Feeding trials were conducted in two years to compare growth performance, carcass characteristics and quality of meat from beef cattle with 0, 50 or 75% Wagyu genetic influence. The cattle types used in the two years of the study were Continental crossbred steers (0% Wagyu), cattle with 50% Wagyu influence (Wagyu/Angus crossbreds) and 75% Wagyu influence (containing 25% from other European breeds). Cattle were housed in individual pens in the first year and in group pens in the second year. Cattle were fed a backgrounding diet containing 35% barley grain and 65% barley silage with protein, mineral and vitamin supplements until they weighed more than 380 kg, after which they were adapted to a finishing diet consisting of 80% barley and 20% barley silage with mineral and vitamin supplements. Cattle were weighed every 4 wk and at the end of the finishing period they were processed and carcass information was obtained. A three-rib section (10-12) was removed from 41 (year 1) and 44 (year 2) carcasses selected randomly from each group, and Warner-Bratzler shear force was determined. The year-by-cattle type interaction was significant for most parameters; thus all the data were also analysed by year using weight of cattle at initiation of the feeding trial as a covariate for the backgrounding and finishing phases of growth and using carcass weight and back fat depth as covariates for carcass and meat-quality parameters. Weight at the start of the trial influenced most growth parameters and age at slaughter. Continental crossbred steers had higher (P < 0.05) ADG than Wagyu crossbred cattle during the finishing phase (1.47 vs. 0.82 kg d(-1)) in year 1, but not in year 2. Carcass weights of 75% Wagyu crossbred cattle were lower (P < 0.05) than those of Continental crossbred steers in both years. Warner-Bratzler shear force values were less than 5.3 kg to shear cores of 19 mm diameter. However, 92% in year 1 and 71% in year 2 of the carcasses from Wagyu crossbred cattle graded Canada AAA, and contained more than a "small" amount of intramuscular fat. Thirty percent of the carcasses from Wagyu crossbred cattle in year 1 graded Canada Prime. Only 30% of Continental crossbred steers in year 1 and 10% in year 2 graded Canada AAA. Wagyu genetic influence enhanced marbling in beef cattle without loss in carcass size for the 50% Wagyu steers.

ABSTRACT The objective of the present study was to develop an immunohistochemical technique to locate preadipocytes in bovine muscle tissue. Preadipocyte factor‐1 (pref‐1) is a transmembrane protein that is part of the family of epidermal growth factor‐like repeat‐containing proteins that are involved in cell fate determination. Pref‐1 is highly expressed in preadipocytes, but expression is completely abolished during differentiation into an adipocyte. In the present study, reverse transcription polymerase chain reaction demonstrated that bovine intramuscular adipose tissue contains the three splice forms of pref‐1 (A, C2 and E). Western blots were used to confirm that the protein for pref‐1 was expressed in intramuscular adipose tissue. Polyclonal antibodies against pref‐1 were tested against a cell culture of bovine preadipocytes from an embryo source to confirm that the antibody would immunolocate bovine preadipocytes. The antibody was applied to sections of longissimus dorsi muscle from Charolais and Holstein cattle. Immunohistochemical results showed that pref‐1 is expressed in the perimycium near mature adipocytes and blood vessels. The pool size of preadipocytes appeared to be low. Previous reports, however, have demonstrated that preadipocytes are known to divide and/or migrate, providing a potentially endless source of adipogenic precursor cells.

Peroxisome proliferator-activated receptor γ(PPARγ) regulates adipogenesis and lipid metabolism-related gene transcripts. The role, however, of PPARγ in different adipose depots and muscle in Holstein and Charolais cattle is still unclear. We used 20 animals (10 from each breed) for semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) to measure PPARγ mRNA levels in subcutaneous (SC), perirenal (PR), omental (OM), and intramuscular (IM) adipose depots as well as longissimus muscle (MU). IM fat was dissected from muscle tissue in MU. Holstein were characterized by their higher OM (P < 0.01) and PR (P < 0.05) fat weights while the Charolais had a higher body weight (P < 0.001) and a larger longissimus muscle area (P < 0.001). The IM fat content and marbling scores tended to be higher in Holstein. No significant differences in PPARγ mRNA expression were observed between these two breeds for any tissue. In both breeds, MU PPARγ had the lowest expressed mRNA level (P < 0.05). In the IM fat depot, expression was higher (P < 0.05) than MU, but lower than the SC, PR, and OM fat depot PPARγ mRNA levels. Only OM PPARγ mRNA levels were higher (P < 0.05) than SC and PR in Charolais. To characterise the role of PPARγ mRNA in bovine adipogenesis, correlations were performed among PPARγ mRNA, carcass characteristics, and adipogenesis-related genes. Key words: Adipose tissue, muscle tissue, PPARγ, cattle

Meaningful estimates of diacylglycerol acyltransferase (EC 2.3.1.20) activity in different tissue samples require effective, unbiased methods of sample storage. Samples of the pars costalis diaphragmatis muscle (skirt muscle of the diaphragm) were obtained from 18- to 20-month-old cattle and assayed for microsomal protein content and diacylglycerol acyltransferase activity after having been stored under various conditions as dissected tissue or microsomes prepared from dissected tissue. There was relative enrichment of diacylglycerol acyltransferase specific activity (p<0.05) when samples prepared from the pars costalis diaphragmatis muscle were dehydrated and stored for 2 weeks, as compared to the control condition (in which the microsome fraction was prepared from fresh pars costalis diaphragmatis muscle and assayed immediately). The results suggested that dehydration was an effective method of storage for bovine muscle samples destined for estimation of the microsomal diacylglycerol acyltransferase activity. The dehydration approach for preparing samples for analysis of diacylglycerol acyltransferase activity might also prove useful to investigators who are interested in obtaining reliable estimates of the activity of other enzymes in tissue samples.

The relationship between hormone-sensitive lipase (HSL; EC 3.1.1.3) activity from pars costalis diaphragmatis (PCD) muscle (skirt muscle of diaphragm) and the lipid content of PCD and longissimus muscle was examined in a herd of Wagyu hybrid cattle. HSL activity from subcutaneous (SC) adipose tissue was examined as control for general fattening. When assayed as a cholesterol esterase (i.e. using cholesteryl oleate as a substrate), cytosolic HSL activity from both PCD muscle and SC adipose tissue displayed similar dependence on bulk substrate concentration. Significant positive correlations were found for PCD HSL specific activity and activity per g wet weight with both the total lipid content of PCD muscle (r=0.49, P<0.01) and longissimus muscle (r=0.41, P<0.01). As well, HSL specific activity and activity per g wet weight of PCD muscle showed significant negative correlations with the Canadian marbling score (an inverse scale; r=−0.42, P<0.01 and r=−0.40, P<0.05, respectively). In contrast, HSL activity of SC adipose tissue showed no significant correlation with any parameter examined including muscle lipid content and backfat depth. The results suggest HSL activity in bovine muscle is associated with the content of intramuscular (IM) fat, and may have predictive value for assessing the potential of cattle to deposit IM fat.

The activity of the triacylglycerol bioassembly enzyme, diacylglycerol acyltransferase (DGAT), was characterized in microsomal fractions prepared from bovine subcutaneous (SC) adipose, intramuscular (IM) adipose, and muscle (pars costalis diaphragmatis) tissue. The activity of DGAT was generally higher from SC adipose tissue than from IM adipose or muscle tissue. The characteristics of DGAT activity from the three bovine tissues resembled the activity characteristics observed in previous studies from various other organisms and tissues; the pH optimum was near neutrality, the activity was almost completely inhibited by pre-incubation with N-ethylmaleimide (NEM), and the enzyme accepted a broad range of acyl-CoAs and sn-1,2-diacylglycerols. In some aspects, the SC adipose tissue DGAT activity was different from the DGAT activity from the other two tissues. The SC adipose tissue DGAT activity was not as susceptible to inhibition by NEM as the enzymes from the two other tissue sources, and it exhibited increased specificity for substrates containing oleoyl moieties. The differences in DGAT properties between the three bovine tissues may account to some extent for the differences in the relative fatty acid composition and the positional distribution of fatty acids in triacylglycerol between bovine tissues. The observed differences in enzymatic properties also support recent biochemical and molecular genetic observations that imply the existence of multiple DGAT genes and/or isoforms.

The deposition of i.m. fat, or marbling, in cattle is recognized as a desirable carcass trait in North American beef grading schemes. In order to investigate the relationship between degree of marbling and fatty acid composition of whole bovine muscle, we extracted the total lipid from pars costalis diaphragmatis (PCD) (n = 23) and longissimus (n 36) muscles from Wagyu crossbred cattle that were assigned Canadian Grading Agency marbling scores ranging from 1 to 8 on an inverse 10-point scale (i.e., a score of 1 indicated "very abundant" marbling and a score of 10 would be assigned to a carcass "devoid" of marbling). Fatty acid methyl esters (FAME) of the total lipid and triacylglycerol fractions were resolved and quantified through GLC. Marbling scores were negatively associated with total lipid from both PCD (r = -.57, P < .01) and longissimus (r = -.80, P < .001). Differences between PCD and longissimus were found for almost all FAME studied from both lipid fractions, but no differences (P > .05) were seen when the monounsaturated:saturated fatty acid (MUFA/SFA) ratios were compared. Heifers had higher (P < .05) oleic acid content and lower (P < .05) palmi tic acid content in lipid extracted from both muscles, resulting in higher (P < .05) MUFA/SFA ratios than those for steers. The relative amount of myristic acid increased as the lipid content (total lipid and triacylglycerol) increased in either longissimus (r values from .48 to .55; n = 36; P < .01) or PCD muscles (r from .67 to .76; n = 23; P < .001). The relative amount of linoleic acid (cis-9, cis-12 isomer) from total lipid was negatively associated with all chemical measurements of lipid from the longissimus (r from -.52 to -.64; n = 36; P < .001) and PCD muscles (r from -.75 to -.85; n = 23; P < .001). This association was not significant (P > .1) for either muscle when linoleic acid from the triacylglycerol fraction was examined, suggesting the negative association between this fatty acid and lipid content was due to a dilution of membrane phospholipids with increasing triacylglycerol. Indices of fatty acid elongase activity, calculated from FAME data, implicated the balance between this enzyme activity and fatty acid synthase as a source of variation between animals displaying various degrees of marbling and worthy of further investigation to better understand the process of marbling fat deposition in beef cattle.

The development of a biochemical or molecular marker for predicting the propensity for cattle to deposit intramuscular (i.m.) fat would reduce the time required to improve for this trait. In the current study, diacylglycerol acyltransferase (DGAT, EC 2.3.1.20) activity was investigated as a potential biochemical marker for predicting the deposition of i.m. fat. The activities of DGAT were determined in microsomal fractions of subcutaneous (s.c.) adipose tissue, i.m. adipose tissue and muscle tissue sampled from Wagyu crossbred cattle at slaughter weight. The i.m. adipose and muscle tissues were dissected from the pars costalis diaphragmatis (p.c.d.) muscle. The lipid content (LC) of the p.c.d. muscle correlated with the LC of the longissimus muscle (r = 0.51; P < 0.05). There was an inverse correlation between p.c.d. i.m. adipose tissue DGAT specific activity and LC of the p.c.d. (r = −0.50; P < 0.05) suggesting that DGAT activity was down-regulated as i.m. adipose tissue became filled. There was a strong correlation (r = 0.74; P < 0.001) between DGAT total activity in i.m. adipose tissue and muscle DGAT total activity, suggesting DGAT activity is coordinately regulated in these two tissues. Key words: Bovine, intramuscular fat, diacylglycerol acyltransferase, Bos taurus

A study was conducted to determine differences in length of intestinal villi and crypts and activity of mucosal carbohydrases of several breeds of beef cattle. Tissue samples, from three locations in the small intestine, were obtained from heifers and steers of Wagyu (W) × Angus (W × A), W × Hereford (W × H) and W × Holstein (W × O) crossbred cattle and from Holstein, Parthenais and Piedmontese steers at slaughter. In W × A cattle villi were shorter and crypts were longer (P < 0.05) than in cattle of other breeds. Mucosal protein concentration was highest (P < 0.05) in the mid- and distal intestinal regions in the W × A cattle. Maltase activity was highest (P < 0.05) for W × H heifers and least (P < 0.05) for W × A heifers in the proximal intestinal region. Holstein and Piedmontese steers had the highest ADG, and Holstein steers had the highest lactase activity in the mid-intestinal region. Lactase activity in the mid-intestinal region appeared to be weakly associated with ADG of cattle (r = 0.454, P = 0.015, n = 28). These data suggest that differences due to breed of cattle exist in villus and crypt lengths and mucosal carbohydrase activity; however, the influence of these intestinal parameters on cattle performance may be relatively small. Key words: Intestinal villi, crypt, cellobioase, maltase, lactase, beef cattle

1. The vascular anatomy of the bovine omasal lamina permitted perfusion of a discrete area of the tissue. As occurs in vivo, oxygen was provided through the vascular system, while the luminal sides of the tissue could be kept in an anaerobic environment, thus allowing study of foregut tissue metabolism under physiologically realistic conditions.2. O2consumption of perfused leaves in the presence of anaerobic buffer was 64.9 and 73.5 nmol O2/mg dry weight per h in Expts 1 and 2 respectively, and was elevated (P< 0.05) when the lumen side of the tissue was exposed to an atmosphere of nitrogen gas.3. In Expt 1, the rate of O2consumption was increased (P< 0.01) by 35% as a result of suspension of a boiled preparation of rumen micro-organisms and particles (< 1 mm) in the anaerobic lumen buffer. Replacement of the boiled preparation with an unboiled suspension increased O2consumption further by 11 %, but this was not statistically significant (P> 0.05).4. In Expt 2, sequential addition of the following substrates or preparations to the lumen chambers all resulted in stepwise increases (P< 0.05) in O2consumption; 8 mM-butyrate, boiled rumen micro-organisms and particles and, finally, unboiled rumen micro-organisms and particles.5. Identities of the heat-labile and heat-stable components of the microbial and particle suspensions that caused enhancement of O2removal across the perfused tissue are discussed.

Oxygen uptake by the perfused omasal leaf was measured in the presence of mixed rumen microbial suspension which was kept anaerobically in the lumen side of a perfusion chamber. The addition of DL-3-hydroxybutyrate at 2, 4 and 8 mM concentrations to the microbial suspension increased oxygen withdrawal by 4.4, 11.1 and 15.5%, respectively. It is proposed that electron transfer through the coupled D(−)-3-hydroxybutyrate-acetoacetate reaction from rumen microorganisms to rumen epithelium yield energy for the latter tissue. Key words: Omasal leaf, oxygen uptake, microbes, DL-3-hydroxybutyrate