Transcriptome analyses indicate that heat stress-induced inflammation in white adipose tissue and oxidative stress in skeletal muscle is partially moderated by zilpaterol supplementation in beef cattle.

Heat stress (HS) triggers oxidative stress, systemic inflammation, and disrupts growth efficiency of livestock. ß-adrenergic agonists supplemented to ruminant livestock improve growth performance, increase skeletal muscle mass, and decrease carcass fat. The objective of this study was to understand the independent and interacting effects of HS and zilpaterol hydrochloride (ZH) supplementation on the transcriptome of subcutaneous white adipose tissue and the longissimus dorsi muscle in steers. Twenty-four Red Angus-based steers were assigned to thermoneutral (TN; Temperature Humidity Index [THI] = 68) or HS (THI = 73-85) conditions and were not supplemented or supplemented with ZH (8.33 mg/kg/d) for 21 d in a 2 × 2 factorial. Steers in the TN condition were pair-fed to the average daily feed intake of HS steers. RNA was isolated from adipose tissue and skeletal muscle samples collected via biopsy on 3, 10, and 21 d and sequenced using 3' Tag-Seq to an achieved average depth of 3.6 million reads/sample. Transcripts, mapped to ARS-UCD1.2, were quantified. Differential expression (DE) analyses were performed in DESeq2 with a significance threshold for false discovery rate of 0.05. In adipose, 4 loci (MISP3, APOL6, SLC25A4, and S100A12) were DE due to ZH on day 3, and 2 (RRAD, ALB) were DE due to the interaction of HS and ZH on day 10 (Padj < 0.05). In muscle, 40 loci (including TENM4 and OAZ1) were DE due to ZH on day 10, and 6 loci (HIF1A, LOC101903734, PDZD9, HNRNPU, MTUS1, and TMCO6) were DE due to environment on day 21 (Padj < 0.05). To explore biological pathways altered by environment, supplement, and their interaction, loci with DE (Praw < 0.05) were evaluated in Ingenuity Pathway Analysis. In adipose, 509 pathways were predicted to be altered (P < 0.01): 202 due to HS, 126 due to ZH, and 181 due to the interaction; these included inflammatory pathways predicted to be upregulated due to HS but downregulated due to the interaction of HS and ZH. In muscle, 113 pathways were predicted to be altered (P < 0.01): 23 due to HS, 66 due to ZH, and 24 due to the interaction of HS and ZH. Loci and pathway data in muscle suggest HS induced oxidative stress and that the stress response was moderated by ZH. Metabolic pathways were predicted to be altered due to HS, ZH, and their interaction in both tissues. These data provide evidence that HS and ZH interact to alter expression of genes in metabolic and immune function pathways and that ZH moderates some adverse effects of HS.

Heat stress (HS) negatively impacts livestock health and carcass quality. Supplementation of livestock with ß-adrenergic agonists (ß-AA) increases muscle mass and decreases fat deposition. The purpose of this study was to understand how HS and zilpaterol hydrochloride (ZH), a ß-AA, alter gene expression in muscle and in adipose of cattle. Twenty-four steers were assigned to thermoneutral (TN) or HS conditions and were not supplemented (NS) or supplemented with ZH for 21 d. RNA was isolated from muscle and adipose collected on days 3, 10, and 21 to identify changes in gene expression. Several individual loci were differentially expressed (DE) due to HS or ZH in both tissues while the interaction of HS and ZH altered expression in adipose. A less stringent definition of DE used to explore biological pathways predicted that both treatments alter metabolism. Pathway analyses also supported that HS increased inflammation in adipose, but that these inflammatory pathways were downregulated by ZH. HS also was predicted to induce oxidative stress in muscle although ZH moderated this response. This study provides information on how HS and ß-AA act independently and interact to alter physiology, lending insight useful for the development of management and mitigation strategies for stress.

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.

Growth, carcass characteristics, cut yields and meat quality of lambs finished with zilpaterol hydrochloride and steroid implant.

Forty hairbreed male lambs were used to evaluate the effects of zilpaterol hydrochloride (ZH, 0 and 0.15 mg/kg BW) and steroid implant (SI, without and with 52.5 mg trenbolone acetate and 7.5 mg 17ß-estradiol) on feedlot performance, carcass characteristics, non-carcass components, wholesale cut yield, and meat quality. Supplemental ZH increased growth rate, feed efficiency, carcass weight, and dressing percentage, with no effect on wholesale cut yields. Feeding ZH increased muscle pH at 24 h. Supplemental ZH increased meat shear force, but decreased lightness, redness, and yellowness after frozen storage followed by a 14-day aging period. The SI administration increased dressing percentage and neck yield, but decreased testicle weight and meat redness, without affecting other variables. The LT area was greater with ZH + SI administration than with individual application of ZH or SI. Compared to individual administration, simultaneous application of ZH and SI did not result in improved growth performance, carcass traits and wholesale cut yields in hairbreed male lambs.

Profiling the Nucleobase and Structure Selectivity of Anticancer Drugs and other DNA Alkylating Agents by RNA Sequencing.

Drugs that covalently modify DNA are components of most chemotherapy regimens, often serving as first-line treatments. Classically, the reactivity and selectivity of DNA alkylating agents has been determined in vitro with short oligonucleotides. A statistically sound analysis of sequence preferences of alkylating agents is untenable with serial analysis methods because of the combinatorial explosion of sequence possibilities. Next-generation sequencing (NGS) is ideally suited for the broad characterization of sequence or structure selectivities because it analyzes many sequences at once. Herein, NGS is used to report on the chemoselectivity of alkylating agents on RNA and this technology is applied to the previously uncharacterized alkylating agent trimethylsilyl diazomethane.

Acute exposure of primary rat soleus muscle to zilpaterol HCl (ß2 adrenergic agonist), TNFα, or IL-6 in culture increases glucose oxidation rates independent of the impact on insulin signaling or glucose uptake.

Recent studies show that adrenergic agonists and inflammatory cytokines can stimulate skeletal muscle glucose uptake, but it is unclear if glucose oxidation is similarly increased. Thus, the objective of this study was to determine the effects of ractopamine HCl (ß1 agonist), zilpaterol HCl (ß2 agonist), TNFα, and IL-6 on glucose uptake and oxidation rates in unstimulated and insulin-stimulated soleus muscle strips from adult Sprague-Dawley rats. Effects on phosphorylation of Akt (phospho-Akt), p38 MAPK (phospho-p38), and p44/42 MAPK (phospho-p44/42) was also determined. Incubation with insulin increased (P<0.05) glucose uptake by ∼47%, glucose oxidation by ∼32%, and phospho-Akt by ∼238%. Insulin also increased (P<0.05) phospho-p38, but only after 2h in incubation. Muscle incubated with ß2 agonist alone exhibited ∼20% less (P<0.05) glucose uptake but ∼32% greater (P<0.05) glucose oxidation than unstimulated muscle. Moreover, co-incubation with insulin+ß2 agonist increased (P<0.05) glucose oxidation and phospho-Akt compared to insulin alone. Conversely, ß1 agonist did not appear to affect basal or insulin-stimulated glucose metabolism, and neither ß agonist affected phospho-p44/42. TNFα and IL-6 increased (P<0.05) glucose oxidation by ∼23% and ∼33%, respectively, in the absence of insulin. This coincided with increased (P<0.05) phospho-p38 and phospho-p44/42 but not phospho-Akt. Furthermore, co-incubation of muscle with insulin+either cytokine yielded glucose oxidation rates that were similar to insulin alone, despite lower (P<0.05) phospho-Akt. Importantly, cytokine-mediated increases in glucose oxidation rates were not concomitant with greater glucose uptake. These results show that acute ß2 adrenergic stimulation, but not ß1 stimulation, directly increases fractional glucose oxidation in the absence of insulin and synergistically increases glucose oxidation when combined with insulin. The cytokines, TNFα and IL-6, likewise directly increased glucose oxidation in the absence of insulin, but were not additive in combination with insulin and in fact appeared to disrupt Akt-mediated insulin signaling. Rather, cytokines appear to be acting through MAPKs to elicit effects on glucose oxidation. Regardless, stimulation of glucose oxidation by these key stress factors did not rely upon greater glucose uptake, which may promote metabolic efficiency during acute stress by increasing fractional glucose oxidation without increasing total glucose consumption by muscle.

Feedlot performance and carcass traits of hairbreed ewe lambs in response to zilpaterol hydrochloride and soybean oil supplementation.

The aim of the present study was to evaluate the impact of zilpaterol hydrochloride (ZH; 0 or 10 mg/lamb daily) and soybean oil (SBO; 0 or 6%) supplementation on feedlot performance, carcass traits, and wholesale cut yield of 32 Dorper × Pelibuey ewe lambs (30.55 ± 2. 57 kg of initial BW). Lambs were blocked by BW and randomly assigned to treatments under a randomized complete block design with a 2 × 2 factorial arrangement. After a 34-d feeding period, all ewes were slaughtered. No ZH × SBO interactions were detected (P ≥ 0.11) for the variables evaluated. In the overall feeding period and first 17 d of experiment, feedlot performance was not affected (P ≥ 0.26) by ZH supplementation, but from d 18 to 34, ZH increased (P ≤ 0.03) total gain, ADG, and G:F without affecting DMI (P = 0.58). Also, ZH increased (P ≤ 0.02) HCW, cold carcass weight, dressing percentage, LM area, and leg perimeter. Lung weight as percentage of final BW decreased (P = 0.05) whereas other noncarcass components and wholesale cut yields were not affected (P ≥ 0.06) by ZH supplementation. Inclusion of SBO did not affect (P ≥ 0.08) feedlot performance or wholesale cut yields. The LM pH at 24 h postmortem as well as liver and peritoneum percentages were decreased (P ≤ 0.05) by SBO supplementation, but no other carcass characteristics or noncarcass components were affected (P ≥ 0.08) by SBO. In conclusion, feedlot performance and carcass characteristics were not altered by the interaction of ZH × SBO. However, ZH alone increased the growth of ewes during the last 17 d of the feeding period. Likewise, carcass characteristics of economic importance (i.e., HCW, dressing percentage, LM area, and leg perimeter) increased with ZH supplementation. In general, feedlot performance, carcass traits, and wholesale cut yields were not altered by including 6% of SBO in the finishing diet of ewe lambs.

The effects of technology use in feedlot production systems on feedlot performance and carcass characteristics.

The objectives of this study were to examine the effects of feedlot production systems with and without the use of a ß-adrenergic agonist compared to an all-natural production program on feedlot performance and carcass characteristics. Crossbred beef steers ( = 336; initial BW = 379 ± 8 kg) were randomized to 1 of 3 treatments in a randomized complete block design (RCBD; 14 steers/pen; 8 pens/treatment). Treatments consisted of an all-natural treatment (NAT), a conventional treatment (CONV), and a conventional treatment with a ß-agonist (CONV-Z). All treatments were fed the same basal diet with NAT cattle receiving no growth promoting technologies. The CONV and CONV-Z cattle were implanted with 40 mg of estradiol and 200 mg of trenbolone acetate (TBA) on d 0 and were fed 33 and 9 mg/kg of monensin and tylosin daily, respectively. The CONV-Z cattle were fed zilpaterol hydrochloride (ZH) at 6.76 mg/kg (90% DM basis) for the last 20 days on feed (DOF) There was no effect of treatment on DMI ( = 0.83); however, CONV-Z steers gained 3.8% faster (1.64 vs. 1.58 kg/d; < 0.01) and were 5.3% more efficient (0.160 vs. 0.152; < 0.01) than CONV steers, and CONV steers gained 32.8% faster (1.58 vs. 1.19 kg/d; < 0.01) and were 26.7% more efficient (0.152 vs. 0.120; < 0.01) than NAT steers. There was a 35.7% improvement in estimated carcass gain (1.29 vs. 0.95 kg/d; < 0.01) and a 32.6% improvement in carcass efficiency (0.126 vs. 0.095; < 0.01) for CONV-Z steers compared to NAT steers. Hot carcass weight was increased by 8 kg for CONV-Z steers compared to CONV steers (394 vs. 386 kg; = 0.05) and 46 kg compared to NAT steers (394 vs. 348 kg; < 0.01). Longissimus muscle area was increased by 3.6 cm for CONV-Z steers compared to CONV steers (92.29 vs. 88.67 cm; = 0.02) and 12.1 cm for CONV-Z steers compared to NAT steers (92.29 vs. 80.16 cm; < 0.01), resulting in a 9.6% unit increase in USDA yield grade (YG) 1 (15.14 vs. 5.52%; < 0.05) and a 21.6% unit reduction in USDA YG 3 for CONV-Z steers compared to CONV steers (30.70 vs. 52.32%; < 0.05). The CONV-Z steers had a lower marbling score compared to the other treatments (432; 0.01), resulting in an 11.7% unit increase (20.70 vs. 9.03%; < 0.05) in USDA Select carcasses compared to CONV steers. The results of this experiment show that CONV-Z and CONV production results in a significant improvement in feedlot performance and USDA YG compared to NAT.

Effects of beef production system on animal performance and carcass characteristics.

The objective of this study was to evaluate conventional (CONV) and natural (NAT) beef production systems from annual pasture through finishing through grazing. Beef steers (n=180, initial BW=250±19 kg) were assigned randomly to 2 treatments in the pasture phase. Steers were implanted with 40 mg of trenbolone acetate (TBA), 8 mg estradiol, and 29 mg tylosin tartrate (CONV), or received no implant (NAT). Steers on the 2 treatments grazed wheat or cereal rye for 109 d. Conventional steers had an 18.5% improvement in ADG (1.22 vs. 1.03 kg/d, P<0.01) and a heavier final BW (385 vs. 366 kg, P<0.01) compared with NAT steers. Following the pasture phase, steers (n=160 steers, 5 steers/pen, 8 pens/treatment) were assigned to a 2×2 factorial in the feedlot phase. Production system (NAT vs. CONV) was maintained from the pasture phase, and the second factor was 7 vs. 12% low-quality roughage (DM basis, LOW vs. HIGH). During finishing, CONV steers were given 120 mg of TBA and 24 mg estradiol at processing, fed monensin and tylosin, and fed zilpaterol hydrochloride for the last 20 d of the experiment. There were no program×roughage level interactions (P>0.07). The CONV steers ate 6.9% more feed (11.8 vs. 11.0 kg/d, P<0.01), gained 28.4% faster (1.90 vs. 1.48 kg/d, P<0.01), and were 24.2% more efficient (0.164 vs. 0.132, P<0.01) compared with NAT steers. The LOW steers had greater G:F (0.153 vs. 0.144, P<0.01) compared with HIGH steers. There was a 28.3% improvement in estimated carcass weight gain (1.36 vs. 1.06 kg/d), 18.6% improvement in carcass efficiency (0.115 vs. 0.097, P<0.01), and 21.6% improvement (1.52 vs. 1.25 Mcal/kg, P<0.01) in calculated dietary NEg for CONV compared with NAT steers. Hot carcass weight was increased by 62 kg (424 vs. 362 kg, P<0.01) and LM area was increased by 16.9 cm2 (100.9 vs. 84.0 cm2, P<0.01), decreasing USDA yield grade (YG, 3.09 vs. 3.54, P<0.01) for CONV steers compared with NAT steers. Natural steers had a greater percentage of carcasses in the upper 2/3 of USDA Choice grade (48.7 vs. 18.7%, P<0.01), a greater percentage of YG 4 and 5 carcasses (25.4 vs. 9.3%, P<0.01), and a greater percentage of abscessed livers (39.6 vs. 10.5%, P<0.01) compared with CONV steers. The results show that CONV production results in more rapid and efficient production that resulted in heavier carcasses with superior YG and desirable quality grades with both roughage levels.

Biological responses of beef steers to steroidal implants and zilpaterol hydrochloride.

British × Continental steers (n = 168; 7 pens/treatment; initial BW = 362 kg) were used to evaluate the effect of dose/payout pattern of trenbolone acetate (TBA) and estradiol-17ß (E2) and feeding of zilpaterol hydrochloride (ZH) on serum urea-N (SUN), NEFA, IGF-I, and E2 concentrations and LM mRNA expression of the estrogen (ER), androgen (ANR), IGF-I (IGF-IR), ß1-adrenergic (ß1-AR), and ß2-adrenergic (ß2-AR) receptors and IGF-I. A randomized complete block design was used with a 3 × 2 factorial arrangement of treatments. Main effects were implant (no implant [NI], Revalor-S [REV-S; 120 mg TBA + 24 mg E2], and Revalor-XS [REV-X; 200 mg TBA + 40 mg E2]) and ZH (0 or 8.3 mg/kg of DM for 20 d with a 3-d withdrawal). Steers were fed for 153 or 174 d. Blood was collected (2 steers/pen) at d -1, 2, 6, 13, 27, 55, 83, 111, and 131 relative to implanting; LM biopsies (1 steer/pen) were collected at d -1, 27, 55, and 111. Blood and LM samples were collected at d -1, 11, and 19 relative to ZH feeding. A greater dose of TBA + E2 in combination with ZH increased ADG and HCW in an additive manner, suggesting a different mechanism of action for ZH and steroidal implants. Implanting decreased (P < 0.05) SUN from d 2 through 131. Feeding ZH decreased (P < 0.05) SUN. Serum NEFA concentrations were not affected by implants (P = 0.44). There was a day × ZH interaction (P = 0.06) for NEFA; ZH steers had increased (P < 0.01) NEFA concentrations at d 11 of ZH feeding. Serum E2 was greater (P < 0.05) for implanted steers by d 27. Serum trenbolone-17ß was greater (P < 0.05) for implanted steers by d 2 followed by a typical biphasic release rate, with a secondary peak at d 111 for REV-X (P < 0.05) implanted steers. Implanting did not affect mRNA expression of the ANR or ER, but the IGF-IR and the ß1-AR and ß2-AR were less (P < 0.05) for REV-S than NI at d 55 and ß2-AR mRNA was less (P < 0.05) for REV-S than for REV-X. Expression of the IGF-IR and the ß1-AR at d 111 was greater (P< 0.05) for REV-X than for REV-S and NI at d 111, and the ß2-AR was less (P< 0.05) for REV-S than for REV-X. Feeding ZH did not affect mRNA expression of the ß1-AR and ß2-AR. Both implanting and feeding ZH decreased SUN, but a greater dose of TBA + E2 did not result in further decreases. In addition, feeding ZH increased serum NEFA concentrations. Metabolic changes resulting from implanting and feeding ZH may aid in explaining steer performance and carcass responses to these growth promotants.

Effects of zilpaterol hydrochloride and soybean oil supplementation on feedlot performance and carcass characteristics of hair-breed ram lambs under heat stress conditions.

Forty Dorper × Pelibuey ram lambs initially weighing 31.7 ± 2.30 kg were stratified by BW and randomly assigned to treatments under a completely randomized design with a 2 × 2 factorial arrangement of treatments to evaluate effects of zilpaterol hydrochloride (ZH; 0 or 10 mg/lamb daily) and soybean oil (SBO; 0 or 6%) on feedlot performance, carcass characteristics, and wholesale cut yield of ram lambs under heat stress conditions. After a 34-d feeding period, all lambs were harvested. Climatic conditions were of moderate heat stress (average temperature 35.7°C) for lambs during the study. Interactions ZH × SBO were not observed (P ≥ 0.11) for any of the variables evaluated. During the first 17 d of experiment, ZH increased (P ≤ 0.05) BW, ADG, and G:F without affecting feed intake (P = 0.40), but from d 18 to 34 and the entire 34-d feeding period, feedlot performance was not affected (P = 0.18) by ZH. Also, ZH decreased KPH, dressing percent, LM area, LM pH at 24 h postmortem, and leg perimeter (P ≤ 0.04). Renal fat (P = 0.03) decreased with ZH while other noncarcass components were not affected (P ≥ 0.06) by ZH supplementation. Leg yield (P = 0.01) and plain loin (P = 0.04) decreased with ZH and yields of other wholesale cuts were not affected (P ≥ 0.10) by ZH. Feedlot performance (P ≥ 0.20) and wholesale cut yield (P ≥ 0.21) were not affected by SBO. Additionally, dressing percentage decreased (P < 0.01) with SBO while other carcass characteristics (P ≥ 0.12) were not affected by SBO. In conclusion, inclusion of both ZH and SBO in feedlot finishing diets did not improve feedlot performance, carcass characteristics, or wholesale cut yield of hair-breed ram lambs under moderate heat stress. Feedlot performance responded only to ZH and only during the first 17 d of the feeding period. In addition, some carcass characteristics of economic importance, such as dressing, LM area, and leg yield, were improved by ZH.

Growth promoting technologies reduce greenhouse gas, alcohol, and ammonia emissions from feedlot cattle.

Increased animal productivity has the potential to reduce the environmental impact per unit of consumable product and is believed to be the most promising and sustainable mitigation technique to meet increasing demand for high quality protein. The feedlot industry uses ionophores, antibiotics, growth implants, and ß2-adrenergic agonists to improve health and growth performance of cattle. These technologies not only increase productivity but also alter microbes in the rumen and increase nitrogen retention in the animal, which may lead to changes in greenhouse gas (GHG), volatile organic compound (VOC), and ammonia (NH3) emissions from feedlot cattle. The present study investigated GHG, VOC, and NH3 emissions from 160 Angus crossbred steers. Steers were blocked by weight in a randomized block design and assigned to 16 pens of 10 animals each. Treatments applied were 1) control (CON; no technology application), 2) monensin and tylosin phosphate (MON), 3) monensin, tylosin phosphate, and growth implant (IMP), and 4) monensin, tylosin phosphate, growth implant, and zilpaterol hydrochloride (fed during the last 20 d of the feeding period; BAA). Cattle were on feed for an average of 107 d. Performance variables (DMI, BW, ADG, and G:F) and carcass traits (HCW, dressing percent, KPH, LM area, fat thickness, marbling score, yield grade, and quality grade) were measured. Gaseous emissions were measured during the last 10 d of the feeding period when animals were housed in 4 totally enclosed identical cattle pen enclosures. To quantify gaseous emissions a 4×4 Latin square design (n=4) was used. Gaseous emissions were analyzed using Proc Mixed in SAS and reported in grams per kilogram HCW per day and grams per kilogram per animal per hour. Treatment with IMP and BAA increased (P<0.05) ADG, final BW, and HCW. Cattle on BAA had greater HCW and LM area (P<0.05) and had lower (P<0.05) CH4, methanol, and NH3 emissions per kilogram HCW than cattle on the remaining treatments. Methane emissions were similar for CON and IMP treated cattle. Nitrous oxide emissions were similar across CON, MON, and IMP treated cattle and were higher in BAA treated cattle (P<0.05). The present study provides a better understanding of how application of growth promoting technologies to feedlot steers affects GHG, VOC, and NH3 emissions per kilogram of product.

Effects of sequential feeding of ß-adrenergic agonists on cull cow performance, carcass characteristics, and mRNA relative abundance.

The objectives of this study were to determine the effects of supplementation with a single ß-adrenergic agonist (ß-AA) or a sequence of ß-AA on cow performance, carcass characteristics, and mRNA relative abundance of cull cows implanted and fed a concentrate diet. Sixty cull cows were implanted with Revalor-200 (200 mg of trenbolone acetate and 20 mg of estradiol) and assigned to 1 of 4 treatments (n = 15/treatment): CON = fed a concentrate diet only; RH = supplemented with ractopamine-HCl for the last 25 d before slaughter; ZH = supplemented with zilpaterol-HCl for 20 d before a 3-d withdrawal before slaughter; RH + ZH = supplemented with RH for 25 d, followed by ZH for 20 d before a 3-d withdrawal before slaughter. Ractopamine-HCl was supplemented at a dose of 200 mg·animal(-1)·d(-1), and ZH was supplemented at 8.33 mg/kg (100% DM basis) of feed. All cows were fed a concentrate diet for 74 d. Each treatment had 5 cows per pen and 3 replicate pens. Body weights were collected on d 1, 24, 51, and 72. Muscle biopsies from the LM were collected on d 24, 51, and at slaughter from a subsample of 3 cows per pen. Carcass traits were evaluated postslaughter. The 2 ZH treatments averaged 15.3 kg more BW gain, 0.20 kg greater ADG, and 7.8 cm(2) larger LM area than CON and RH treatments, and 21 kg more HCW than CON, but these differences were not significant (P > 0.10), likely due to a sample size of n = 15/treatment. The sequence of RH followed by ZH tended to optimize the combination of HCW, LM area, percent intramuscular fat, and lean color and maturity compared with the ZH treatment. Abundance of ß(2)-adrenergic receptor (AR) mRNA was not altered in the RH + ZH treatment during RH supplementation from d 24 to 51 of feeding. However, the abundance of ß(2)-AR mRNA increased (P < 0.05) the last 23 d of feeding for the RH treatment and tended (P = 0.10) to increase in ZH cows during ZH supplementation. For all cows, abundance of type IIa myosin heavy chain (MHC-IIa) mRNA decreased (P < 0.05) after 24 d of feeding. Abundance of MHC-IIx mRNA increased (P < 0.05) for ZH and RH + ZH treatments the last 23 d of feeding during ZH supplementation. Although few significant differences were observed in performance or carcass traits, mRNA quantification indicated that ß-AA supplementation elicited a cellular response in cull cows. Implanting and feeding cull cows for 74 d, regardless of ß-AA supplementation, added economic value by transitioning cows from a cull cow to what is referred to in industry as a white cow market in which cows have white fat resulting from grain feeding.

Performance of finishing beef steers in response to anabolic implant and zilpaterol hydrochloride supplementation.

Our objectives were to evaluate the dose/payout pattern of trenbolone acetate (TBA) and estradiol-17ß (E(2)) implants and feeding of zilpaterol hydrochloride (ZH) on performance and carcass characteristics of finishing beef steers. A randomized complete block design was used with a 3 × 2 factorial arrangement of treatments. British × Continental steers (n = 168; initial BW = 362 kg) were blocked by BW and allotted randomly to 42 pens (7 pens/treatment; 6 pens/block; 4 steers/pen). The main effects of treatment were implant [no implant (NI); Revalor-S (REV-S; 120 mg of TBA + 24 mg of E(2)); and Revalor-XS (REV-X; 200 mg of TBA + 40 mg of E(2))] and ZH (0 or 8.3 mg/kg of DM for 20 d with a 3-d withdrawal before slaughter). Blocks were split into 2 groups, and block groups were fed for either 153 or 174 d. No implant × ZH interactions were noted for cumulative performance data. Overall, shrunk final BW (567, 606, and 624 kg for NI, REV-S, and REV-X, respectively), ADG (1.25, 1.51, and 1.60 kg), and G:F (0.14, 0.16, and 0.17) increased (P < 0.05) as TBA and E(2) dose increased. Implanting increased (P < 0.05) DMI, but DMI did not differ (P > 0.10) between REV-S and REV-X (8.8 for NI vs. 9.4 kg/d for the 2 implants). From d 1 to 112 of the feeding period, implanting increased (P < 0.05) ADG and G:F, but REV-S and REV-X did not differ (P > 0.10). From d 112 to end, ADG increased by 19% (P < 0.05) and G:F was 18% greater (P < 0.05) for REV-X vs. REV-S. Carcass-adjusted final BW (29-kg difference), ADG (0.2-kg/d difference), and G:F (0.02 difference) were increased (P < 0.05) by ZH, but daily DMI was not affected by feeding ZH. Hot carcass weight was increased (P < 0.05) by ZH (19-kg difference) and implant, with REV-X resulting in the greatest response (HCW of 376 for NI vs. 404 and 419 kg for REV-S and REV-X, respectively; P < 0.05). An implant × ZH interaction (P = 0.05) occurred for dressing percent (DP). Without ZH, implanting increased DP, but DP did not differ (P > 0.10) between REV-X and REV-S. With ZH, REV-X increased (1.7%; P < 0.05) DP vs. NI and REV-S. Marbling score, 12th-rib fat, and KPH were not affected (P > 0.10) by implant or ZH. Overall, treatment increased steer performance and HCW in an additive fashion, suggesting different mechanisms of action for ZH and steroidal implants. In addition, a greater dose of TBA + E(2) and extended payout improved steer performance and HCW.

The design, synthesis and antiviral evaluation of a series of 5-trimethylsilyl-1-beta-D-(arabinofuranosyl)uracil phosphoramidate ProTides.

BACKGROUND: Nucleoside analogues always require phosphorylation to be active. This appears to be a particular limitation for uridine-based nucleosides. Our ProTide method allows the direct use of masked membrane-soluble preformed nucleoside phosphates, bypassing the need for the initial phosphorylation step. We herein applied it to some novel 5-trimethylsilyl arabinosyl uridines. METHODS: 5-Trimethylsilyl-1-beta-D-(arabinofuranosyl)uracil was prepared in six steps starting from uridine, and five phosphoramidate ProTide derivatives were synthesized. These compounds were investigated for activity against a range of DNA and RNA viruses, including herpes simplex virus type-1 and type-2, vaccinia virus and HIV. RESULTS: Overall, these compounds did not show significant antiviral activity against any of the viruses tested. CONCLUSIONS: The inactivity of the ProTides of this nucleoside could correspond with poor ProTide activation in vitro, poor onward metabolism or low activity of the putative monophosphate metabolite.

Additive effects of a steroidal implant and zilpaterol hydrochloride on feedlot performance, carcass characteristics, and skeletal muscle messenger ribonucleic acid abundance in finishing steers.

This experiment investigated the effects of zilpaterol hydrochloride (ZH) and the steroidal implant Revalor-S (RS; 120 mg of trenbolone acetate and 24 mg of estradiol-17beta) on finishing steer performance and the mRNA concentration of beta-adrenergic receptors (beta-AR) types I and II, and types I, IIA, and IIX myosin heavy chain (MHC) isoforms. A total of 2,279 feedlot steers weighing 426 +/- 6.4 kg were administered no implant or RS on d 0, and fed 0 or 8.3 mg of ZH/kg of diet DM during the last 30 d with a 3-d withdrawal. Treatments were randomly assigned to 24 pens (n = 6 pens/treatment). At slaughter, semimembranosus muscle tissue was excised for RNA isolation from 4 carcasses per pen. No interactions were detected for any of the variables measured in the experiment. Administration of ZH during the last 30 d of the feeding period increased (P < 0.01) ADG, G:F, HCW, and LM area; decreased (P < 0.01) 12th-rib fat depth and marbling; and improved (P < 0.01) yield grade. Treatment had no effect on beta1-AR mRNA levels, but there was an increase (P = 0.01) in beta(2)-AR mRNA levels due to ZH inclusion. Myosin heavy chain-I (MHC-I) mRNA levels were unaffected by treatment. For MHC-IIA mRNA concentrations, administration of RS tended (P = 0.08) to increase mRNA levels, whereas ZH feeding the last 30 d tended (P = 0.08) to decrease mRNA levels for this isoform of myosin. Feeding ZH the last 30 d before slaughter increased (P < 0.01) mRNA concentrations of MHC-IIX in semimembranosus muscle of steers. These data indicate the combined use of ZH and RS additively contributes to BW and carcass gain in finishing feedlot steers and decreases marbling scores and USDA quality grades. The LM area increased and fat thickness decreased. In addition, ZH feeding changes the mRNA levels of MHC isoforms to a faster, more glycolytic fiber type in bovine skeletal muscle. These changes in mRNA concentrations of MHC isoforms, due to ZH feeding, could be affecting skeletal muscle hypertrophy.

Contribution of AP-1 interference induced by TAC-101 to tumor growth suppression in a hepatocellular carcinoma model.

TAC-101, 4-[3,5-bis(trimethylsilyl)benzamido] benzoic acid, is a synthetic ligand for retinoic acid receptor (RAR)-alpha. Here, we demonstrate the contribution of TAC-101-induced AP-1 interference to stabilization of tumor growth. TAC-101 induced transcriptional activation of RAR, resulting in marked elevation of RARbeta, a representative retinoid response marker, and it also significantly repressed the transcriptional activity of AP-1 in JHH-7 cells. In contrast to JHH-7, JHH-6 is another RARalpha-expressing human hepatocellular carcinoma (HCC) cell line with constitutive activation of AP-1, but it is retinoid insensitive and did not respond to the TAC-101-induced RAR signal. TAC-101 did not inhibit AP-1 activity of the JHH-6 cell line, showing that AP-1 interference by TAC-101 must be in parallel with RAR activation. Interleukin-8 (IL-8), one of the AP-1-regulated factors which correlate with a poor prognosis in HCC patients, was found to be overexpressed in JHH-7 cells. TAC-101 reduced IL-8 production without cytotoxicity and inhibited the progression of HCC in the orthotopic mouse model with decreased tumor IL-8 level. These results suggest that downregulation of the extracellular biomarker for AP-1 interference via the induction of retinoid signals will enhance the pharmacological effect of TAC-101 on HCC and it could be useful as a surrogate biomarker of therapeutic efficacy.

A synthetic retinoid, TAC-101 (4-[3,5-bis (trimethylsilyl) benzamido] benzoic acid), plus cisplatin: potential new therapy for ovarian clear cell adenocarcinoma.

OBJECTIVE: A novel retinoid, TAC-101 (4-[3,5-bis (trimethylsilyl) benzamido] benzoic acid), induces apoptosis of ovarian clear cell adenocarcinoma. The antitumor effect of TAC-101 alone or combined with cisplatin was tested using human ovarian carcinoma. METHODS: Induction of genes related to apoptosis by TAC-101 or cisplatin was assessed by DNA microarray analysis. TAC-101 (8 mg/kg/day orally for 21 days), cisplatin (7 mg/kg intravenously on day 1), or a combination of both drugs at the same dosages was administered to nude mice implanted subcutaneously with RMG-I or RMG-II clear cell adenocarcinoma cells. The antitumor effect was evaluated by calculating the treated/control tumor volume ratio at 21 days after implantation. The histoculture drug response assay was also performed using fresh surgical specimens of human ovarian cancer to determine the 50% inhibitory concentration (IC50). RESULTS: Different apoptosis-related genes were induced by TAC-101 and cisplatin. Compared with control mice, the volume of both RMG-I and RMG-II tumors was significantly reduced (p<0.05) by either drug. The IC50 values of cisplatin and TAC-101 showed a significant correlation (p<0.01). CONCLUSION: These in vitro findings suggest that a combination of TAC-101 and cisplatin may be a potential new treatment for ovarian clear cell adenocarcinoma.

4- [3,5-bis(trimethylsilyl)benzamido] benzoic acid (TAC-101) induced fas expression and activated caspase-3 and -8 in a DLD-1 colon cancer cell line.

BACKGROUND: 4-[3,5-Bis (trimethylsilyl) benzamido] benzoic acid (TAC-101) is a novel retinobenzoic acid derivative which has a specific binding affinity to the retinoic acid receptors (RAR)alpha and RARbeta. Using time-dependent FACScan analysis, it was observed that TAC-101 induced apoptosis in a DLD-1 human colon cancer cell line. In this study, the induction of apoptosis-related proteins and the activities of caspases in a DLD-1 cell line under medication with TAC-101 were investigated. MATERIALS AND METHODS: DLD-1 cells were cultured with different concentrations of TAC-101 for 12, 24 and 48 h. The expressions of Fas, TNF-R1, DR3, bcl-2, Bax and Bid were measured using a Western blot analysis. The activities of caspase-3, -8 and -9 were measured using a colorimetric protease assay kit. RESULTS: The Western blot analysis showed that TAC-IO1 had almost no effect on the level of Bcl-2, Bax or Bid protein. Although TAC-101 did not change the expression of TNF-R1 and DR3, TAC-101 increased the expression of Fas in both a time- and a dose-dependent manner. A 3-fold increase in caspase-3 activity and a 1.5-fold increase in caspase-8 activity were observed in cells treated with TAC-101 in comparison to the control cells (p<0.01). CONCLUSION: Our data indicate that the death receptor root of the apoptotic signal transduction in DLD-1 cells mainly participates in the apoptotic induction of TAC-101. Because the compounds inducing apoptotic activity are frequent targets of cancer therapy, TAC-101 may be a good candidate for use in the treatment of colon cancer.

Integration of two-dimensional LC-MS with multivariate statistics for comparative analysis of proteomic samples.

LC-MS-based proteomics requires methods with high peak capacity and a high degree of automation, integrated with data-handling tools able to cope with the massive data produced and able to quantitatively compare them. This paper describes an off-line two-dimensional (2D) LC-MS method and its integration with software tools for data preprocessing and multivariate statistical analysis. The 2D LC-MS method was optimized in order to minimize peptide loss prior to sample injection and during the collection step after the first LC dimension, thus minimizing errors from off-column sample handling. The second dimension was run in fully automated mode, injecting onto a nanoscale LC-MS system a series of more than 100 samples, representing fractions collected in the first dimension (8 fractions/sample). As a model study, the method was applied to finding biomarkers for the antiinflammatory properties of zilpaterol, which are coupled to the beta2-adrenergic receptor. Secreted proteomes from U937 macrophages exposed to lipopolysaccharide in the presence or absence of propanolol or zilpaterol were analysed. Multivariate statistical analysis of 2D LC-MS data, based on principal component analysis, and subsequent targeted LC-MS/MS identification of peptides of interest demonstrated the applicability of the approach.

Beta-adrenergic receptor agonists induce the release of granulocyte chemotactic protein-2, oncostatin M, and vascular endothelial growth factor from macrophages.

Vascular endothelial growth factor (VEGF), oncostatin M (OSM), and granulocyte chemotactic protein-2 (GCP-2/CXCL6) are up-regulated in U937 macrophages and peripheral blood macrophages exposed to LPS, beta-adrenergic receptor (beta2-AR) agonists (e.g. zilpaterol, and clenbuterol) and some other agents that induce intracellular cAMP (prostaglandin E2, forskolin, and butyryl cAMP). LPS in combination with beta2-agonists and cAMP elevating agents had an additional effect on the release of VEGF, OSM, and CXCL6. These proteins are up-regulated after 16-24 h of exposure and this is mediated by the beta2-AR, as determined by time course experiments and the use of a specific beta2-AR antagonist (ICI 118551). Beta2-AR agonists are used as bronchodilators in the treatment of asthma, but appear to have no effect on the chronic inflammation of the disease. However, the up-regulation of VEGF, OSM, and CXCL6 may have adverse effects on the inflammatory process of asthma. These mediators are involved in the recruitment of neutrophils, airway remodelling and angiogenesis, known features of chronic inflammatory diseases. We propose that the up-regulation of these proteins could play a role in the adverse effects of prolonged excessive usage of beta2-AR agonists on the airways besides the desensitization of the beta2-AR.