeny of Pinus sibirica Du Tour trees at biological age of 31-35 years, grown on the territory of the arboretum of Reshetnev University, the site «Prodolzhenie geoshkoly» (N 55.9666666667, E 92.6166666667, 287 meters above sea level). Initially, five plus trees growing in Irkutsk (60/24) and Novosibirsk regions (98/62, 102/66, 106/70 and 110/74) for reproduction among certified plus trees by seed productivity were selected. The growing conditions and indicators of plus trees from the certificate drawn up by the commission during their certification are given. Cones from these trees were collected, then seeds were sown and planting material was grown, which was transplanted to the site of the arboretum of Reshetnev University. In 2018, 2021 and 2022, the number of cones and microstrobiles in early reproductive development semis by using binoculars was determined. The variability by early reproductive development of half-sibs in the formation of cones both between families of plus trees (7.8-25.3 psc.), and within families (60/24 – 2-22 psc.; 98/62 – 2-115 psc.; 102/66 – 4-45 psc.;106/70 – 1-54 psc.; 110/74 – 1-15 psc.) is noted. The selection of specimens by the size of the cones was not carried out. Some specimens formed both cones and pollen: 82-7, 85-10 and 87-13 in the family of the plus trees 60/24; 72-11, 79-2, 79-3, 79-4, 79-8, 80-3, 81-4, 85-22 (family 98/62); 83-1,83-6, 83-9, 84-2, 84-4, 86-9, 86-1 (family 102/66); 78-3, 78-9, 81-8, 81-9, 81-10, 85-19 (family 106/70); 79-9, 84-12, 86-5 (family 110/74). Selected specimens have a huge value and can be widely used in their vegetative reproduction and the creation of high-yielding plantations of early reproductive development.

Transcriptional Signatures in Contrasting Cultivars of Barley During Early Reproductive Development of Meristem

Transcriptional Signatures in Contrasting Cultivars of Barley During Early Reproductive Development of Meristem

Effect of corn straw or corncobs in total mixed ration during peri-puberty on testis development in Hu lambs

Corn straw and corncobs contain large amounts of crude fibers and are widely used in mutton sheep husbandry in northwest China. The aim of this study was to determine whether feeding with corn straw or corncobs affects lamb testis development. A total of 50 healthy Hu lamb at two-month-old (average body weight of 22.3 ± 0.1 kg) were randomly and equally divided into two groups, and the lambs were equally allocated to five pens in each group. The corn straw group (CS) received a diet containing 20% corn straw, whereas the corncobs group (CC) received a diet containing 20% corncobs. After a 77-day feeding trial, the lambs, except the heaviest and lightest in each pen, were humanely slaughtered and investigated. Results revealed no differences in body weight (40.38 ± 0.45 kg vs. 39.08 ± 0.52 kg) between the CS and CC groups. Feeding diet containing corn straw significantly (P < 0.05) increased testis weight (243.24 ± 18.78 g vs. 167.00 ± 15.20 g), testis index (0.60 ± 0.05 vs. 0.43 ± 0.04), testis volume (247.08 ± 19.99 mL vs. 162.31 ± 14.15 mL), diameter of seminiferous tubule (213.90 ± 4.91 μm vs. 173.11 ± 5.93 μm), and the number of sperm in the epididymis (49.91 ± 13.53 × 108/g vs. 19.34 ± 6.79 × 108/g) compared with those in the CC group. The RNA sequencing results showed 286 differentially expressed genes, and 116 upregulated and 170 downregulated genes were found in the CS group compared with the CC group. The genes affecting immune functions and fertility were screened out. Corn straw decreased the mtDNA relative copy number in the testis (P < 0.05). These results suggest that compared with corncobs, feeding corn straw in the early reproductive development stage of lambs increased the testis weight, diameter of seminiferous tubule and the number of cauda sperm.

Water quality and the CO2-carbonate system during the preconditioning of Pacific oyster (Crassostrea gigas) in a recirculating aquaculture system

The continued increase of the demand for seed of the Pacific oyster (Crassostrea gigas) has driven the aquaculture industry to produce land-based hatcheries using broodstock conditioning. This has led to the need to create closed systems to control the main factors involved in reproduction (temperature and food). Additionally, reproductive synchronization of broodstocks may be considered to ensure homogeneous maturation and spawning among the organisms. In this work, we synchronized the broodstock reproductive stage of Pacific oysters in a recirculating aquaculture system (RAS) using a “preconditioning” process and evaluated the effect of the water quality and the CO2-carbonate system on preconditioned broodstock. The oysters were kept at 12 °C for 45 days in a RAS containing a calcium reactor (C2) or without a calcium reactor (C1, control). Water quality parameters were measured daily, and the oyster’s condition and reproductive development were monitored using condition index, biometrics, and histology, on Days 0, 20, and 45. C1 and C2 systems kept the water quality within the ranges reported as favorable for bivalves. The calcium reactor kept the pH (8.03–8.10), alkalinity (200 mg/L as CaCO3), CO32− (≤ 80 µmol/kg), and Ω aragonite (≤ 1) closer to the ranges reported as optimal for bivalves. However, no significant differences were detected in the total weight and the condition index in C1 and C2. The preconditioning allowed to maintain the organisms in early reproductive development, allowing gametogenesis synchronization to start maturation.

The energetic cost of early reproductive development in juvenile Antarctic krill at the Western Antarctic Peninsula

The overwinter survival mechanisms of Antarctic krill, Euphausia superba, are poorly characterized, especially for juveniles. It has been suggested that juveniles adopt a mix of strategies characteristic of both larvae and adults. Like larvae, they may feed opportunistically throughout winter when food is available, and like adults they may be able to suppress their metabolism when food is scarce. In this study we look at the overwinter strategies of juvenile krill and how their reproductive development changes when energy input exceeds what is necessary for survival. We take a closer look at how the sexual maturation of juvenile krill progresses in response to different environmental conditions throughout the fall and winter. We exposed juvenile Antarctic krill to four different “food environment scenarios”, supplementing them with various diets from May to September 2019 that were representative of environmental conditions that they may encounter in different regions of the Western Antarctic Peninsula during autumn and winter. Each month, we measured the physiology and condition of the krill, and assessed the reproductive development of females. We found that when female juvenile krill have greater energy reserves than what is needed to survive the winter, they will begin to sexually mature. Further, when there are sufficient levels of the fatty acids eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and 16:4 (n-1), krill are likely to be in a more reproductive advanced stage. However, when lipids, EPA, DHA and 16:4 (n-1) are depleted throughout the winter, juvenile female krill lose their ability to develop reproductively. We also found that sexual development is an energy intensive process that requires high respiration rates in juvenile krill. Furthermore, when juvenile females expend energy maturing, their physiological condition declines. This trade-off between early reproductive development and condition in juvenile female krill has important implications for individual health and population fecundity. Gaining a better understanding of the mechanisms behind juvenile krill winter survival strategies and their consequences will allow us to predict how future change at the western Antarctic Peninsula may affect krill population dynamics, especially in light of a warming climate.

A rice single cell transcriptomic atlas defines the developmental trajectories of rice floret and inflorescence meristems.

Rice inflorescence development determines yield and relies on the activity of axillary meristems (AMs); however, high-resolution analysis of its early development is lacking. Here, we have used high-throughput single-cell RNA sequencing to profile 37 571 rice inflorescence cells and constructed a genome-scale gene expression resource covering the inflorescence-to-floret transition during early reproductive development. The differentiation trajectories of florets and AMs were reconstructed, and discrete cell types and groups of regulators in the highly heterogeneous young inflorescence were identified and then validated by in situ hybridization and with fluorescent marker lines. Our data demonstrate that a WOX transcription factor, DWARF TILLER1, regulates flower meristem activity, and provide evidence for the role of auxin in rice inflorescence branching by exploring the expression and biological role of the auxin importer OsAUX1. Our comprehensive transcriptomic atlas of early rice inflorescence development, supported by genetic evidence, provides single-cell-level insights into AM differentiation and floret development.

Gene expression profiling by cDNA‐AFLP reveals potential candidate genes for imazapyr‐induced male sterility in sunflower

AbstractChemically induced male sterility (CIMS) is a valuable tool for hybrid breeding in crops. In this context, the acetohydroxyacid synthase (AHAS)‐inhibiting imazapyr has been proposed to induce male sterility in Imisun sunflower (Helianthus annuus L.) when applied during early reproductive development in a two‐fold higher than recommended field dose. This study aimed to perform a comparative transcriptomic analysis to find out the molecular bases of the gametocidal effect of imazapyr on two Imisun genotypes differing in their herbicide resistance level: completely resistant and intermediate resistant. The CIMS of treated plants was confirmed at physiological and anatomical levels. A total of 948 differentially expressed transcript‐derived fragments (TDFs) were identified by complementary DNA amplified fragment‐length polymorphism (cDNA‐AFLP) analysis at the three most informative developmental stages of anther development. Several genes related to xenobiotic metabolism and stress, including cytochrome P450 monooxygenases and ATP‐binding cassette transporters, were identified in the completely resistant genotype. Interestingly, a group of genes, including protein kinases, sulfonylurea receptor 1, aspartic signal peptide‐peptidase, and specific membrane transporters, were recognized from the differential TDFs simultaneously detected in both genotypes. Some of these transcripts were validated by quantitative real‐time polymerase chain reaction (qPCR) in both genotypes. Therefore, these transcripts could be exclusively related to the CIMS in sunflower. Based on the results, we suggest a probable model of action for the gametocidal effect of imazapyr. Our research provides new insights into the molecular mechanisms of male sterility induction in sunflower.

Dietary supplementation of grape seed tannin extract stimulated testis development, changed fatty acid profiles and increased testis antioxidant capacity in pre-puberty hu lambs

Grape seed tannin extract (GPE) from wine grape pomace has many effective anti-oxidative effects and is used as a promising natural feed additive in the animal feed industry. This study investigated the effect of GPE as a source of tannin on the antioxidant capacity and testis development in Hu lambs. Twenty-seven 3-month-old ram lambs were randomly assigned to three groups. For each treatment group, nine lambs were allocated to nine pens (one lamb per pen). The lambs in the control group were fed a control diet without GPE for 61 days from D21 to D80. Group I (TAN1) was fed with 0.36% GPE diet, and Group II (TAN2) was fed with 0.72% GPE diet. After an 81-day feeding trial, all lambs except the heaviest and lightest in each group were humanely slaughtered and investigated. Results showed that feeding GPE did not affect the body weight, average daily gain, dry matter intake, scrotal circumference, and testis index. Meanwhile, feeding with 0.36% GPE diet increased testis weight, testis volume, and epididymis weight (P ≤ 0.05) compared with those of the control, but no difference was found between TAN1 and TAN2 groups. Copper-zinc superoxide dismutase (Cu-ZnSOD), steroid acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (P450scc), follicle-stimulating hormone receptor (FSHR), elongation of very long chain fatty acid protein 2 (ELOVL2), fatty acid desaturase (FADS2), and proliferating cell nuclear antigen (PCNA) mRNA in TAN1 and TAN2 groups were significantly up-regulated (P < 0.05). GPE also markedly increased the antioxidant status of testis. Compared with the control group, the treatment groups showed significantly increased superoxide dismutase (SOD) activity (314.23 ± 18.64 U/mg prot in control, 505.22 ± 63.47 U/mg prot in TAN1 and 587.88 ± 55.94 U/mg prot in TAN2, P < 0.05) and total antioxidant capacity (T-AOC) (98.23 ± 18.99 U/g prot in control, 202.15 ± 34.19 U/g prot in TAN1 and 189.57 ± 18.95 U/g prot in TAN2, P < 0.05). Consuming 0.72% GPE also changed the fatty acid profiles in testis with increased C15:1, C22:6n3, and total n-3 fatty acids (P < 0.05) but decreased C22:5n3 (P < 0.05). Therefore, feeding lambs with GPE stimulated testis seminiferous tubule development and increased the number of Sertoli cells (10.56 ± 0.44 in control, 14.10 ± 0.57 in TAN1 and 13.60 ± 0.42, P < 0.05), and seminiferous tubule diameter (109.30 ± 4.56 μm in control, 164.49 ± 5.37 μm in TAN1 and 146.56 ± 4.53 μm in TAN2, P < 0.05). These results suggested that feeding GPE in the early reproductive development stage of lambs upregulated the expression of antioxidative, steroidogenesis, and polyunsaturated fatty acid metabolism-related genes, changed the fatty acid profiles, increased the antioxidant capacity in lamb's testis, and contributed to testis development and spermatogenesis.

Reproductive Hormone Concentrations and Associated Anatomical Responses: Does Soy Formula Affect Minipuberty in Boys?

Soy formula feeding is common in infancy and is a source of high exposure to phytoestrogens, documented to influence vaginal cytology in female infants. Its influence on minipuberty in males has not been established. To assess the association between infant feeding practice and longitudinally measured reproductive hormones and hormone-responsive tissues in infant boys. The Infant Feeding and Early Development study was a prospective cohort of maternal-infant dyads requiring exclusive soy formula, cow milk formula, or breast milk feeding during study follow-up. In the 147 infant boy participants, serum testosterone, luteinizing hormone, stretched penile length, anogenital distance, and testis volume were longitudinally assessed from birth to 28 weeks. We examined feeding-group differences in age trajectories for these outcomes using mixed-effects regression splines. Median serum testosterone was at pubertal levels at 2 weeks (176 ng/dL [quartiles: 124, 232]) and remained in this range until 12 weeks in all feeding groups. We did not observe differences in trajectories of hormone concentrations or anatomical measures between boys fed soy formula (n = 55) and boys fed cow milk formula (n = 54). Compared with breastfed boys (n = 38), soy formula-fed boys had a more rapid increase in penile length (P = .004) and slower initial lengthening of anogenital distance (P = .03), but no differences in hormone trajectories. Reproductive hormone concentrations and anatomical responses followed similar trajectories in soy and cow milk formula-fed infant boys. Our findings suggest that these measures of early male reproductive development do not respond to phytoestrogen exposure during infancy.

Effect of feeding linseed diet on testis development, antioxidant capacity, and epididymal cauda sperm concentration in Chinese Hu lamb

Polyunsaturated fatty acids (PUFAs) are essential for mammalian testis development and sperm function. However, PUFAs that are contained in linseed oil are easily oxidized in the diet and biohydrogenated in the rumen. In this study, we investigated the effect of linseed as a source of PUFAs on the antioxidant capacity and testis development in Hu lamb. Seventy-five 3-month-old lambs were randomly assigned to three groups. Within each treatment group, 25 lambs were allocated to five pens (five lambs per pen). The lambs in the control group were fed a control diet without linseed for 42 days from D22 to D63. Group I (BS28) was fed a control diet from D22 to D35 and 8% linseed diet from D36 to D63. Group II (BS42) was fed an 8% linseed diet for 42 days from D22 to D63. After 63-day feeding trial, all lambs except the heaviest and lightest in each pen were humanely slaughtered and investigated. Results revealed that feeding linseed did not affect the body weight, scrotal circumference, and testis weight, whereas feeding linseed for 42 days increased the epididymis weight (37.85 ± 1.61 g vs. 32.09 ± 1.06 g, P < 0.05) compared with the control group. Feeding lambs with linseed for 42 days also significantly upregulated the expression of antioxidative (glutathione peroxidase 4 and copper-zinc superoxide dismutase), steroidogenesis (3β-hydroxysteroid dehydrogenase and steroid acute regulatory protein), and PUFA metabolism-related genes (fatty acid desaturase 2 and elongation of very long-chain fatty acid protein 2) and proliferating cell nuclear antigen mRNA (P < 0.05). It also increased the relative expression of mitochondrial DNA (P < 0.05), total antioxidant capacity (0.230 ± 0.019 mmol/mgprot vs. 0.175 ± 0.011 mmol/mgprot, P < 0.05), and superoxide dismutase (1661.467 ± 147.117 U/mgprot vs. 1158.891 ± 98.850 U/mgprot, P < 0.05) in testicular tissue but decreased the cholesterol concentration (0.331 ± 0.073 mmol/mgprot vs. 0.671 ± 0.092 mmol/mgprot, P < 0.05) compared with the control group. Therefore, feeding lambs with linseed for 42 days stimulated seminiferous tubule development and increased the number of Sertoli cells (20.71 ± 0.89 vs. 17.6 ± 0.73, P < 0.05), epididymal cauda lumina diameter (638.26 ± 22.32 μm vs. 444.41 ± 34.80 μm, P < 0.05), and the number of sperm in the epididymal cauda (68.91 ± 7.06 × 108/g vs. 36.61 ± 7.50 × 108/g). All these results suggested that feeding linseed in the early reproductive development stage of lambs upregulated the expression of antioxidative, steroidogenesis, and PUFA metabolism-related genes; increased the antioxidant capacity in lamb’s testis; and contributed to testis development and spermatogenesis.

Molecular and morphological sex differentiation in sablefish (Anoplopoma fimbria), a marine teleost with XX/XY sex determination.

Phenotypic sex of an organism is determined by molecular changes in the gonads, so-called molecular sex differentiation, which should precede the rise of cellular or anatomical sex-distinguishing features. This study characterized molecular and morphological sex differentiation in sablefish (Anoplopoma fimbria), a marine teleost with established XX/XY genotypic sex determination. Next generation sequencing was conducted on sablefish ovarian and testicular mRNAs to obtain sequences for transcripts associated with vertebrate sex determination and differentiation and early reproductive development. Gene-specific PCRs were developed to determine the distribution and ontogenetic gonadal expression of transcription, growth, steroidogenic and germline factors, as well as gonadotropin and steroid receptors. Molecular changes associated with sex differentiation were first apparent in both XY- and XX-genotype sablefish at~60mm in body length and prior to histological signs of sex differentiation. The earliest and most robust markers of testicular differentiation were gsdf, amh, dmrt1, cyp11b, star, sox9a, and fshr. Markedly elevated mRNA levels of several steroidogenesis-related genes and ar2 in differentiating testes suggested that androgens play a role in sablefish testicular differentiation. The earliest markers of ovarian differentiation were cyp19a1a, lhcgr, foxl2, nr0b1, and igf3. Other transcripts such as figla, zp3, and pou5f3 were expressed predominantly in XX-genotype fish and significantly increased with the first appearance and subsequent development of primary oocytes. This study provides valuable insight to the developmental sequence of events associated with gonadal sex differentiation in marine teleosts with XX/XY sex determination. It also implicates particular genes in processes of male and female development and establishes robust molecular markers for phenotypic sex in sablefish, useful for ongoing work related to sex control and reproductive sterilization.

The regulatory landscape of early maize inflorescence development

BackgroundThe functional genome of agronomically important plant species remains largely unexplored, yet presents a virtually untapped resource for targeted crop improvement. Functional elements of regulatory DNA revealed through profiles of chromatin accessibility can be harnessed for fine-tuning gene expression to optimal phenotypes in specific environments.ResultHere, we investigate the non-coding regulatory space in the maize (Zea mays) genome during early reproductive development of pollen- and grain-bearing inflorescences. Using an assay for differential sensitivity of chromatin to micrococcal nuclease (MNase) digestion, we profile accessible chromatin and nucleosome occupancy in these largely undifferentiated tissues and classify at least 1.6% of the genome as accessible, with the majority of MNase hypersensitive sites marking proximal promoters, but also 3′ ends of maize genes. This approach maps regulatory elements to footprint-level resolution. Integration of complementary transcriptome profiles and transcription factor occupancy data are used to annotate regulatory factors, such as combinatorial transcription factor binding motifs and long non-coding RNAs, that potentially contribute to organogenesis, including tissue-specific regulation between male and female inflorescence structures. Finally, genome-wide association studies for inflorescence architecture traits based solely on functional regions delineated by MNase hypersensitivity reveals new SNP-trait associations in known regulators of inflorescence development as well as new candidates.ConclusionsThese analyses provide a comprehensive look into the cis-regulatory landscape during inflorescence differentiation in a major cereal crop, which ultimately shapes architecture and influences yield potential.

Quantitative real-time PCR based evaluation and validation of reference genes in Gossypium arboreum

Estimation of gene expression levels plays a crucial role in understanding the function of the target gene(s). Intersample variance in gene expression can be more precisely measured if transcripts levels are accurately normalized. Normalization is pre-requisite step prior to the determination of candidate gene expression by qPCR. In this study conducted at ICAR-Central Institute for Cotton Research, Nagpur during 2015–16, six candidate reference genes, viz. actin4 (ACT4), actin7(ACT7), RNA Helicase (RNAH), Serine/threonine-protein phosphatase PP2A-1(PP2A1), ubiquitin7 (UBQ7) and α tubulin (αTUB) were systematically analysed for their expression patterns in different tissues pertaining to three development stages of cotton namely seedling, early reproductive and fiber development. The study has identified actin-4/actin-7/ubiquitin-7 as the most ideal reference genes for fiber development stages whereas actin-4/ ubiquitin-7 and actin-7/RNA helicases for seedling and early reproductive development stages, respectively. Validation of identified reference genes for relative expression analysis of Gacobl9, a COBRA-like protein, demonstrated their usefulness in qPCR analysis in Gossypium arboreum.