Performing Department
(N/A)
Non Technical Summary
Cattle with high pulmonary arterial pressure (PAP) have a 20% higher feed-to-gain ratio in the feedlot and a 30% faster discoloration in postmortem products (2 days earlier) compared with cattle with low PAP. Although cattle with high PAP are usually identified in high-altitude beef production systems in the US, recent studies have reported more cases at low altitudes. The US high-altitude beef production system produces about 1.5 million calves per year, and issues with high PAP have cost about $60 million annually since 2010. On the other hand, worldwide feed price has reached its highest index since 2013, and the waste due to beef discoloration (194.70 million kg/year) is becoming a significant environmental concern to stakeholders and policymakers. A general intervention is needed immediately to manage high PAP animal production inefficiency and product defects. Our current proposal applies supranutritional VE supplementation to counter feed inefficiency and meat discoloration in feedlot cattle with high PAP. Successful completion of this objective will allow us to provide immediate guidance to manage high PAP cattle during the feedlot period and will enable us to evaluate the unknown effects of high PAP on muscle growth, mitochondrial function, and meat quality. This study will also examine how VE supplementation affects postmortem proteolysis and its relation to meat quality developments during industrial meat production. Successful completion of this objective will allow us to understand the linkage between dietary VE supplementation, proteome degradation, and beef quality.
Animal Health Component
45%
Research Effort Categories
Basic
45%
Applied
45%
Developmental
10%
Goals / Objectives
Our long-term goal is to apply affordable and consumer-favored feed supplementation to minimize beef quality defects to ensure the competitiveness and sustainability of US beef production. In the current proposal, our overall objective is to improve beef production efficiency and reduce meat quality defects of feedlot cattle with high pulmonary arterial pressure (PAP), along with redefining the linkage between dietary vitamin E supplementation and muscle mitochondria, proteome, and peptidome as it relates to overall beef quality. Our major objectives are to determine:the effect of supranutritional vitamin E supplementation (1000 IU for 225 days) and PAP level [low (< 50 mmHg) or high (≥ 50 mmHg)] on:muscle gene expression, mitochondrial function, histology, and proteome, as well as feed efficiency of steers during the finishing period;muscle mitochondrial proteome, early postmortem mitochondrial function, carcass characteristics, and meat quality of feedlot steers;the effect of vitamin E supplementation (1000 IU for 225 days) on the muscle-specific peptidome during industrial production (harvest-packing-shelf life) and its relation to meat quality development.
Project Methods
Following initial weighing, steers will be housed in one group pen with ad libitum access to long-stem grass hay and water overnight. The following day, steers will be weighed and blocked by body weight within low and high PAP scores. The heaviest 8 steers from the high and low PAP groups will be considered replicate 1, the next 8 heaviest from each PAP classification will be considered replicate 2, and the lightest 8 steers from each PAP classification will be considered replicate 3. Within a replicate, steers (n=16) will be stratified by body weight into one of two pens. Each pen will contain 4 low and 4 high PAP steers of similar body weight across both pens. Pens will then be randomly assigned to VE treatment (control-no supplemental VE or 110.0 IU of dl-α-tocopheryl acetate/kg DM). This process will be repeated for each replicate allowing for 3 control pens and 3 VE-supplemented pens, each containing 4 low and 4 high PAP steers per pen of similar body weights within a replicate. Each pen (15m x 43m) of 8 animals is equipped with 5 SmartFeed feeding systems, an automatic waterer, concrete bunk pad, and a metal roof (15m x 3m) covering approximately 7% of each pen. The experimental design will be split-plot design to evaluate the effects of PAP score (low <50 mmHg and high ≥50 mmHg) and VE supplementation (no supplemental VE or 110 IU of supplemental dl-α-tocopheryl acetate/kg DM) on the attributes of individual animals. In total, 4 treatment combinations will be used (n=12 steers/treatment): 1) no supplemental vitamin E (basal diet will contain approximately 25 IU of VE/kg DM) and low PAP (CON + LPAP); 2) 110 IU of supplemental VE/kg DM and low PAP (VET + LPAP); 3) no supplemental VE and high PAP (CON + HPAP); and 4) 110 IU of supplemental VE/kg DM and high PAP (VET + HPAP). The total feeding trial duration is targeted at 225 days but will be adjusted depending on body weight.All samples will be collected from each animal on day 0 and day 150 of VE feeding (75 days before slaughter to recover its effect on animal stress). Blood samples will be collected from jugular venipuncture. The supernatant samples will be immediately snap-frozen in liquid nitrogen and shipped to PD Zhai's lab at UConn, followed by storage at −80°C for no longer than 30 days prior to the total antioxidant capacity analysis. Muscle biopsy samples will be collected from the midpoint of the left longissimus muscle of each steer, distributed, and processed for different analyses. Muscle samples for mitochondrial function analysis will be immediately stored in an ice-cold BIOPs solution and analyzed by PD Zhai's team at collaborator Chicco's lab within 48 hours of collection. Muscle samples for gene expression and proteomics will be immediately snap-frozen in liquid nitrogen, while muscle samples for histology will be embedded in Tissue-Tek OCT and frozen in dry ice-cooled iso-pentane. RNA, histology, and proteomics samples will be shipped to UConn on dry ice and stored at −80°C until further analysis. Samples from six steers (n=6) of each treatment will be randomly selected for proteome analysis, resulting in a total of 48 muscle samples from 4 treatments (2 VE × 2 PAP levels) at 2 time points (day 0 and day 150 of the feeding trial).After the feeding trial, steers will be transported approximately 16 km to a USDA-inspected meat laboratory in the Global Food Innovation Center of the Department of Animal Sciences at Colorado State University. After resting time in the lairage, the animals will be stunned, bled, and dehided according to commercial practice under USDA inspection. Fresh longissimus muscle tissue will be collected from the left side of each carcass between the 12th and 13th rib at 1h and 24h postmortem and prepared for mitochondrial function and mitochondrial isolation (proteomic analysis). Histology samples will be collected from LL, ST, and PM at 1 h postmortem. After carcass evaluation at 24 h postmortem, 14 cm sections of LL, ST, and PM will be excised from each side of the carcasses and cut into four 3.5 cm steaks. The steaks will be individually vacuum packed and randomly assigned to be aged 1d, 7d, 14d, or 21d postmortem (2°C in the dark). All steaks will then be shipped to Zhai's lab overnight on ice. At the end of each aging period, the assigned steaks will be further cut into 1 cm and 2.5 cm steaks for biochemical analysis and quality evaluation, respectively. Steaks from the right side of carcasses will be used for measurements of instrumental tenderness and cooking loss, while steaks from the left side of carcasses will be sampled for measurements of instrumental color (shelf display), MB concentration, MetMB reducing activity, and lipid oxidation.For feed efficiency, growth performance, and carcass characteristics, steer will serve as the experimental unit as individual data will be collected. A split-plot will be used to evaluate the effects of PAP and diet on these attributes, where PAP (high or low), diet (control or VE), and their interactions will be fixed effects, and the random effect in the model will be block. For serum, each type of mitochondrial respiration, gene expression, and muscle histology during growth as well as each type of mitochondrial respiration at postmortem, steer will serve as the experimental unit as individual data were collected. A three-factor repeated measurement will be used to evaluate the effects of PAP, diet, and sampling time on these attributes, where PAP (high or low), diet (control or VE), sampling time (d0 or d150 during growth; 1 h or 24 h at early postmortem), and their interactions will be fixed effects, and the random effect in the model will be individual animal. For fiber type composition of each postmortem muscle, individual animal will serve as the experimental unit. A split-split-plot measurement will be used to evaluate the effects of PAP and diet on fiber type composition of each muscle (LL, PM, or ST), where PAP (high or low), diet (control or VE), and their interactions will be fixed effects, and the random effect in the model will be individual animal. For meat quality analysis of each muscle, individual 3.5-cm steak of each muscle will serve as the experimental unit. A split-split-plot measurement will be used to evaluate the effects of PAP, diet, and sampling time on a single attribute of each muscle, where PAP (high or low), diet (control or VE), aging time (1d, 7d, 14d, and 21d aging), and their interactions will be fixed effects, and the random effect in the model will be individual animal. For all above analyses, data analysis will be performed by R using the lme4 package as a mixed model, and the differences between least-square means (P < 0.05) will be determined by Tukey's multiple comparisons.Proteomic and peptidomic analysis: The spectrum count/abundance of identified proteins (for proteomics)/peptides (for peptidomics) will be outputted by Scaffold Q+S v5 and inputted into R for further statistical analysis using limma package. Briefly, after log2 transformation and median normalization, the identified proteins that have 50% missing values or more (more than half the observations) will be removed. A moderated t-test will be used for pairwise comparisons between treatments and feeding time points (or postmortem time). Benjamini-Hochberg multiple testing adjustment will be used to control the false discovery rate and control for multiple testing at P < 0.05. Log2 fold change threshold ±0.585 (1.5 times) will be applied for differentially abundant protein/peptide identification. Detailed functions of identified proteins or parent proteins of identified peptides will be annotated based on UniProt database with verification through literature search.