The mycoestrogen zeranol at high dosage antagonizes transient receptor potential channel activities in 3T3 L1 cells.

Recent findings have revealed that exposure to environmental contaminants may result in obesity and pose a health threat to the general public. As the activity of transient receptor potential channels (TRPs) plays a permissive role in adipogenesis, the interactions between TRPs and some food pollutants, i.e. bisphenol A, di (2-ethylhexyl) phthalate, zearalenone, and zeranol at 10 µM were investigated in the present study. TRP-V1,-V3, -C4 and -C6 are reported to be differentially expressed in the adipocyte differentiation, and immunoblotting was performed to quantify changes in these TRPs affected by the pollutants. Our result indicated that the mycoestrogen zeranol or α-zearalanol suppressed the expression of the V1 and C6 isoforms. Subsequently, confocal microscopy was used to measure the calcium inflow repressed by zeranol from 0.1 µM to 10 µM. Oil Red O staining was used to determine the differentiation of 3T3 L1 preadipocytes. Zeranol could suppress the expression of TRP-V1 and -C6 protein and inhibit the associated flow of calcium into the cytosol of 3T3 L1 cells. Its IC50 value for inhibiting calcium inflow stimulated by 40 µM capsaicin or 10 µM GSK1702934A was estimated to be around 6 µM. Reduced TRP-V1 or -C6 activity might result in promoting adipogenesis. In conclusion, this study demonstrated that zeranol could potentiate fat cell differentiation through antagonizing TRP-V1 and -C6 activities.

Synergistic estrogenic effects of Fusarium and Alternaria mycotoxins in vitro.

Mycotoxins are toxic secondary metabolites formed by various fungal species that are found as natural contaminants in food. This very heterogeneous group of compounds triggers multiple toxic mechanisms, including endocrine disruptive potential. Current risk assessment of mycotoxins, as for most chemical substances, is based on the effects of single compounds. However, concern on a potential enhancement of risks by interactions of single substances in naturally occurring mixtures has greatly increased recently. In this study, the combinatory effects of three mycoestrogens were investigated in detail. This includes the endocrine disruptors zearalenone (ZEN) and α-zearalenol (α-ZEL) produced by Fusarium fungi and alternariol (AOH), a cytotoxic and estrogenic mycotoxin formed by Alternaria species. For evaluation of effects, estrogen-dependent activation of alkaline phosphatase (AlP) and cell proliferation were tested in the adenocarcinoma cell line Ishikawa. The estrogenic potential varied among the single substances. Half maximum effect concentrations (EC50) for AlP activation were evaluated for α-ZEL, ZEN and AOH as 37 pM, 562 pM and 995 nM, respectively. All three mycotoxins were found to act as partial agonists. The majority of binary combinations, even at very low concentrations in the case of α-ZEL, showed strong synergism in the AlP assay. These potentiating phenomena of mycotoxin mixtures highlight the urgent need to incorporate combinatory effects into future risk assessment, especially when endocrine disruptors are involved. To the best of our knowledge, this study presents the first investigation on synergistic effects of mycoestrogens.

Cytotoxicity induced by ochratoxin A, zearalenone, and α-zearalenol: effects of individual and combined treatment.

This study investigated the cytotoxicity of combined mycotoxins of ochratoxin A (OTA), zearalenone (ZEA), and/or α-zearalenol (α-ZOL). The cytotoxicity of two mycotoxin combinations (two two-toxin combinations and one three-toxin combination) on human Hep G2 cells was evaluated using a tetrazolium salt (MTT) assay and isobologram analysis. Our results demonstrated significant cytotoxic effects of the two-toxin combination and the three-toxin combination on Hep G2 cells in a time- and concentration-dependent manner. The combination indexes (CI) were 2.73-7.67 for the OTA+ZEA combination and 1.23-17.82 for the OTA+α-ZOL combination after 24 h, 48 h, and 72 h of exposure at all inhibit concentration (IC) levels (IC10-IC90), indicating an antagonism. The CIs of the ZEA+α-ZOL combination were 1.29-2.55 after 24 h and 72 h of exposure (IC10-IC90), indicating an antagonism. The CIs of the ZEA+α-ZOL combination were 0.74-1.68 after 48 h of exposure, indicating synergism (IC80-IC90), additive effects (IC50-IC70), or antagonism (IC10-IC40). For the OTA+ZEA+α-ZOL combination, the CIs were 1.41-14.65 after 24 h, 48 h, and 72 h of exposure (IC10-IC90), indicating an antagonism.

Alleviation of plasma homocysteine level by phytoestrogen α-zearalanol might be related to the reduction of cystathionine ß-synthase nitration.

Hyperhomocysteinemia is strongly associated with cardiovascular diseases. Previous studies have shown that phytoestrogen α-zearalanol can protect cardiovascular system from hyperhomocysteinemia and ameliorate the level of plasma total homocysteine; however, the underlying mechanisms remain to be clarified. The aim of this research is to investigate the possible molecular mechanisms involved in ameliorating the level of plasma homocysteine by α-zearalanol. By the successfully established diet-induced hyperhomocysteinemia rat models, we found that, after α-zearalanol treatment, the activity of cystathionine ß-synthase, the key enzyme in homocysteine metabolism, was significantly elevated and level of nitrative stress in liver was significantly reduced. In correlation with this, results also showed a decreased nitration level of cystathionine ß-synthase in liver. Together data implied that alleviation of plasma homocysteine level by phytoestrogen α-zearalanol might be related to the reduction of cystathionine ß-synthase nitration.

Interactive effects of zearalenone and its metabolites on cytotoxicity and metabolization in ovarian CHO-K1 cells.

Zearalenone (ZEA) is a non-steroidal estrogen mycotoxin with high binding affinity to estrogen receptors. ZEA is rapidly absorbed and metabolized in vivo to α-zearalenol (α-ZOL) and ß-zearalenol (ß-ZOL). So, mixtures of them may be present in biological systems and suppose a hazard to animals and human health. The aims of this study were to determine the cytotoxic effects of ZEA and its metabolites, alone and in combination in ovarian (CHO-K1) cells during 24, 48 and 72h by the MTT assay; and to investigate the metabolism of the CHO-K1 cells on ZEA, and its conversion into α-ZOL and ß-ZOL by CHO-K1 cell after 24 and 48h of exposure. The IC50 value obtained for individual mycotoxins range from 60.3 to >100.0µM, from 30.0 to 33.0µM and from 55.0 to >75.0µM for ZEA, α-ZOL and ß-ZOL, respectively. Cytotoxic interactions were assayed by the isobologram method, which provides a combination index (CI) value as a quantitative measure of the degree of the three mycotoxin interaction. The CI values for binary combinations ranged from 0.56±0.15 (synergism at low concentrations) to 5.25±5.10 (addition at high concentrations) and tertiary combinations from 2.95±0.75 (antagonism at low concentrations) to 0.41±0.23 (synergism at high concentrations). The concentration of ZEA and its metabolites was determined with liquid chromatography coupled to the mass spectrometer detector-linear ion trap (LC-MS-LIT). The percentage of ZEA degradation ranged from 4% (24h) to 81% (48h). In the same conditions, α-ZOL and ß-ZOL concentration decreased from 8% to 85%. No conversion of ZEA in α-ZOL and ß-ZOL was detected. However, at 24h of exposure other degradation products of ZEA and its derived were detected.

Zeranol enhances the proliferation of pre-adipocytes in beef heifers.

BACKGROUND: The high morbidity and mortality of breast cancer among women is a serious problem. The adverse effects of the consumption of beef with zeranol (Z, a growth promoter widely used in beef industry in North American) residue on human health are still unknown. MATERIALS AND METHODS: The effects of Z implantation on the growth of heifer pre-adipocytes were evaluated. The stimulatory effects of Z and estradiol-17beta (E(2)) on the proliferation of pre-adipocytes isolated from control heifers and Z-implanted heifers were measured. Real-time PCR and Western-blotting analysis were performed to evaluate the expression of cyclin D1 and p53 at both mRNA and protein levels. RESULTS: The growth of pre-adipocytes from heifers bearing for 2 months of Z-implants was about 12-fold faster than that observed in control heifers. The pre-adipocytes isolated from Z-implanted heifers were more sensitive to treatment with Z and E(2). Z up-regulated the expression of cyclin D1 and down-regulated p53 in pre-adipocytes isolated from Z-implanted heifers. CONCLUSION: The implantation of Z increases body weight gain by enhancing growth of pre-adipocytes. The stimulation of pre-adipocytes division by Z and E(2) might be partially mediated by up-regulation of cyclin D1 and down-regulation of p53 at mRNA and protein levels.

Effects of Zeranol upon luteal maintenance and fetal development in peripubertal gilts.

Eighty gilts were utilized to determine whether zeranol implants could maintain hCG-induced corpora lutea (CL) in peripubertal gilts and to examine the effects of a Zeranol implant on fetal development. Crossbred gilts (171+/-0.3 days of age, 109.1+1.4 kg) were blocked by weight and ancestry to control (n=40) or treatment (n=40) groups. To induce ovulation and CL maintenance, treated gilts received 500 IU of hCG i.m. and a Zeranol ear implant (Ralgro, 36 mg; day 0). All gilts were checked once daily for estrus with a mature boar from days 3-58 of the experiment. On day 42, treated gilts received two 10 mg injections of Lutalyse (PGF(2)alpha) spaced 6 h apart. Treated gilts not displaying estrus within 7 days of PGF(2)alpha received two additional 10 mg of PGF(2)alpha spaced 6 h apart on day 49. On days 44-58, gilts detected in estrus were inseminated twice, 24 h apart with pooled semen via AI. Blood samples were obtained on days 0, 7, 18 and 42 and analyzed for serum progesterone (P(4)). Bred gilts were slaughtered on days 58-62 of gestation. Ovulation, as determined by serum concentrations of P(4) on day 7 of the experiment, was induced by hCG in 79.5% of treated gilts. Zeranol implants, however, failed to increase (P>0.05) the proportion of gilts available for breeding (treated, 21/39; control, 18/40). Of gilts inseminated on days 44-58, 16/21 treated gilts and 16/18 control gilts were pregnant at slaughter on days 58-62 of gestation. Number of fetuses (7.5 versus 12), fetal weight (83 versus 121 g), fetal length (117 versus 132 mm) and fetal survival (45% versus 78%) were reduced (P<0.001) by Zeranol implants. These data indicate that treatment of peripubertal gilts with a 36 mg Zeranol implant did not increase the proportion of gilts available for breeding while causing deleterious effects upon the fetuses.

Assessment of proliferative activity by AgNOR and PCNA in prostatic tissues of ram lambs implanted with zeranol.

The aim of this study was to assess cellular proliferation using silver-stained nucleolar organizer regions (AgNOR) and the proliferating cell nuclear antigen (PCNA) in various tissues in the prostate of ram lambs implanted with increasing zeranol doses and to compare the sensitivity of different tissues of lamb prostate to zeranol. Twenty-four Akkaraman lambs were implanted with increasing zeranol doses, including 12 mg (n = 8), 24 mg (n = 8) and 96 mg (n = 8), with eight lambs serving as controls. After 33 days, the prostate tissues of the lambs were stained using AgNOR and PCNA techniques. The prostate tissues were divided into two compartments--the epithelial tissues, including glandular acinus, collecting duct and penile urethra, and the non-epithelial tissues, including interstitial tissue and striated muscle. AgNOR dots and PCNA index on each prostatic tissue were counted under a light microscope and were evaluated statistically. AgNOR staining in the treatment groups showed a higher score in the non-epithelial tissues than the epithelial components, whereas the PCNA index was significant in the epithelial tissues and non-epithelial tissues had very low PCNA immunostaining. According to the PCNA index, collecting duct epithelium showed more sensitivity to increasing zeranol doses and according to AgNOR counts, there was no difference of sensitivity to zeranol among tissues of the same origin. Both AgNOR counts and PCNA indexes seem to be valuable proliferating markers for the epithelial components of ram prostate, but PCNA index had no significance in relation to the non-epithelial components in contrast to AgNOR counts. Therefore, the controversial results arising from the combined use of both techniques as proliferating markers for the ram prostate should be considered in further studies.

Transformation of MCF-10A human breast epithelial cells by zeranol and estradiol-17beta.

Among the endocrine factors associated with breast cancer, estrogens are considered to play a central role in human breast carcinogenesis. Breast cancer risks are increased by long-term exposure to estrogens. Zeranol (Ralgro) is a nonsteroidal agent with estrogenic activity that is used as a growth promoter in the U.S. beef and veal industry. To determine whether zeranol and estradiol-17beta play a role in the neoplastic transformation of human breast and to compare the estrogenic potency of zeranol to that of estradiol-17beta in human breast, we treated human breast epithelial cell MCF-10A with different doses of zeranol or estradiol-17beta for 10 repeated treatment cycles. By utilizing the doubling time assay, soft agar assay, and reverse transcriptase polymerase chain reaction (RT-PCR) assay, we showed that 10 repeated estradiol-17beta or zeranol treatment cycles to MCF-10A cells decrease the doubling time of the cells by 30 to 40% and stimulate colony formation in soft agar and induce estrogen receptor beta (ER-beta) mRNA expression, all of which are not dose related in our tested dose range. Furthermore, we show that zeranol and estradiol-17beta have a similar potency in the stimulation and inhibition of gene expressions in human breast cancer cell line MCF-7 by RT-PCR. These results indicate that both zeranol and estradiol-17beta can induce human breast epithelial cell neoplastic transformation with similar potency in the long-term exposure through the oxidation-reduction (redox) pathway and/or ER-beta-mediated pathway.

Effects of implant regimens (trenbolone acetate-estradiol administered alone or in combination with zeranol) and vitamin D3 on fresh beef color and quality.

In the first oftwo experiments, 123 calf-fed steers were used over a 2-yr period to evaluate the effects of trenbolone acetate (TBA)-based implants administered alone or in combination with zeranol implants on fresh beef muscle quality, color, and physiological maturity of the carcass. Implant treatments decreased (P < 0.05) a* values (d 0 and d 3 of retail display) and b* values (d 0, d 1, and d 3 of retail display) after 14 d of aging. Carcasses from cattle initially implanted with Revalor-S and reimplanted with Revalor-S on d 60 of the finishing period showed increased lean and bone maturity scores and ash content of the 9th to 11th thoracic buttons and Warner-Bratzler shear force values (WBS) compared to those initially implanted with Ralgro and subsequently reimplanted with Revalor-S or control cattle. In addition, implants decreased (P < 0.05) marbling, percentage of the carcasses grading Choice, and kidney, pelvic, and heart fat (KPH). Implant treatments increased (P < 0.05) ADG, hot carcass weights, and longissimus muscle (LM) area. In the second experiment over a 2-yr period, 166 steers fed as yearlings were allotted to one of two implant treatments and one of two vitamin D3 preharvest supplementation treatments. Implanted steers had heavier (P < 0.05) final body weights and higher (P < 0.05) ADG, less (P < 0.05) KPH fat, and larger (P < 0.05) LM. Also, implanted steers had more (P < 0.05) advanced bone maturity scores, higher (P < 0.05) ash content of the 9th to 11th thoracic buttons, and higher (P < 0.05) WBS values on 5-d postmortem loin steaks. Vitamin D3 feeding decreased (P < 0.05) final live weight, ADG (P < 0.05), and LM (P < 0.05), but did not significantly improve WBS values. In Experiment 2, neither implant treatment nor vitamin D3 supplementation had significant effects on L*, a*, or b* values of muscles in steaks before or during simulated retail display.

Impact of doramectin treatment at the time of feedlot entry on the productivity of yearling steers with natural nematode infections.

OBJECTIVE: To measure the reduction in fecal nematode egg counts and productivity impact of treatment of yearling steers with doramectin at entry into the feedlot, compared with control steers treated only with fenthion. ANIMALS: 6,096 crossbred yearling steers with a mean (+/- SD) body weight of 377.0 (+/- 37) kg. PROCEDURE: Steers were implanted with zeranol and alternately separated to fill each of 24 pens. Groups of steers within 12 matched pairs of pens were randomly allocated to treatment with doramectin or no treatment with doramectin for internal nematodes. Fecal samples were collected from approximately every twentieth steer from each pen at day 0 and at reimplant (approx day 60). Each steer was weighed on day 0 and at reimplant and then mean body weights of steers per pen were determined at 120 to 140 days after trial initiation. RESULTS: Treatment steers had a significantly lower fecal egg count at reimplant than control steers. Treatment steers had a significantly greater mean daily gain during the study, significantly greater feed consumption, significantly lower feed-to-gain ratio, and significantly better quality carcass grades at slaughter. CONCLUSIONS AND CLINICAL RELEVANCE: Under the conditions of our trial, there was a significant fecal egg count reduction response to doramectin treatment, which resulted in significantly improved productivity. Results of economic analysis of return on investment indicated that even with low egg counts in heavy body weight cattle, nematode egg count reduction with doramectin significantly improved returns.

Hormone contents in peripheral tissues after correct and off-label use of growth promoting hormones in cattle: effect of the implant preparations Filaplix-H, Raglo, Synovex-H and Synovex Plus.

Certain hormonal growth promoters are licensed in several beef producing countries outside the European Union (EU). Use in compliance with Good Veterinary Practice is mandatory. As risk assessment of hormone residues in animal tissues up to now has neglected potential off-label use, the present study dealt with two topics: 1) multiple treatment with the implant preparations Finaplix-H (200 mg trenbolone acetate), Ralgro (36 mg zeranol) and Synovex-H (200 mg testosterone propionate plus 20 mg estradiol benzoate) in heifers (1-fold, 3-fold and 10-fold dose), and 2) non-approved treatment of female veal calves (1-fold dose of Synovex-H or Synovex Plus with 200 mg trenbolone acetate plus 28 mg estradiol benzoate). Residues of estradiol-17beta, estradiol-17alpha, estrone and testosterone, trenbolone-17beta, trenbolone-17alpha and trendione or zeranol, respectively, were measured in loin, liver, kidney and peri-renal fat by high performance liquid chromatography/enzyme immunoassay (HPLC/EIA) after liquid-liquid extraction and solid-phase clean-up. The hormone residues in the multiple-dose experiments were dose-dependent and partially exceeded the threshold values: in the liver in one animal after 3-fold dose and in two animals after 10-fold dose of Finaplix-H, and in the liver and kidney after 3-fold and 10-fold dose of Synovex-H. Mean hormone residues in calves were mainly below those of heifers and did not infringe threshold values.

Quantification of rainbow trout (Oncorhynchus mykiss) zona radiata and vitellogenin mRNA levels using real-time PCR after in vivo treatment with estradiol-17 beta or alpha-zearalenol.

Estrogen receptor-mediated induction of zona radiata (ZR) and vitellogenin (VTG) mRNA and protein in rainbow trout (Oncorhynchus mykiss) was compared to assess their utility as biomarkers for exposure to estrogenic compounds. Partial sequences of rainbow trout ZR and beta-actin were cloned by reverse transcriptase polymerase chain reaction (RT-PCR) using degenerate primers based on conserved regions across a number of species. A 549 bp fragment of the rainbow trout ZR-gene showed a high degree of amino acid sequence identity to that of salmon (77%), winter flounder (64%), carp ZP2 (63%) and medaka (61%) ZR-proteins. The 1020 bp beta-actin fragment was approximately 100% identical to sequences from several species. Real-time PCR was used to quantify the induction of ZR-gene and VTG in rainbow trout liver after in vivo exposure to estradiol-17 beta (E(2)) (0.01, 0.1, 1.0 or 10 mg/kg body weight (bw) fish) or alpha-zearalenol (alpha-ZEA) (0.1, 1.0 or 10 mg/kg bw). Real-time PCR and indirect enzyme-linked immunosorbent assay (ELISA) showed that ZR and VTG were induced in both the liver and the plasma after a single injection of E(2) or alpha-ZEA. ZR was more responsive to low levels of E(2) and alpha-ZEA than VTG, and real-time PCR was shown to be more sensitive than the ELISA. Rainbow trout ZR-gene and proteins provide a sensitive biomarker for assessing estrogenic activity.

Efecto del zeranol sobre la maduración de piel en chinchilla lanígera (Eryomis laniger) / Zeranol effect over chinchilla's (Eryomis laniger) fur maturation

Con el objeto de evaluar el efecto de la lactona del ácido resorcílico (zeranol) sobre la maduración de la piel de chinchilla, se realizó un experimento en un diseño completamente al azar por grupo de edad y sexo. El estudio duró 4 meses en la época de verano-otoño (junio-septiembre). Se incluyeron 33 hembras vírgenes de 12 a 24 meses de edad, 60 machos de 12 a 18 meses; y 29 hembras en producción de 24 a 48 meses de edad; a la mitad de los animales se les implantó subcutáneamente 12 mg de zeranol. La evaluación visual se hizo cada 2 semanas y se obtuvieron las pieles cuando las 3 franjas del color del pelo presentaban el mismo nivel en toda la superficie del cuerpo, momento en el cual se considera como piel madura. Se realizó un análisis de sobrevida mediante el método de Log Rank ajustado por grupos, en el cual se encontró un efecto anagénico significativo (P< 0.01) del zeranol. El porcentaje de maduración en los animales implantados fue 84 por ciento, 94 por ciento y 50 por ciento en los machos, hembras jóvenes y hembras adultas, respectivamente; mientras que para los no tratados la maduración fue de 20 por ciento, 38 por ciento y 13.3 por ciento, para los mismos grupos. Se hicieron dos análisis de Log Rank ajustado por grupos de tratamiento, los cuales mostraron un efecto de edad y sexo; en ambos casos las hembras jóvenes presentaron un mayor porcentaje de maduración que las hembras adultas y que los machos (P< 0.01)

Implant sequence effects in intact male Holstein veal calves: live and slaughter traits.

Seven sequences of growth promotant implants were used in special-fed intact male Holstein veal calves (n = 443). Calves received implants 4 d after arrival at the veal barn, 42, and 84. The following implants were used: placebo (0), Z (36 mg zeranol), ET (20 mg estradiol, 200 mg testosterone), EP/2 (10 mg estradiol, 100 mg progesterone), EP (20 mg estradiol, 200 mg progesterone), and EBA (24 mg estradiol, 120 mg trenbolone acetate). The following sequences were compared: 0-0-0 (negative control), 0-ET-ET, Z-ET-ET, 0-EP-EP, Z-EP-EP, 0-EP/2-EBA, and Z-0-EBA. From 0 to 42 d, Z implants increased (P<.05) ADG by 3.4% compared to placebo. However, implant schemes without an initial Z implant (0-ET-ET and 0-EP-EP) had higher (P<.05) mean ADG for the period from d 42 to 84. From 84 d to the end of the experiment, only the 0-EP/2-EBA treatment increased (P<.05) ADG compared to 0-0-0. Over the entire trial 0-ET-ET, 0-EP-EP, Z-EP-EP, and 0-EP/2-EBA implant sequences increased (P<.05) ADG by 3.2, 3.2, 2.4, and 4.7%, respectively, compared to the 0-0-0 sequence. Blood traits measured within 2 wk before slaughter were not affected by implant sequence, except that sequences with EP had higher (P<.05) leukocyte counts than were observed for the other sequences. Testicular weight was less (P<.01) for all of the implant sequences than for the negative control and less (P<.05) for Z-ET-ET than for 0-ET-ET, 0-EP-EP, 0-EP/2-EBA, and Z-0-EBA. The type and frequency of medical treatments did not differ among implant sequences for any of the 42-d phases, or over the entire trial. Generally, the growth promotant implants currently approved for beef cattle resulted in approximately 50% of the increase in growth rate in Holstein intact bull calves, as has been observed in beef-type steers or heifers.

Implant sequence effects in intact male Holstein veal calves: carcass characteristics.

Seven sequences of growth promotant implants were used in intact male Holstein veal calves (n = 443). Implants were administered on d 0 (within 4 d after arrival at the veal barn), 42, and 84. The implants used were placebo (0), Z (36 mg zeranol), ET (20 mg estradiol, 200 mg testosterone), EP/2 (10 mg estradiol, 100 mg progesterone), EP (20 mg estradiol, 200 mg progesterone), and EBA (24 mg estradiol, 120 mg trenbolone acetate). The following sequences were compared: 0-0-0 (negative control), 0-ET-ET, Z-ET-ET, 0-EP-EP, Z-EP-EP, 0-EP/2-EBA, and Z-0-EBA. Sequences 0-EP-EP, Z-EP-EP, and 0-EP/2-EBA increased (P<.05) carcass weight from 3.3 to 3.9% compared to nonimplanted controls. There were no differences (P>.05) in percentage of carcass weight accounted for by the fore vs. rear halves of carcasses, suggesting there was no difference in the distribution of weight. Although there were differences in longissimus area, the results were not consistent, except that there was a trend for longissimus area to be increased by the use of estrogenic-androgenic implants (ET and EBA). There were no differences among implant sequences for carcass conformation, fat cover, muscle texture, marbling/ feathering, muscle color, or muscle chemical composition. Of four implant sequences (0-0-0, 0-ET-ET, 0-EP-EP, and 0-EP/2-EBA) tested for differences in Warner-Bratzler shear force tenderness, the latter two sequences averaged higher (P<.05) for shear force than did the negative control. These results suggest that aggressive implant strategies in young, intact Holstein bull calves (raised as veal) have minimal effects on carcass characteristics.

Effects of in utero exposure to nonsteroidal estrogens on mouse testis.

Male mice exposed in utero to alpha-zearalanol (zeranol) or diethylstilbestrol (DES) were analyzed postnatally to evaluate the possible changes on their testicular morphology as part of an examination of the effects of transplacental exposure to non-steroidal estrogens on sensitive tissues. Pregnant NMRI mice were injected subcutaneously with ethyl oleate (0.1 mL) alone (negative control) or with 150 micrograms/kg of body weight of zeranol or DES (positive control) on days 9 and 10 of gestation. Experimental and control male offspring were euthanized at days 45 (n = 47), 90 (n = 44), 180 (n = 40) and 365 (n = 26) after birth and their gonads were examined by light and electron microscopy. The results suggested that prenatal zeranol or DES exposure induced more severe and earlier (at 45 d) testicular abnormalities than in negative control (at 6 mo). These age-related alterations were characterized by regressive changes in the germinal epithelium and Sertoli's cells as well as foci of Leydig's cells around atrophied seminiferous tubules and dysplasia of the rete testis epithelium. On the contrary, the presence of Leydig's cells with immature morphology and their arrangement in sheet could be attributable exclusively to estrogen treatment. The presence of no neoplasm was confirmed.

Use of melengestrol acetate-based treatments to induce and synchronize estrus in seasonally anestrous ewes.

Ten experiments utilizing 1,659 ewes were conducted to determine the potential of melengestrol acetate (MGA)-based treatments in inducing and synchronizing estrous activity in ewes during the nonbreeding season. In May or June, ewes were given .25 mg of MGA.ewe-1.d-1 or control diet and(or) a subsequent i.m. injection of zeranol, estradiol-17 beta, or oil vehicle. Fertile rams were introduced, and estrous and(or) lambing responses were determined. In Exp. 1, more (P < .01) ewes given MGA for 14 d followed by 5 mg of zeranol (i.m.) were in estrus than controls. Melengestrol acetate and zeranol independently induced estrus in Exp. 2, with MGA-treated ewes exhibiting the greatest lambing response (P < .05). The optimum duration of feeding MGA was 8 d vs 11 or 14 d (Exp. 3). In Exp. 4, as the dose of zeranol increased, synchrony of estrus increased but lambing response decreased. A dose of 1.25 mg of zeranol represented a compromise in estrous and lambing response and was determined to be optimally effective when given 54 h after the last feeding of MGA (Exp. 5). Shortcomings in lambing responses were treated in Exp. 6 by supplementing ewes with MGA after breeding, which proved unsuccessful, and in Exp. 7 by substituting estradiol-17 beta for zeranol, which proved effective when given 54 h after the last feeding of MGA (Exp. 8). In Exp. 9, use of MGA + estradiol increased (P < .05) the estrous and subsequent lambing response of 30-d postpartum ewes temporarily weaned of their lambs. In contrast, lambing response of 90-d postpartum ewes treated with MGA + estradiol was not increased by temporary removal of lambs (Exp. 10). These data collectively support the hypothesis that treatment of ewes with MGA + estradiol is an effective and practical approach for inducing and synchronizing a fertile estrus in ewes during the non-breeding season.

Effect of implant sequence and dose on feedlot cattle performance.

Studies were conducted to evaluate the effects of delayed implanting or the use of a low-dose implant followed by a higher-dose implant in feedlot cattle. In the first study, 150 steers were allotted to 15 pens (three pens/treatment) and assigned to a nonimplant treatment (control), a single zeranol (36 mg) implant (R), or a double zeranol implant (DR) administered at the start of a 140-d finishing period, or a single zeranol implant administered at the start of an 80-d growing period, followed by a single (RR) or double (RDR) zeranol implant administered at the start of the finishing period. Steers managed under the DR, RR, and RDR implant schemes had greater (P < .10) finishing period gains and intakes than the control steer group. However, only DR and RDR steer groups had improved (P < .10) finishing period feed conversions compared with control steers. In combined growing and finishing periods, the RDR steer group displayed the lowest (P = .12) feed:gain ratio. In a second trial, conducted concurrently to the zeranol trial, steers that did not receive an initial implant containing 20 mg of estradiol benzoate plus 200 mg of progesterone (S) but were subsequently implanted twice, once at the start of the finishing period and again 80 d later, had a lower (P < .11) finishing period feed:gain ratio (6.08 vs 6.51) than steers implanted all three times.(ABSTRACT TRUNCATED AT 250 WORDS)

Growth and slaughter characteristics of ram and wether lambs implanted with zeranol.

Forty-nine Columbia ram and wether lambs born in April 1990 and 46 born in April 1991 were studied to determine the effects of zeranol implants on growth, difficulty of pelt removal, and carcass characteristics. Implanting ram and wether lambs once (1990) or twice (1991) with 12 mg of zeranol did not change live weight or ADG but gain/feed decreased (P < .05) in ram lambs slaughtered at approximately 50 kg. Testes weight was reduced approximately 50% by implanting. Two implants reduced (P < .05) the force needed to pull the pelt from the hind legs of ram lambs, but implanting tended to increase the force required to pull the pelt from wether lambs. Data for pelt weight, force required to pull the pelt, percentage of the carcass in the shoulder or splenius muscle, and Warner-Bratzler shear values showed that zeranol implants resulted in ram lambs becoming more like wethers and wether lambs becoming more like rams. Implanting with zeranol did not affect closure of the metacarpal growth plate in ram or in wether lambs. Difficulty of pelt removal can be reduced by implanting ram lambs with 12 mg of zeranol at approximately 114 d of age and reimplanting zeranol 28 d later.