17ß-Trenbolone activates androgen receptor, upregulates transforming growth factor beta/bone morphogenetic protein and Wnt signaling pathways, and induces masculinization of caudal and anal fins in female guppies (Poecilia reticulata).

Sexually mature female guppies (Poecilia reticulata) were exposed to environmentally relevant concentrations (20, 200, and 2000 ng/L) of 17ß-trenbolone for four weeks. As evidenced by the increased caudal fin index and anal fins developing into gonopodium-like structures, exposed females displayed masculinized secondary sexual characteristics. Differential gene expression and subsequent pathway analysis of mRNA sequencing data revealed that the transcription of transforming growth factor beta/bone morphogenetic protein signaling pathway and Wnt signaling pathway were upregulated following 17ß-trenbolone exposure. Enzyme-linked immunosorbent assays showed that the bone morphogenetic protein 7 protein content was elevated after 17ß-trenbolone exposure. Finally, real-time PCR revealed that 17ß-trenbolone treatment significantly increased androgen receptor mRNA levels, and molecular docking showed potent interaction between 17ß-trenbolone and guppy androgen receptor. Furthermore, 17ß-trenbolone-induced masculinization of caudal and anal fins in female guppies, concomitant to the upregulated expression of differentially expressed genes involved in the above-mentioned two signaling pathways, was significantly inhibited by flutamide (androgen receptor antagonist). These findings demonstrated that 17ß-trenbolone masculinized fins of female guppies by activating the androgen receptor. This study revealed that 17ß-trenbolone could upregulate signaling pathways related to fin growth and differentiation, and eventually cause caudal and anal fin masculinization in female guppies.

Impact of a constant current electrical stimulation (CCES) system and hormonal growth-promoting (HGP) implants on meat quality and palatability of finished steers.

This study evaluated the effects of a constant current electrical stimulation (CCES) system and hormonal growth-promoting (HGP) implants on the quality and palatability of the longissimus thoracis et lumborum (LTL) from yearling-finished steers. The experiment used a total of 46 Angus cross steers, which were either non-implanted (n = 20) or implanted with trenbolone acetate and estradiol benzoate (n = 26). The CCES was applied to one side of each carcass during the slaughter process, whereas the other side remained unstimulated. Regardless of the application of HGP implants, the CCES reduced pH at 3 and 72 h post-mortem and shear force at all ageing times (P < 0.05), improved colour at 72 h post-mortem and during the retail display (P < 0.05), increased initial and overall tenderness (P < 0.01), and decreased the amount of perceived connective tissue and the proportion of trained panelists detecting spongy texture (P < 0.05) compared to meat from unstimulated carcass sides. Although CCES increased meat purge losses and reduced moisture content (P < 0.05), this did not affect meat juiciness (P > 0.10). CCES interacted with HGP to prevent increase in drip loss (P > 0.10), increase frequency of panelists detecting bloody/serumy flavour and typical texture, and reduce the proportion of panelists detecting rubbery texture in meat (P < 0.05). Regardless of stimulation treatment, meat from implanted animals had a more pronounced pH decline at 72 h post-mortem (P < 0.05) and a higher proportion of panelists finding no off-flavours (P < 0.05) or bloody/serumy flavour (P < 0.01) than non-implanted cattle. The CCES system tested in this study improved LTL quality and palatability of heavier beef carcasses.

Anabolic implants alter abundance of mRNA involved in muscle growth, metabolism, and inflammation in the longissimus of Angus steers in the feedlot.

The majority of beef cattle in the United States often receive at least one anabolic implant resulting in improved growth, feed efficiency, and environmental and economic sustainability. However, the physiological and molecular mechanisms through which anabolic implants increase skeletal muscle growth of beef cattle remain elusive. The objective of this study was to identify transcriptional changes occurring in skeletal muscle of steers receiving anabolic implants containing different steroid hormones. Forty-eight steers were stratified by weight into 1 of 4 (n = 12/treatment) implant treatment groups: (1) estradiol (ImpE2; 25.7 mg E2; Compudose, Elanco Animal Health, Greenfield, IN), (2) trenbolone acetate (ImpTBA; 200 mg TBA; Finaplix-H, Merck Animal Health, Madison, NJ), (3) combination (ImpETBA; 120 mg TBA + 24 mg E2; Revalor-S, Merck Animal Health), or (4) no implant (CON). Skeletal muscle biopsies were taken from the longissimus 2 and 10 d post-implantation. The mRNA abundance of 94 genes associated with skeletal muscle growth was examined. At 10 d post-implantation, steers receiving ImpETBA had greater (P = 0.02) myoblast differentiation factor 1 transcript abundance than CON. Citrate synthase abundance was increased (P = 0.04) in ImpETBA steers compared to CON steers. In ImpE2 steers 10 d post-implantation, muscle RING finger protein 1 decreased (P = 0.05) compared to CON steers, and forkhead box protein O4 decreased (P = 0.05) in ImpETBA steers compared to CON steers. Interleukin-6 abundance tended to be increased (P = 0.09) in ImpE2 steers compared to both ImpETBA and CON steers. Furthermore, interleukin-10 mRNA abundance tended to be increased (P = 0.06) in ImpTBA steers compared to ImpETBA steers. Leptin receptor abundance was reduced (P = 0.01) in both ImpE2 and ImpTBA steers when compared to CON steers. Abundance of phosphodiesterase 4B was increased (P = 0.04) in ImpTBA steers compared to CON steers 2 d post-implantation. Taken together, the results of this research demonstrate that estradiol increases skeletal muscle growth via pathways related to nutrient partitioning and mitochondria function, while trenbolone acetate improves steer skeletal muscle growth via pathways related to muscle growth.

Altrenogest treatment reduces the stress response of three-year-old warmblood mares during their initial equestrian training.

Horse mares are frequently treated with the progestin altrenogest with the aim to suppress estrous behavior and its negative impact on equestrian performance. Progestogens, however, also have sedative effects in males, and females of different species. The aim of our study was therefore to investigate altrenogest-induced changes in the stress response of female horses during initial equestrian training. Three-yr-old Warmblood mares were randomly assigned to treatment with altrenogest (ALT; 0.044 mg/kg once daily; n = 6) or sunflower oil (CON; n = 5) for 12 wk during training. At predefined steps of the training program (free movement, lunging without and with side reins, lunging with saddle, mounting of a rider, free riding, riding by an unfamiliar rider) salivary cortisol concentration, and heart rate were determined from 60 min before to 120 min after training. The same procedures were performed during repeated gynecologic examinations and 2 novel object tests. Bodyweight and body condition scores (BCS) were assessed at 4-wk intervals. During all training units, salivary cortisol concentration and heart rate increased (P < 0.001), but the increase was smaller in group ALT mares (time x treatment P < 0.001). Gynecologic examinations and novel object tests induced a much smaller increase in cortisol and heart rate (P < 0.001) than equestrian training with no difference between groups ALT and CON. Initially, bodyweight, and BCS decreased during training. The subsequent increase was larger in group ALT vs CON (time x treatment P < 0.05). In conclusion, altrenogest reduced the stress response of 3-yr-old mares to equestrian training. The use of altrenogest during equestrian competitions should therefore be reconsidered.

Effects of increased ambient temperature and supplemental altrenogest before pregnancy establishment in gilts.

Heat stress (HS) mitigation strategies are critically needed to combat the substantial economic effects on animal agriculture. The manifestations of seasonal infertility include delayed puberty onset, reduced conception rates, decreased litter size, and increased wean to estrus interval. To assess the effects of HS during early gestation and evaluate the benefit of supplemental altrenogest (ALT) as a mitigation strategy, 30 crossbred postpubertal gilts (157 ±â€…11 kg body weight) were subjected to estrous synchronization via 14 d oral administration of ALT. Artificial insemination during estrus was performed, and gilts were then placed into one of four treatment groups: HS (35 ±â€…1 °C for 12 h/31.60 ±â€…1 °C for 12 h) with (HSALT, n = 7) or without (HSCON, n = 7) 15 mg/d ALT supplementation or thermal neutral (TN; 20 ±â€…1 °C) conditions with (TNALT, n = 8) or without (TNCON, n = 8) 15 mg/d ALT supplementation until 12 d post-estrus (dpe). Administrating ALT occurred at 0600 hours from 3 to 12 dpe, and rectal temperatures (TR) and respiration rates (RR) were recorded. Blood was collected via jugular venipuncture on 0, 4, 8, and 12 dpe. Gilts were euthanized humanely at 12 dpe followed by the collection of ovarian tissue, and uterine flushing for conceptus collection. In HS compared with TN gilts, RR and TR were increased (P < 0.01) but unaffected by ALT supplementation. Feed intake was reduced (P < 0.01) by HS but unaltered by the ALT treatment. Corpora lutea (CL) weight was reduced (P < 0.01) in HSCON gilts when compared with TNCON and HSALT gilts despite progesterone concentrations in serum and luteal tissue not being affected by treatment (P ≥ 0.10). CL diameter was reduced (P ≤ 0.05) in HSALT gilts compared with other treatments. Interleukin-1ß (IL1B) uterine flush concentration was not affected (P > 0.20) by environment or ALT supplementation, although moderate (P = 0.06) interaction between environment and ALT existed, as IL1B concentration in TNALT was increased (P = 0.03) compared with TNCON gilts. While environment did not affect conceptus development (P = 0.90), ALT supplementation advanced conceptus elongation (P < 0.01). Collectively, these data demonstrate that HS may affect luteal development before pregnancy establishment, and ALT increases conceptus elongation by 12 dpe.

Control of parturition in hyperprolific sows by using altrenogest and double administrations of PGF2α.

The aim of the present study was to develop a protocol to reduce the variation in gestation length and synchronise the onset of parturition in sows by using altrenogest in combination with double administrations of prostaglandin F2alpha (PGF2α). In total, 188 Landrace x Yorkshire crossbred sows with parity numbers 3.1 ± 1.6 were included in the experiment. The sows were classified into two groups: CONTROL (n = 94) and TREATMENT (n = 94). CONTROL sows were allowed to farrow naturally, and TREATMENT sows were orally administered 20 mg/day of altrenogest starting when they entered the farrowing house (107.0 ± 2.0 days) until 113 (TREAT-113, n = 18), 114 (TREAT-114, n = 29) and 115 (TREAT-115, n = 47) days of gestation. The altrenogest-treated sows were administered PGF2α twice 6 h apart at 24 h after the withdrawal of altrenogest. The litters were randomly selected (25 and 26 litters from CONTROL and TREATMENT groups, respectively) to determine individual body weight at birth and at 24 h after birth. Gestation length of sows that farrowed naturally averaged 115.1 days (range 111-118), whereas gestation length of altrenogest-treated sows averaged 115.1-116.3 days (range 114-118). The colostrum yield of sows averaged 4.25 ± 1.19 kg and was not affected by the treatment (P > 0.05). Colostrum IgG in the CONTROL group was higher than in the TREAT-114 and TREAT-115 groups (P < 0.05) but did not differ significantly compared to the TREAT-113 group. The proportion of sows that farrowed during working hours (0700-1700 h) in the TREAT-113 group (72.3%) tended to be higher than in the CONTROL (46.4%, P = 0.053). The interval from the last altrenogest treatment until farrowing in the TREAT-113 group was longer than in the TREAT-114 and TREAT-115 groups (62.8, 40.7 and 34.6 h, respectively, P < 0.05). Similarly, the intervals from the first PGF2α administration to the onset of parturition in the TREAT-113 group (38.8 ± 3.8 h) was longer than TREAT-114 (21.9 ± 3.5 h, P = 0.002) and TREAT-115 (25.5 ± 3.7 h, P = 0.016) groups. However, the incidence of stillbirths in the TREAT-113, TREAT-114 and TREAT-115 groups was higher than in the CONTROL (16.4, 17.2, 11.8 and 5.8%, respectively, P < 0.05). In conclusion, altrenogest supplementation in combination with double administrations of PGF2α can reduce the variation in gestation length and synchronise the onset of parturition in sows. However, its side effects on the incidence of stillbirths should be considered.

Altrenogest treatment during the last week of lactation on ovarian traits and subsequent reproductive performance of primiparous and multiparous sows.

High-quality follicles result in larger corpora lutea (CL), producing more progesterone, and having a fundamental role in pregnancy maintenance. For some sows, follicular growth takes place during lactation, and follicle selection occurs under a catabolic environment. As altrenogest inhibits follicular development, this study aimed to evaluate follicular growth, CL size, estrus expression, and subsequent reproductive performance of sows treated with altrenogest during the last seven days of a three-week lactation. A total of 81 primiparous and 319 multiparous sows were allocated to two treatments: CONT (control group) and ALT (20 mg of altrenogest/day during the last seven days of lactation). Subsamples of 20 primiparous sows and 97 multiparous were randomly selected to evaluate follicular growth and 26 multiparous sows for serum progesterone analysis at day 21 of gestation. On day 21 of pregnancy, CL measurement was performed by ultrasound. Once in estrus, sows were post-cervically inseminated with pooled semen doses with 1.5 × 109 sperm cells at estrus onset and every 24 h during the standing reflex period. Sows not showing estrus until 10 days after weaning were considered in anestrus. The variables weaning-to-estrus interval, CL size, litter size in the subsequent cycle, and piglet birth weight were evaluated using the GLIMMIX procedure and compared using the Tukey-Kramer test. Anestrus, pregnancy, farrowing, and adjusted farrowing rate were evaluated as binary responses using logistic regression. Follicular size was analyzed as a repeated measure during treatment and after weaning. Treatment was considered as a fixed effect. During the treatment period, follicular size was smaller in ALT sows than CONT sows (3.29 vs. 3.52 mm; P < 0.001). However, after treatment, ALT sows showed a larger follicular size than CONT sows (5.30 vs. 5.03 mm; P ≤ 0.01). There were less ALT sows showing estrus than CONT sows on days three (1.03 vs. 4.57%) and four (55.38 vs. 68.02%) after weaning (P ≤ 0.05), respectively. At 21 days after insemination, ALT sows showed larger CL size and lower CL size variation (P < 0.01) than CONT sows. Anestrus rate, pregnancy rate, farrowing rate, adjusted farrowing rate, litter size in the subsequent cycle, piglet birth weight, litter birth weight, and birth weight variation did not differ between treatments (P ≥ 0.14). In conclusion, altrenogest treatment during the last week of lactation concentrated estrus expression on day five after weaning, larger follicle and CL sizes; however, with no improvement in reproductive performance.

Outcome of bilateral equid laparoscopic ovariectomies.

OBJECTIVE: To assess outcomes and behavior changes associated with bilateral laparoscopic ovariectomies. STUDY DESIGN: Retrospective study. SAMPLE POPULATION: Fifty-one equids. METHODS: Medical records were evaluated from equid bilateral laparoscopic ovariectomies from January 2012 to October 2018 with a potential of 6 months follow-up. Follow-up information obtained by telephone interviews included behavior before and after surgery. Likelihood ratio chi-square tests and odds ratios (OR) with 95% CI were calculated where applicable, with statistical significance at p < .05. RESULTS: Bilateral ovariectomy was performed in 51 cases, with elective (no pathologic ovaries) ovariectomies performed in 41/51 cases. Occasional estrus-like behavior was observed postoperatively in 14/51 (27%) mares, but the behavior was mild and manageable in all cases. There was no age effect on outcome in all bilateral (p = .56) or elective only (p = .36) cases. In 37/41 (90%) elective cases, improvement was observed in the reason for presentation. Some response to altrenogest administration for behavior modification was observed preoperatively in 12/18 (67%) elective cases. Response to altrenogest was not associated with (p = .31) or able to predict a beneficial response to surgery (OR = 5.5; 95% CI = 0.38-78.57; p = .21). CONCLUSION: Response to altrenogest in elective cases may not predict behavioral outcome with ovariectomy. Occasional estrus-like behavior in mares postoperatively was not problematic for any owners. Bilateral ovariectomy is a viable treatment option for owners seeking to alleviate undesirable behavior in mares. CLINICAL SIGNIFICANCE: This study should aid veterinarians and horse owners in case selection for bilateral ovariectomy.

Localization of kisspeptin, NKB, and NK3R in the hypothalamus of gilts treated with the progestin altrenogest.

Mechanisms in the brain controlling secretion of gonadotropin hormones in pigs, particularly luteinizing hormone (LH), are poorly understood. Kisspeptin is a potent LH stimulant that is essential for fertility in many species, including pigs. Neurokinin B (NKB) acting through neurokinin 3 receptor (NK3R) is involved in kisspeptin-stimulated LH release, but organization of NKB and NK3R within the porcine hypothalamus is unknown. Hypothalamic tissue from ovariectomized (OVX) gilts was used to determine the distribution of immunoreactive kisspeptin, NKB, and NK3R cells in the arcuate nucleus (ARC). Almost all kisspeptin neurons coexpressed NKB in the porcine ARC. Immunostaining for NK3R was distributed throughout the preoptic area (POA) and in several hypothalamic areas including the periventricular and retrochiasmatic areas but was not detected within the ARC. There was no colocalization of NK3R with gonadotropin-releasing hormone (GnRH), but NK3R-positive fibers in the POA were in close apposition to GnRH neurons. Treating OVX gilts with the progestin altrenogest decreased LH pulse frequency and reduced mean circulating concentrations of LH compared with OVX control gilts (P < 0.01), but the number of kisspeptin and NKB cells in the ARC did not differ between treatments. The neuroanatomical arrangement of kisspeptin, NKB, and NK3R within the porcine hypothalamus confirms they are positioned to stimulate GnRH and LH secretion in gilts, though differences with other species exist. Altrenogest suppression of LH secretion in the OVX gilt does not appear to involve decreased peptide expression of kisspeptin or NKB.

Downregulation of testicular function in the goat by altrenogest.

BACKGROUND: The present study investigated whether the administration of the progestin altrenogest provides noninvasive, temporary, and reversible suppression of gonadal function in the goat as a potential alternative to chirurgical castration, which is related with irreversibility, risks of complications till death of the animal and welfare issues. Eight sexually mature Peacock goats were randomly divided into two groups. The experimental group was administered altrenogest (0.088 mg/kg) orally once daily for 7 weeks. The remaining four goats received an oral glucose solution and served as the control group. After completing the administration period, the reversibility of the medication was evaluated for another 7 weeks (observation phase). The treatment effects were assessed by clinical examination; ultrasound examination of the testes, including one-dimensional grayscale analysis, blood testosterone levels, analysis of semen parameters and libido. At the end of the observation period, the animals were castrated and the testicles were examined histologically. RESULTS: Altrenogest treatment had no significant effect on the physical development of the goats, the sonographic appearance of the testes, the gray values measured in the ultrasound images, or the blood testosterone levels. The effects of treatment on the testicular and semen parameters varied widely in the experimental animals; the testicle volume was significantly lower and the number of pathologically altered sperm in the ejaculate was significantly higher in treated animals. CONCLUSION: These findings indicate that daily altrenogest administration at a dose of 0.088 mg/kg does not reliably suppress gonadal function in the goat.

Muscle fiber morphometry and physicochemical characteristics of the Longissimus thoracis muscle of hair male lambs fed zilpaterol hydrochloride and implanted with steroids.

Muscle fiber morphometry and physicochemical characteristics were evaluated in LT muscles obtained from entire male lambs treated with zilpaterol hydrochloride (ZH, 0 and 0.15 mg/kg body weight) and/or steroidal implant (SI, with and without trenbolone acetate/estradiol). ZH and SI acted synergistically to increase LT area, type-IIb fiber cross-sectional area and soluble collagen content, likewise to decrease metmyoglobin concentration and insoluble collagen content. Ash content and ultimate pH showed a decrease due to an antagonistic effect between ZH and SI. Content of total collagen, protein, fat, moisture, oxidized lipids and water-holding capacity were unaffected by ZH and SI. Supplemental ZH, but not SI, decreased all color parameters and tended to increase shear force. Overall, the SI implantation of male lambs followed by a ZH supplementation promoted greater LT hypertrophy, without affecting protein and fat content, and physicochemical characteristics in their meat.

Structurally different anabolic androgenic steroids reduce neurite outgrowth and neuronal viability in primary rat cortical cell cultures.

The illicit use of anabolic androgenic steroids (AAS) among adolescents and young adults is a major concern due to the unknown and unpredictable impact of AAS on the developing brain and the consequences of this on mental health, cognitive function and behaviour. The present study aimed to investigate the effects of supra-physiological doses of four structurally different AAS (testosterone, nandrolone, stanozolol and trenbolone) on neurite development and cell viability using an in vitro model of immature primary rat cortical cell cultures. A high-throughput screening image-based approach, measuring the neurite length and number of neurons, was used for the analysis of neurite outgrowth. In addition, cell viability and expression of the Tubb3 gene (encoding the protein beta-III tubulin) were investigated. Testosterone, nandrolone, and trenbolone elicited adverse effects on neurite outgrowth as deduced from an observed reduced neurite length per neuron. Trenbolone was the only AAS that reduced the cell viability as indicated by a decreased number of neurons and declined mitochondrial function. Moreover, trenbolone downregulated the Tubb3 mRNA expression. The adverse impact on neurite development was neither inhibited nor supressed by the selective androgen receptor (AR) antagonist, flutamide, suggesting that the observed effects result from another mechanism or mechanisms of action that are operating apart from AR activation. The results demonstrate a possible AAS-induced detrimental effect on neuronal development and regenerative functions. An impact on these events, that are essential mechanisms for maintaining normal brain function, could possibly contribute to behavioural alterations seen in AAS users.

Improving ovulation in gilts using anti-inhibin serum treatment combined with fixed-time artificial insemination.

For successful batch farrowing, porcine oestrus and ovulation must be synchronized using fixed-time artificial insemination (FTAI). However, exogenous gonadotropins, which are currently used in FTAI, negatively affect gilt ovulation. Here, we aimed to improve sexually mature gilt superovulation efficiency using passive immunization against inhibin during FTAI. Altrenogest-treated gilts were challenged with 10 ml anti-inhibin serum (AIS group, n = 6), 1,000 IU pregnant mare serum gonadotropin (PMSG group, n = 6), or 10 ml goat serum (control group, n = 6). Gilts in the AIS and PMSG groups were inseminated according to the FTAI protocol, and gilts in the control group were inseminated during natural oestrus. When PMSG was replaced by AIS during FTAI of gilts, ovulation rate and embryos recovered were significantly greater in the AIS group as compared to the other two groups (p < .05). Especially the average number of 6-8-cell embryos in the AIS group was significantly higher than that in the PMSG group (p < .01). Moreover, the blastocyst number in the AIS group was significantly higher than that in the PMSG group and the control group (p < .05). But there was no significant difference in the blastocyst number between the PMSG group and the control group (p > .05). Besides, plasma levels of estradiol-ß (E2) and progesterone (P4) were significantly greater in the AIS group as compared to the other two groups on Day 23 and D 27, respectively (p < .01). In summary, we devised an improved high-yield FTAI protocol for sexually mature gilts using AIS; this protocol had a greater superovulation efficiency than the FTAI using PMSG.

Altrenogest affects expression of galectin-3 and fibroblast growth factor 9 in the reproductive tract of sows.

A synthetic progestin altrenogest (ALT) is used to synchronize the estrus cycle of swine for fixed-time artificial insemination (AI) and has been shown to improve follicular development and reproductive performances in post-weaning sows. However, the effects of ALT treatment on reproductive tracts, including the ovaries, oviducts and uterus have not been yet clarified. In this study, we examined the expression of genes involved in endometrial responses in ALT-treated sows. ALT did not significantly alter luteinizing hormone (LH), follicle-stimulating hormone (FSH) and estradiol profiles in blood compared to untreated control. Quantitative RT-polymerase chain reaction (qRT-PCR) analysis showed that the expression of genes encoding galectin-3 (LGALS3) and fibroblast growth factor 9 (FGF9) was upregulated in the reproductive tracts of ALT-treated sows, including the ovaries, oviducts and uteri. Moreover, ALT treatment induced the expression of FGF9 and galectin-3 proteins, and promoted their localization to the luminal epithelium of the oviducts and uterus. Our findings suggest that the enhancement of reproductive performance shown by ALT-treated sows is associated with the upregulation of galectin-3 and FGF9, which are essential for endometrial receptivity, successful implantation, and pregnancy.

Unravelling androgens in sport: Altrenogest shows strong activation of the androgen receptor in a mammalian cell bioassay.

Altrenogest is a commonly used progestogen for the suppression of oestrus and associated distracting behaviours that interfere with training and performance of female racehorses. The steroid is derived from 19-nor testosterone and is structurally similar to the anabolic androgenic steroid, trenbolone. In this study, the relative androgen potency of altrenogest was determined by a kidney (HEK293) cell androgen bioassay. The HEK293 bioassay shows that in its pure form, altrenogest has a high relative potency compared with testosterone but is not as strong as ß-trenbolone. Our results also show that altrenogest is able to activate the androgen receptor at the concentrations relevant to the administration regime of racehorses and retains its activity ex vivo. Thus, we show unequivocally that altrenogest, a progestogen used widely in female racehorses, acts as a strong androgen in a mammalian cell bioassay.

Understanding the influence of trenbolone acetate and polyamines on proliferation of bovine satellite cells.

Approximately 90% of beef cattle on feed in the United States receive at least one anabolic implant, which results in increased growth, efficiency, and economic return to producers. However, the complete molecular mechanism through which anabolic implants function to improve skeletal muscle growth remains unknown. This study had 2 objectives: (1) determine the effect of polyamines and their precursors on proliferation rate in bovine satellite cells (BSC); and (2) understand whether trenbolone acetate (TBA), a testosterone analog, has an impact on the polyamine biosynthetic pathway. To address these, BSC were isolated from 3 finished steers and cultured. Once cultures reached 75% confluency, they were treated in 1% fetal bovine serum (FBS) and/or 10 nM TBA, 10 mM methionine (Met), 8 mM ornithine (Orn), 2 mM putrescine (Put), 1.5 mM spermidine (Spd), or 0.5 mM spermine (Spe). Initially, a range of physiologically relevant concentrations of Met, Orn, Put, Spd, and Spe were tested to determine experimental doses to implement the aforementioned experiments. One, 12, or 24 h after treatment, mRNA was isolated from cultures and abundance of paired box transcription factor 7 (Pax7), Sprouty 1 (Spry), mitogen-activated protein kinase-1 (Mapk), ornithine decarboxylase (Odc), and S adenosylmethionine (Amd1) were determined, and normalized to 18S. No treatment × time interactions were observed (P ≥ 0.05). Treatment with TBA, Met, Orn, Put, Spd, or Spe increased (P ≤ 0.05) BSC proliferation when compared with control cultures. Treatment of cultures with Orn or Met increased (P ≤ 0.01) expression of Odc 1 h after treatment when compared with control cultures. Abundance of Amd1 was increased (P < 0.01) 1 h after treatment in cultures treated with Spd or Spe when compared with 1% FBS controls. Cultures treated with TBA had increased (P < 0.01) abundance of Spry mRNA 12 h after treatment, as well as increased mRNA abundance of Mapk (P < 0.01) 12 h and 24 h after treatment when compared with 1% FBS control cultures. Treatment with Met increased (P < 0.01) mRNA abundance of Pax7 1 h after treatment as compared with 1% FBS controls. These results indicate that treatments of BSC cultures with polyamines and their precursors increase BSC proliferation rate, as well as abundance of mRNA involved in cell proliferation. In addition, treatment of BSC cultures with TBA, polyamines, or polyamine precursors impacts expression of genes related to the polyamine biosynthetic pathway and proliferation.

Altrenogest affects the development and endocrine milieu of ovarian follicles in prepubertal and mature gilts†.

Altrenogest with gonadotropins is commonly used to synchronize the estrous cycle, but it can also lead to follicular cyst formation, especially in prepubertal gilts. Here, we aimed to investigate how maturity and altrenogest treatment affect the development, endocrine milieu, and molecular control of ovarian follicles. Crossbred prepubertal and mature gilts were challenged or not (control) with altrenogest, and ovaries were collected in the morning on the first day of behavioral estrus. In prepubertal gilts, altrenogest decreased the percentage of primordial and atretic small follicles, but increased large antral follicles when compared with controls. In mature gilts, altrenogest reduced the percentage of primary follicles and elevated the total number of antral follicles. Maturity affected the estradiol level in the follicular fluid of preovulatory follicles, luteinizing hormone (LH)-stimulated cyclic adenosine monophosphate (cAMP) generation, and LH receptor messenger RNA (mRNA) expression in granulosa. Moreover, cytochrome P45017A1 (CYP17A1) mRNA levels in the theca layer were affected and correlated with follicular androstendione and estradiol concentration. Altrenogest negatively affected follicular fluid progesterone concentration and decreased levels of prostaglandin (PG) E2 in prepubertal gilts and PGF2alpha metabolite in mature gilts. LH-stimulated cAMP release in granulosa cells of mature gilts as well as human chorionic gonadotropin- and forskolin-induced cAMP were also affected. In addition, altrenogest downregulated CYP17A1 mRNA in the prepubertal theca layer and PGF2alpha synthase expression in the granulosa and theca layer of mature gilts. To the best of our knowledge, this is the first study to report multiple effects of maturity and altrenogest on the endocrine milieu and molecular regulations governing ovarian follicle development in gilts.

Effects of oestrous synchronization with altrenogest in gilts on endometrial and embryonic characteristics.

The use of altrenogest (ALT) supplementation for oestrous synchronization improves subsequent reproductive performance of gilts and sows. However, the causes of this improvement in reproductive performance after ALT treatment are not fully/clearly understood. The objective of this study was to evaluate the effects of ALT supplementation for oestrous synchronization in gilts on the endometrial glands and embryonic development characteristics at 28 days of pregnancy. Pregnant gilts were divided into two experimental treatments: Control (did not receive ALT; n = 9 gilts) and ALT (ALT feeding at 20 mg/day for 18 days; n = 9 gilts). At 28 days of pregnancy, six gilts from each treatment were slaughtered, and reproductive tracts were immediately evaluated. There was no statistical difference (P > 0.05) between treatments regarding ovulation rate, number of embryos, number of vital embryos and number of non-vital embryos. Embryo weight, length and embryonic vesicle weight were lower in ALT treatment compared with Control (P < 0.01), and it was lower in the cervical uterine region compared with apex uterine region, respectively (P < 0.05). Higher values of gland duct area, gland duct perimeter, percentage of the glandular area and total endometrial area were observed in ALT treatment compared with Control (P < 0.05). The use of ALT during 18 days for oestrous synchronization in gilts increased the gland duct area, perimeter and total endometrial area but did not increase the embryo number and embryo size at day 28 of pregnancy.

Supplemental progesterone during early pregnancy exerts divergent responses on embryonic characteristics in sows and gilts.

Progesterone (P4) plays a key role in pregnancy establishment and maintenance; during early pregnancy, P4 stimulates the production and release of uterine secretions necessary for conceptus growth prior to implantation; therefore, exogenous P4 supplementation may improve embryo development. This study evaluated the effects of supplementation during early pregnancy with long-acting injectable progesterone or altrenogest on embryonic characteristics of sows and gilts. Thus, a total of 32 sows and 16 gilts were used. On day 6 of pregnancy sows and gilts were allocated to one of the following groups: non-supplemented; supplemented with 20 mg of altrenogest, orally, from days 6 to 12 of pregnancy; supplemented with 2.15 mg/kg of long-acting injectable progesterone on day 6 of pregnancy. Animals were killed on day 28 of pregnancy, and ovulation rate, embryo survival, embryo weight, crown-to-rump length, uterine glandular epithelium and endometrial vascularization were assessed. Treatments had no effect on pregnancy rate, embryo survival or endometrial vascular density (P > 0.05). Non-supplemented gilts presented larger and heavier embryos compared to gilts from supplemented groups (P < 0.05). Sows in the altrenogest group presented larger and heavier embryos compared to non-supplemented sows and sows supplemented with long-acting injectable progesterone. In conclusion, supplementation of sows and gilts with progestagen from day 6 of pregnancy can be used as a means to improve embryo survival without deleterious effects.

Effects of 17ß-trenbolone exposure on sex hormone synthesis and social behaviours in adolescent mice.

17ß-Trenbolone (17ß-TBOH) is an endocrine disruptor that has been widely reported in aquatic organisms. However, little is known about the effect of 17ß-TBOH on mammals, particularly on the development of adolescents. Through a series of behavioural experiments, exposure to at 80 µg kg -1 d -1 and 800 µg kg -1 d -1 17ß-TBOH during puberty (from PND 28 to 56, male mice) increased anxiety-like behaviours. Exposure to the low dose of 80 µg kg -1 d -1 resulted in a clear social avoidance behaviour in mice. The two doses affected testicular development and endogenous androgen synthesis in male mice. In addition, 17ß-TBOH exposure altered the differentiation of oligodendrocytes and the formation of the myelin sheath in the medial prefrontal cortex (mPFC). These results reveal the effects of 17ß-TBOH on the behaviours, gonadal and neurodevelopment of adolescent mammals. In addition, the inhibition of the secretion of endogenous hormones and decrease in the formation of the myelin sheath in mPFC may be associated with the 17ß-TBOH-induced behavioural changes in mice.