Effect of nutritional factors on biochemical, structural and metabolic characteristics of muscles in ruminants, consequences on dietetic value and sensorial qualities of meat.

Ruminant meat is an important source of nutrients and is also of high sensory value. However, the importance and nature of these characteristics depend on ruminant nutrition. The first part of this review is focused on biochemical and dietetic value of this meat. It offers a panel of quantitative and qualitative contributions, especially through its fatty acids characteristics. Since saturated and trans-monounsaturated fatty acids are considered as harmful to human health, their amount in muscles can be reduced by increasing the proportions of dietary polyunsaturated fatty acids (PUFA) absorbed by the animals. On the contrary, some fatty acids (n-6 and n-3 PUFA, conjugated linoleic acid) specifically incorporated in muscle tissues would play a favourable role in the prevention or reduction of major diseases in human (cancers, atherosclerosis, obesity) and therefore be recommended. The second part of this review treats different aspects of the sensorial qualities of meat. Skeletal muscle structure and its biochemical components influence muscle transformation to meat and sensorial qualities including tenderness, colour, flavour and juiciness. This paper shows how nutrition can influence, in ruminants, metabolic activity as well as muscle structure and composition, and thereby affect meat quality.

Age-related changes and location of types I, III, XII and XIV collagen during development of skeletal muscles from genetically different animals.

The ontogenesis of total collagen and of different collagen types was studied in four muscle types from genetically different cattle. Hydroxyproline content was 1.2-fold higher in muscles from cross-bred foetuses with normal muscle growth compared to those of the other genetic types (pure bred with different growth rates, double-muscled breed). A similar tendency was observed for type III collagen content. In all muscles of each animal studied, type XII and XIV collagens were colocated in perimysium. Immunolabelling obtained for type XII collagen was higher during foetal life than after birth, while for type XIV collagen, the opposite result was obtained. Whatever the muscle studied, but especially in semitendinosus muscle, during the foetal and the post-natal period until 15 months of age, immunolabelling with antibody anti-type XIV collagen tended to be more intense in muscles of animals from fathers selected for a low muscle growth capacity compared to those from fathers selected for a high muscle growth capacity. In conclusion, this study shows, that during foetal life, selection according to muscle growth capacity has no significant effect on the contents of total hydroxyproline or type III collagen, but minor effects on collagen localization.

Effects of muscle type, castration, age, and compensatory growth rate on androgen receptor mRNA expression in bovine skeletal muscle.

The effect of testosterone on sexual dimorphism is evident by differential growth of forelimb and neck muscles in bulls and steers. Divergent hormone sensitivites may account for the differential growth rates of individual muscles. Therefore, the objective of this study was to compare androgen receptor (AR) expression in three different muscles of bulls and steers at various ages and growth rates. Thirty Montbéliard bulls and 30 steers were assigned to four slaughter age groups. Four or five animals of each sex were slaughtered at 4 and 8 mo of age. Animals in the remaining two slaughter groups (12 and 16 mo) were divided into groups of either restricted (R) or ad libitum (AL) access to feed. Five animals of each sex and diet were slaughtered at the end of the restricted intake period at 12 mo of age. To simulate compensatory growth, the remaining animals (R and AL) were allowed ad libitum access to feed until slaughter at 16 mo of age. Total RNA was extracted from samples of semitendinosus (ST), triceps brachii (TB), and splenius (SP) muscles. Androgen receptor mRNA was quantified in 200-ng total RNA preparations using an internally standardized reverse transcription (RT) PCR assay. Data were analyzed using 18S ribosomal RNA concentrations as a covariable. Steers had higher AR mRNA levels per RNA unit than bulls (P < .01). Androgen receptor mRNA levels differed between muscles (P < .05), with lowest expression in the SP. The pattern of AR expression differed (P < .05) for each muscle with increasing age. Between 4 and 12 mo of age, AR mRNA levels increased (P < .05) in SP but remained unchanged in the ST and TB. Feeding regimen had no effect on muscle AR expression, but steers exhibiting compensatory growth had higher AR mRNA levels than AL steers (P < .01) or bulls (P < .01). Our results show that AR expression is muscle-specific and may be modulated by circulating testicular hormones. These data suggest that the regulation of AR expression may be linked to allometric muscle growth patterns in cattle and compensatory gain in steers.

Specificity of different anti-myosin heavy chain antibodies in bovine muscle.

Muscle fibre types play an important role in bovine meat tenderness, and conditions the speed of ageing. Two generations of myotubes appear during foetal life. Their differentiation results in several types of fibres differing in their contractile and metabolic properties, namely type I fibres and type II fibres (IIA and IIB). Fibre types were identified using antibodies by immunohistochemistry and western-blotting techniques. The aim was to test different antibodies on foetal and adult muscles by these two techniques. Ten monoclonal antibodies reported to be specific to different myosin heavy chain (MHC) isoforms in other species were tested. Only three antibodies were selected: BF.35, specific for I and IIa MHCs, F1.652 specific for foetal MHC and SC.71 an anti II MHCs (IIa and IIb) antibody. The use of these antibodies will enable a more precise characterisation of muscle fibres and will have applications in the study of bovine muscle ontogenesis.

Characterisation and location of insulin-like-growth factor (IGF) receptors in the foetal bovine Semitendinosus muscle.

We characterised IGFI and IGFII receptors and located them in bovine muscle during foetal growth. Semitendinosus muscle samples were taken from foetuses ranging from 80 to 270 days post-conception. The relative affinities of 125I-IGFII and 125I-IGFI mark the presence of typical type II and type I receptors in foetal muscle membranes. IGFII-specific binding is consistently five times greater than that of IGFI. The patterns of 125I-IGFII- and 125I-IGFI-specific binding are similar. They increase up to 110 and 170 days post-conception, respectively (P < 0.05); thereafter, they decrease (P < 0.05). This decrease was due to a fall in the number of receptors without any change in affinity. At the adult stage the specific binding of both the 125I-IGF is very low. In foetal muscle, type II receptors are located both in the muscle bundles and in the connective tissue while type I receptors are only located in the muscle bundles.

Changes in the metabolic and contractile characteristics of muscle in male cattle between 10 and 16 months of age.

Samples of semitendinosus muscle from 28 male cattle (18 Salers and 10 Limousins) were taken at 10 months (biopsy) and at 16 months of age (at slaughter). The animals had received the same diet and were slaughtered after the same duration of fattening. The activities of isocitrate dehydrogenase and lactate dehydrogenase were measured in the muscle samples. The five lactate dehydrogenase isoenzymes were separated by electrophoresis under non-denaturing conditions and assayed by densitometry. Fibres were identified by histochemistry by myofibrillar ATPase and succinate dehydrogenase activities as SO (slow oxidative), FOG (fast oxidative glycolytic) or FG (fast glycolytic), and by immunohistochemistry by their reaction to monoclonal antibodies specific to slow and fast myosin heavy chain reactions in I, IIC, IIA, IIAB and IIB type fibres. The isocitrate dehydrogenase activity was not modified between 10 and 16 months of age; the lactate dehydrogenase activity decreased and was correlated with an increase in the proportion of the H isozyme to the detriment of the proportion of the M form. This period was characterized by an increase in fibre size, increased expression of MHC IIa, resulting in more IIA fibres, less IIB fibres, and an increase in the percentage of type IIAB fibres, however the proportions of SO, FOG and FG, when analysed statistically, were not modified between 10 and 16 months of age.

Age-related changes and location of type I, III, IV, V and VI collagens during development of four foetal skeletal muscles of double-muscled and normal bovine animals.

The aim of this work was to study the variability in the quantities of hydroxyproline, of type I and III collagens and in the location of types, I, III, IV, V, VI in four muscles of normal and double-muscled (DM) cattle. Samples were collected from foetuses at different ages post-conception. Both in the two genetic types and in muscles, from 110 days, types I, III, V, VI were located in perymisium and types I, IV, V, VI in endomysium. The amounts of hydroxyproline and of type I collagen increased from 150 to 180 or 230 days then decreased up to 260 days, with a trend to lower quantities in muscles of DM animals. Depending on the muscle and of the genetic type, amounts of type III, or changed as those of type I, or remained stable. Whatever the genetic type, at the end of gestation, proportions of type I and III in the total collagen are not identical in the four muscles, differences between muscles being particularly marked for type III, CT and MA muscles being the richer in this type. In addition, these two muscles contained less type III in DM animals than in normal ones.

Comparison of foetal metabolic differentiation in three cattle muscles.

Metabolic differentiation of Semitendinosus (ST), Cutaneus trunci (CT) and Masseter (MA) in cattle foetuses aged from 110 to 260 days was studied by measuring isocitrate dehydrogenase (ICDH, oxidative) and lactate dehydrogenase (LDH, glycolytic) activities. The five LDH isoenzymes were separated by electrophoresis and assayed by densitometry. ICDH activity increased from 210 days onwards in the three muscles but more intensively in MA (oxidative). LDH activity increased from 170 days onwards in ST, 180 days onwards in CT and only from 210 days onwards in MA and was higher in the glycolytic muscles (ST and CT). The proportion of the LDH-M subunit increased during foetal life in glycolytic muscles. At 110 days, it was higher in CT, intermediate in ST and lower in MA. These results show that 1) metabolic differentiation of bovine muscle begins during the last third of foetal life and 2) the proportion of the LDH-M subunit seems to be related to the contractile type of adult muscle from the first stages of foetal life.

Insulin-like growth factor II (IGF-II) mRNA expression during skeletal muscle development of double-muscled and normal bovine foetuses.

We investigated the IGF-II gene expression in developing Semitendinosus muscle in foetal normal and double-muscled cattle. Samples from normal and double-muscled foetuses ranging from 90 to 210 d post-conception were collected and total RNA extracted. Northern blot analysis was performed using the human IGF-II cDNA probe. Five IGF-II transcripts, 5.1, 4.4, 3.7, 2.6 and 1.7 kb, were detected in muscle samples. Throughout gestation, all transcripts, except for the 5.1 kb one, decreased similarly in both genetic types. In double-muscled foetuses, the amount of the 5.1 kb transcript was higher than those of the other transcripts and its expression remained stable throughout the gestational stages analysed. These results indicated that the regulation of IGF-II gene transcription was distinct in both genetic types. The IGF-II foetal plasma concentrations increased throughout gestation. In bovine foetuses, the first muscle cell differentiation was concomitant with a high autocrine IGF-II mRNA expression and low plasma IGF-II levels in both genetic types. The second step of muscle cell differentiation was associated with high IGF-II plasma concentrations and the autocrine expression of IGF-II was reduced.

Fat partitioning and biochemical characteristics of fatty tissues in relation to plasma metabolites and hormones in normal and double-muscled young growing bulls.

Plasma metabolites and hormones, and the biochemical characteristics of four fatty tissues (FT) were studied in two groups of six normal (N) or six double-muscled (DM) Belgian Blue young growing bulls fed the same net energy amount at the same live weight and slaughtered at 10 months of age. Average daily gain and feed efficiency did not significantly differ between the two groups. However, the DM bulls exhibited a higher proportion of muscles (+22%, P < 0.01) and a reduced proportion of fat (-49%, P < 0.01) mainly in the subcutaneous FT (-80%, P < 0.05). Triiodothyronine, insulin and glucose plasma concentrations tended to be lower in DM bulls (-24%, P < 0.02; -27%, P = 0.14; -7%, P = 0.06, respectively) and were positively related to the higher fat development in N bulls. From the results of total protein. DNA, lipid and TG contents of FT, it appeared that a reduction in fat storage per fat cell (hypotrophy) or a reduction in total fat cell number (hypoplasia) could explain, in DM bulls, two-thirds and one-third of the reduction of perirenal and subcutaneous FT weights, respectively, as compared to N bulls. In contrast, either hypotrophy or hypoplasia was the main cause of omental or intermuscular FT weight reduction in DM animals.

Regulation of bovine satellite cell proliferation and differentiation by insulin and triiodothyronine.

Satellite cells activity contributes to postnatal muscle growth. Herein, we have studied the respective influence of insulin and triiodothyronine (T3) on the proliferation and differentiation of primary bovine satellite cells isolated from Semitendinosus muscle of Montbéliard steers. Under basal conditions, satellite cells proliferated until the fifth day of culture, began to fuse into myotubes and expressed differentiation markers such as connectin, myogenin, and myosin heavy chain (MHC) isoforms. Insulin behaved as an effective mitogen. Moreover, it promoted extensive myotube formation and enhanced differentiation as shown by an increase in the accumulation of differentiation markers. Maximal differentiation occurred with insulin physiological range concentrations. A delay in the stimulation of differentiation was registered with a high dose that promoted maximal proliferation. Conversely, T3 decreased cell proliferation in a dose-dependent manner. In addition, fusion and biochemical differentiation (accumulation of connectin, MyoD1, myogenin, and myosin heavy chain isoforms) were also enhanced. Bovine satellite cells seemed to respond differentially to insulin and T3 for proliferation. Interestingly, both hormones displayed a myogenic influence. Our observations suggest that both hormones could influence bovine satellite cells in vivo and contribute to the regulation of postnatal muscle growth.

Contractile differentiation of foetal cattle muscles: intermuscular variability.

off actile differentiation was studied in six foetal muscles exhibiting different contractile characteristics in adult cattle: the Masseter, Diaphragma, Biceps femoris, Longissimus thoracis, Semitendinosus and Cutaneus trunci. These muscles were excised from foetuses aged 60-260 days. Fibre types were identified by immunohistochemistry using three monoclonal antibodies raised against types 1, 2a, 2b (or 2x) and foetal myosin heavy chains. The different myosin isoforms were also separated by electrophoresis, identified by immunoblotting and quantified by ELISA. At least two generations of cells were observed in all the muscles studied. The primary, early differentiated one, gave rise to type II fibres in Cutaneus trunci and type I fibres in all remaining muscles. The secondary generation of cells differentiated later than the first generation of cells. Its pattern of differentiation was more complex in particular from 150 to 210 days. It formed slow fibres in slow adult muscles, fast fibres in fast adult muscles and both types in mixed muscles. Precocity of differentiation was muscle-type dependent and related to muscle function at birth.

Electrophoretic separation of bovine muscle myosin heavy chain isoforms.

This study concerns the definition of the optimum conditions for separation of adult and developmental myosin heavy chain (MHC) isoforms in bovine muscle. The various techniques published do not result in good separation of the MHC in this species. The trials carried out concerned the concentration of acrylamide and N, N'-methylene-bis-acrylamide, and more particularly the concentration of Tris in the separating gel. The finding was that analysis of adult isoforms and developmental isoforms require different conditions. A acrylamide gradient of 3.5-10% with 200 mM Tris pH 8.8 gives good resolution for adult isoforms. Under these conditions 3 fast adult isoforms are revealed. However, study of MHC isoforms throughout foetal life in bovines is complex, and requires the combined use of more than one gel (gradient 3.5-10% at 200 mM Tris and gradient 3.5-10% at 250 mM Tris).

Effect of the type of diet on muscle characteristics and meat palatability of growing Salers bulls.

The effect of the type of diet (hay vs grass silage) on body composition and characteristics and palatability of semitendinosus (ST) and longissimus thoracis (LT) muscles of 16 month old Salers bulls fed at the same energy levels were studied. Animals fed hay had a lower daily weight gain and carcass weight and were leaner. There were no significant differences in the proportions of fibre types in the ST or LT due to diet. ST muscle of hay fed animals had a lower oxidative metabolism, but contained similar amounts of total and type I collagen and greater amounts, and proportions of soluble collagen and of type III collagen, than those of animals fed grass silage. ST muscles of hay-fed animals were more tender than those of silage-fed animals.

Age-related changes and location of type I, III and IV collagens during skeletal muscle development of double-muscled and normal bovine foetuses.

The aim of this study was to investigate age-related changes in collagen content in muscles of normal and double-muscled (DM) bovine foetuses. Psoas major (PM) and triceps brachii (TB) muscles were collected from foetuses at 110 to 260 days post-conception (p.c.), frozen and powdered. Cyanogen bromide (CNBr) digestion and hydroxyproline measurements were carried out on the powder. CNBr peptides underwent SDS-polyacrylamide gel electrophoresis, and type I and III collagen relative variations were measured by densitometric analysis. Type I and III procollagen mRNA were located by in situ hybridization and types I, III and IV collagen located by indirect immunofluorescence. Although there was no significant difference between normal and DM animals in the amounts of collagen (except in PM muscle at 180 and 230 days p.c.), there was nevertheless a tendency to lower collagen content in muscles of DM animals. Amounts of hydroxyproline in PM and TB muscles from 110 to 230 days p.c. increased two- and threefold to 8.5 micrograms per mg of dry matter (d.m.) and 12 micrograms per mg d.m., respectively, and then decreased up to 260 days p.c., when they were twice as high in TB muscle (9.1 micrograms per mg d.m.) as in PM (4.5 micrograms per mg d.m.). The same difference in hydroxyproline levels was observed between normal adult PM and TB muscles. These variations were explained, mainly, by those of type I. In foetal muscle, in both genotypes and as in adult muscle, perimysium was types I and III, and endomysium type I, III and IV. Procollagen type I and III mRNA were located in perimysium. In conclusion, this study emphasizes that at the end of foetal life, collagen content is representative of what it will be in adult in muscle of both normal and DM animals.

Influences of the method of housing bulls on their body composition and muscle fibre types.

The influence of the type of housing (loose or tying-type) was studied in relation to body composition and muscular characteristics of bulls. 34 young bulls (18 Salers and 16 Limousins) were divided into two groups with equal breed representation and equal mean growth rates, to 10 months of age. One group was housed in tying-type housing (short stalls 1·8m(2) per animal) and the other in loose housing (6·5m(2) per animal). Samples of semitendinosus (ST) and longissimus thoracis (LT) muscle were taken at slaughter at 16 months. The lactate dehydrogenase (LDH) and isocitrate dehydrogenase (ICDH) activities were measured. The proportion of isoforms LDH-M and LDH-H were determined for each muscle. Total collagen content and solubility were measured for ST alone. Fibres were classified by ATPase myofibrillar and succinate dehydrogenase activities into SO (slow oxidative), FOG (fast oxidative glycolytic) or FG (fast glycolytic), and by immunohistochemistry by reaction with monoclonal antibodies specific to slow and fast myosin heavy chain reactions into I, IIC, IIA, IIAB and IIB fibres. Compared with animals in tying-type housing, animals in loose housing presented the following: fewer carcass adipose deposits (p<0·01); for ST muscle, more collagen (p<0·01) and a reduced glycolytic metabolism, as indicated by lower LDH activity (p<0·10) and a lower proportion of FG fibres (p<0·10). In both ST and LT muscles, loose housing resulted in an increased percentage of IIC fibres (p<0·05) but did not alter the proportions of fast fibres. Modifications in ST alone were increased percentage of IIA fibres (p<0·10) and IIAB fibres (p<0·01) and a lower percentage of IIB fibres (p<0·01). Thus, for a given rate of growth, the type of housing (loose or tying-type) influenced contractile characteristics, especially in muscles involved in movement.

Regional variations of muscle fibre characteristic in m. semitendinosus of growing cattle.

The objectives of the present study were to elaborate an intra-muscular profile of metabolic enzyme equipment, contractile and morphometric features along the longitudinal axis of m. semitendinosus at various ages throughout the growth phase. Thirty-seven male Montbéliard cattle, about half of them castrated, were representatively allocated to various slaughter dates, scheduled at 4, 8, 12 and 16 months of age. Samples were collected from proximal, medial and distal locations of m. semitendinosus. Isocitrate dehydrogenase (aerobic metabolism) and lactate dehydrogenase (anaerobic metabolism) were measured spectrophotometrically. Contractile muscle type was classified by quantification of myosin heavy chain I isoform proportion using the sensitive enzyme-linked immunosorbent assay. Mean muscle fibre area was obtained on histologically-stained cross-sections utilizing an image analysis system. Our results indicate the existence of a regular intra-muscular pattern of muscle fibre traits along the length of m. semitendinosus, with decreasing glycolytic activities and concomitantly an increase in oxidative capacity towards the distal extremity. The metabolic characteristics were in good agreement with decreasing cross-sectional muscle fibre areas and the slow myosin heavy chain I isoform proportion becoming gradually more abundant from proximal to distal regions of the muscle. Moreover, the observed gradient was found to be closely related to age and diminished with advanced physiological maturity. At the final slaughter age (16 months) no differences among the distinct portions were detected, m. semitendinosus was longitudinally homogeneous in all the characteristics studied.

Comparative study of metabolic differentiation of foetal muscle in normal and double-muscled cattle.

Studies of the contractile differentiation of foetal muscles in normal and double-muscled cattle have shown that double-muscled foetal muscles display comparative delayed contractile differentiation. We compared the metabolic differentiation of foetal muscles in normal and double-muscled cattle, for which no data were available. The semitendinosus (ST) muscle was excised from normal and double-muscled cattle foetuses aged 110-260 days. Isocitrate dehydrogenase (ICDH) and lactate dehydrogenase (LDH) activities, respectively representative of aerobic oxidative and anaerobic glycolytic metabolism, were assayed. The five LDH isozymes were separated by electrophoresis under non-denaturing conditions and assayed by densitometry. ICDH activity was weaker in the ST of double-muscled foetuses than in that of normal ones throughout gestation, and it increased later in the double-muscled foetuses than in normal ones. In contrast, the LDH activity increased at 180 days in both. It was greater in the double-muscled beyond that time. The proportion of LDH-M subunit was higher at all foetal ages in the double-muscled. Overall, these results show that: (1) the metabolic differentiation of muscle fibre occurs during the last third of foetal growth in both normal and double-muscled cattle; and (2) the differentiation of aerobic oxidative metabolism is delayed in double-muscled foetuses compared with normal ones.

Comparison of the foetal development of muscle in normal and double-muscled cattle.

Muscle differentiation was studied in foetal Semitendinosus muscle from normal cattle and those with the 'culard' gene of muscular hypertrophy sampled at 90, 130, 170 and 210 days of foetal life. The different fibre types were characterized by immunohistochemistry with antibodies specific to different isoforms of myosin heavy chains. The isoforms were separated by electrophoresis, identified by immunoblotting and quantified by ELISA. In double-muscled animals, there was a slower rate of differentiation in the first generation of cells, most markedly apparent at 90 days. At 130 days, differentiation was retarded mainly in the second generation, while at 170 days there was no longer any difference between the two animal types in the differentiation of first generation cells, which were totally slow in both. At the same stage however, type IIC fibres in double-muscled animals were much slower in appearing and continued to be so at 210 days, albeit to a lesser extent. These findings show that differentiation of the muscle fibres occurs at a slower rate in double-muscled foetuses particularly during the first two-thirds of foetal life.

Enzyme-linked immunosorbent assay for myosin heavy chains in the horse.

The content in slow and fast myosin heavy chains (MHC 1 and MHC 2) of 5 equine muscles was determined using an enzyme-linked immunosorbent assay. The results obtained with this immunoenzymatic method were compared with complementary techniques: electrophoresis and immunohistochemistry. Slices of masseter, diaphragm, tensor faciae latae, semitendinosus and cutaneus trunci were obtained from a 12-year-old saddle horse after slaughter. Muscular proteins were specifically extracted to be analysed by ELISA. The technique used 2 complimentary monoclonal antibodies (MAb). MAb 1 was prepared from a human atrium specimen that reacted specifically against MHC 1. Mab 2 was prepared from myosin of rabbit psoas muscle and reacted against MHC 2. The masseter muscle contained solely MHC 1 (100%) and this was confirmed by electrophoresis and immunohistochemistry. By contrast, the cutaneus trunci was very poor in MHC 1 (1.3%) and was entirely composed of MHC 2 (98.7%) which was confirmed by the other techniques. The diaphragm, tensor fasciae latae and semitendinosus contained 89, 40 and 2% of MHC 1, respectively. It was concluded that this ELISA method made it possible to measure a wide range of MHC contents in equine muscles with a good reproducibility. The results were consistent with those of the other fibre typing techniques. Moreover, this immunoenzymatic method is less time consuming than histological techniques and therefore offers new perspectives for muscle fibre typing in the horse.