Effect of ruminally protected amino acids and zilpaterol hydrochloride on performance, carcass, and skeletal muscle fiber characteristics of finishing beef steers
Hosford, Andrew David
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Metabolic modifiers have profound effects on muscle and fat development, meat quality, and economic value. A series of experiments were conducted to gain a better understanding of how zilpaterol hydrochloride (ZH) affected skeletal muscle growth in feedlot cattle. The objective of the first experiment (Exp. 1) was to evaluate the effect of ZH on gene an protein expression in muscle; in order to gain a better understanding of the mode of action for ZH. The objective of the second experiment (Exp. 2) was studying a major cellular energy regulator AMP-activated protein kinase (AMPK). Additionally, to see if there was a method to enhance the typical ZH response utilizing ruminally-protected lysine and methionine. The objective of the last experiment (Exp. 3) was to evaluate three different ratios of lysine to methionine in culture media on bovine skeletal muscle satellite cells. Skeletal muscle tissue was collected from cattle fed either 0 or 8.3 mg/kg ZH for 20 d (Exp. 1). Total RNA and protein were extracted and analyzed to detect differences in gene and protein expression. Additionally, bovine skeletal muscle satellite cells were collected to evaluate 0 or 10 µM concentrations of ZH on cellular gene and protein expression and differing ratios of lysine to methionine. Crossbred steers (n = 180) were blocked by weight and then randomly assigned to treatments (Exp. 2). Treatment groups consisted of no ZH and no supplemental amino acids (Cont-), ZH and no supplemental amino acids (Cont+), ZH and ruminally-protected Lysine (Lys), ZH and ruminally-protected methionine (Met), and ZH with ruminally-protected lysine and methionine (Lys+Met). Zilpaterol hydrochloride was supplemented at a rate of 8.3 mg/kg for the last 20 d with a 3 d withdrawal. Lysine and methionine were top dressed daily for the 134 d feeding period in order to provide 12 or 4 g•hd-1•d-1 respectfully. Bovine Satellite cells isolated from semimembranosus muscle were plated on tissue cultures (Exp. 3). Cells grown in proliferation media until 80% confluence and then a custom made DMEM media with varying ratios of lysine to methionine were added (3:1, 1.5:1, and 1:1) with or without 1 µM ZH for cellular differentiation. Cells were then harvested at either 48, 96, r 120 h post-differentiation. Total RNA and protein were isolated, RTQPCR and Western blotting techniques were conducted in order to observe defenses in gene and protein expression. Our findings indicated increased AMPK and phosphorylated AMPK gene and protein expression in steers supplemented with ZH as compare to control cattle. Myosin heavy-chain IIX (MHCIIX) was also increased with ZH fed steers. AMPK activation is known to cause shifts in MHC expression and we think that ZH's effects on muscle fiber type is mediated via direct activation of AMPK. During the ZH feeding period specifically, but also the entire 134 d feeding period, Lys+Met treated cattle had better average daily gain and gain:feed. Additionally, Met and Lys+Met supplemented cattle had greater hot carcass weight (HCW) as compared to Cont-. Lysine and ZH supplemented cattle were the fattest carcasses but also had the toughest meat up to 14 day of aging when compared to cattle that received no amino acid supplement with or without ZH. The Lys+Met cattle had thicker MHCIIX muscle fiber as compared to control. Suggesting differential response with amino acids and ZH supplementation. Furthermore, nuclei density was elevated with control cattle as compared to all ZH supplemented cattle. This is a clear dilution effect of less nuclei into larger muscle fibers. Lastly, with bovine satellite cells, no clear ratio of lysine to methionine demonstrated an advantage with ZH supplementation. Potentially our cellular lysine:methionine were not at the correct levels to enhance the ZH response and needs to be investigated further. Our data shows that when ZH is supplemented, AMPK protein is up-regulated and activated. This suggests that ZH's effects on fiber type and diameter could be through its effects on AMPK. Additionally, that lysine and methionine may be limiting during ZH feeding and performance and carcass characteristics were improved when these limiting amino acids were administered.