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Histochemical and structural characteristics were investigated in Gastrocnemius pars interna (GN) and Iliofiburalis (IF) limb muscles of Rhea americana. The average myofibre area cross-section was greater in GN than IF muscle (p

Postmortem (PM) meat tenderization is a complex biochemical process that involves multiple endogenous proteolytic enzymes and that is not completely understood. Cell death by apoptosis is recently proposed as a novel mechanism in this process. The signification of the caspases implication in muscle proteins degradation is that the cells are dying through apoptosis, a major cell death programme. Our purpose was therefore to verify this statement using different well known hallmarks of the apoptotic process including cell shrinkage, phosphatidylserine (PS) externalization. Cells start to shrink few minutes after animal death and reach a maximum shrinkage about 24 h postmortem while they showed the typical rounding appearance of apoptotic cells. Externalization of PS is detectable within 1 h postmortem and increased gradually with time. The present work is therefore the first to provide direct evidence supporting the onset of apoptosis in post-mortem muscle. The present findings should lead us to reconsider as a whole, the mechanisms contributing of the development of meat qualities (tenderness, flavor, juiciness …) by a detailed analysis of the apoptosis associated biochemical and physiological modifications taking place within the cells.

To investigate the soft flesh problem in rainbow trout, the dynamics of water in postmortem white muscle were studied by magnetic resonance imaging, histology, and rigor index (θr) measurements. At 24-h postmortem, the trout identified as having the soft flesh problem (θr≥ 40°) differed from the others (0°≤θr≤ 20°) by longer transverse relaxation time (T2≥ 45 ms) and smaller diffusion anisotropy (DA ≤ 1.3), which proved to be the most relevant nuclear magnetic resonance parameter for an early diagnosis of the soft flesh problem. To interpret these results, we hypothesized an early and exacerbated tenderization phenomenon because of protein denatur-ation, inducing important modifications in fine connections that anchor the 3-dimensional structure of the tissue. The incidence of 3 slaughtering procedures on rigor were tested, and no differences were detected for trout presenting the soft flesh problem.

A magnetic resonance imaging technique based on susceptibility-induced contrast was used to visualise the spatial distribution of connective tissue in meat. Magnetic resonance imaging of bovine meat samples was carried out with a high-field 4.7 T imager. Magnetic resonance images obtained with spin-echo and gradient-echo sequences were compared to elucidate the role of connective tissue in the additional signal losses observed in the gradient-echo images. maps were reconstructed from the multiple gradient-echo images, which provide quantitative information. Comparison with histological pictures indicates that these maps exhibit the overall organisation of the primary perimysium at the scale of the whole muscle. The distinct perimysial organisation shown between the Gluteo biceps and Pectoralis profundis muscles illustrates the potential of magnetic resonance imaging for characterising the muscle connective tissue structure. © 2000 Society of Chemical Industry

International audience; The thermal denaturation of proteins in skeletal muscle was studied and characterised for the first time taking into account the in situ metabolic and contractile fibre types. From serial histological sections, collagen, elastin, various type I, IIa and IIx fibres and type I-IIa and IIa-IIx hybrids were identified by immunohistofluorescence. Histological sections were incubated in buffer solutions at increasing temperatures (40, 50, 60, 70 and 80 degrees C). Protein secondary structure was investigated by synchrotron radiation FT-IR microspectroscopy on connective tissue and in muscle fibres rigorously identified for sub-type. Whatever the target protein components, increasing temperature resulted in a decrease in alpha-helix secondary structure and an increase in beta-sheet structure. This phenomenon was more pronounced for intracellular proteins than for connective tissue. Although hybrid fibres were generally somewhat less sensitive to unfolding than the pure types, the amplitude of the thermal denaturation of intracellular proteins was practically independent of fibre type.

A histological and ultrastructural study was conducted to characterize changes in beef muscle structure after heating. Pieces of rectus abdominis muscle were heated at 100 degrees C over varying time frames from 15 min to 60 min and at 270 degrees C for 1 min; samples were then prepared for optical and transmission electron microscopy. After 15 min of heating, at 100 degrees C, a lateral shrinkage in fibre of 48% and an increase in gaps between the myofibrillar masses of 27% was noted. No more significant evolution was observed as heating duration escalated. The ultrastructure showed strong myofibril to sarcolemma detachments in which granular aggregates, coming in part from myofibrillar proteins, are stored. Neighbouring muscle Fibres showed strong heterogeneity in morphological behaviour after thermal treatment, suggesting that differences in composition and structure of the cytoskeleton proteins in the different fibres can cause denaturation/shrinkage of the proteins at different times along the timescale of microstructural changes during heating. Short heating at high temperatures expanded the gaps between myofibrillar mass, but the overall changes in the ultrastructure were similar to those obtained when heating at 100 degrees C.

International audience; An accurate characterisation of muscle fibres is essential for studying muscle plasticity. During some transient events such as ageing, myogenesis, physical activity or conversion of muscle to meat, the morphological parameters and/or the fibre type distribution may change. Nowadays, this information is generally obtained using immunohistology techniques, but these analyses are acknowledged to be laborious and time-consuming. In fact, each myofibre, from thousands, must be measured individually and its expression profile in response to different anti-myosin antibodies must be established step by step. In this paper, we describe a new histological approach using double-labelling (laminin, myosin) serial sections, fluorescence microscopy visualisation and, finally, semi-automatic image analysis. The goal of the study was to propose a tool allowing faster fibre type characterisation, including the identification of hybrid fibres from pure ones. The steps in the image processing prone to subjectivity have been fully automated. On the other hand, the expert retained control of all image analysis procedures requiring visual diagnosis. The tool that we developed with the Visilog software allowed a rapid and objective fibre typing and morphometric characterisation of two different bovine muscles. The results were in agreement with our previous histological and densitometric assays. The method and the tool proved to be potentially more efficient than other techniques used in our institute or described in the literature. A more global evaluation will be considered in other laboratories as well as on other animal species.

International audience; Several reports suggested the activation of caspases in postmortem muscle implicating the onset of a caspase-dependent cell death process after animal bleeding. It has been further well established that apoptosis and necrosis are the two major cell death pathways. The questions addressed in the present work were as follows: (a) in postmortem muscle, do cells die as in vivo? and (b) if so, by which dying process this goal is achieved? Selected hallmarks of apoptosis (phosphatidylserine externalization (PS), cell shrinkage, actin degradation) were analyzed in postmortem rat muscles and compared to usual cell behavior in apoptotic and necrotic processes. Results presented clearly demonstrate a rapid PS externalization and cell shrinkage extending during the first 24 h postexsanguination together with a progressive degradation of cytoskeletal and thin filaments of actin. It was therefore concluded that, in postmortem muscle, cells commit suicide soon after animal bleeding through apoptosis.

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International audience; A histological and ultrastructural study was conducted to characterize changes in the muscle fibre structure of three fresh sausage preparations, depending on meat composition, sodium chloride (NaCl) and potassium lactate (K-lactate) contents. After addition of 0.8% and 1.6% NaCl, 65% and 51%, respectively, of the area observed showed well preserved fibres (histological data). The altered regions presented a large disorganization of the myofilaments and a solubilization of the sarcolemma and of the Z lines. K-lactate addition had no marked effects on meat structure. The preparation containing some sheep meat was more sensitive to salt than the others containing only bovine meat. The level of alteration was much lower than those obtained in pork meat in another study. Technological conditions used to modify the internal muscle fibre structure during sausage processing depend on the species used. Therefore, the classification of the sausage preparation to "meat preparation" or "meat product" under the EU regulation (EC) No. 853/2004 (which assign meat preparations to meat if the product has undergone a process insufficient to modify the internal muscle fibre structure of the meat) must be systematically controlled when changing the meat sausage composition.

International audience; In meat, no detailed studies on the intracellular distribution of oxidized proteins during oxidative stress have been performed, to our knowledge. Therefore, we used fluorescence microscopy to detect and locate protein carbonyls, oxidation products of basic amino acids, generated in bovine M. Rectus abdominis during either exposition to a chemical free radical generating system, or refrigerated storage, or cooking. The technique consisted of an immunohistochemical detection of carbonyls by reaction with the specific probe DNPH (2,4-dinitrophenylhydrazine) followed by the sequential addition of a first antibody against DNPH-carbonylated proteins and a CY3-labeled secondary antibody. The fluorescence of the CY3 probe increased regularly with level of free radical generating system and storage time. Moreover, an important heterogeneity of carbonyl distribution was observed, with a higher oxidation level at the periphery than inside the muscle cells. Cooking induced fluorescence increase only at the periphery of cells. Specific coloration of collagen by Sirius red showed that collagen was not involved in fluorescence. We can deduce that accumulation of oxidized proteins observed in the cell periphery was linked to membrane protein oxidation and not to connective tissue oxidation. Biochemical assays were performed in parallel on membrane and myofibrillar proteins to provide complementary quantitative data on level of oxidized proteins.

The main function of mastication is to transform a solid food into a bolus that can be swallowed safely. The bolus characteristics such as particles size or cohesiveness, are continuously sensed during mastication and they are important in initiating deglutition. This study examined the following question: What is the condition of the bolus just before swallowing? Ten subjects with normal dentition aged 37.5 +/- 3.7 years were asked to chew without swallowing six different foods (three nuts and three vegetables) while the number of cycles and the duration of the sequence were recorded. The particle size distribution shown by the expectorated food bolus just before swallowing was examined by image analysis. The results showed that, for a given food, the sizes of the bolus particles just before swallowing were comparable in all subjects. However, the number of cycles and duration of the sequence varied between subjects. Taken together these data strongly suggest that the granularity of the bolus before swallowing has to reach a predetermined state which is obtained by using an individual chewing strategy. This suggests that the bolus structure reflects a key factor for homeostasis and explains the large interindividual variability of the mastication physiologic parameters.

International audience; Pig semimembranosus muscles, sampled from normal hams or from PSE-zones of defective hams, were analysed by histochemistry and electrophoretic techniques. PSE zones were characterised by a disorganisation of fibre alignment and a significant increase of inter fibre spacing (26.2% vs. 16.9%, p < 0.05). Protein solubility was significantly lower in defective muscle (55.4 vs. 91.5 mg/g, p < 0.001). SDS-PAGE evidenced in such samples a lower abundance of the 97, 40 and 26 kDa bands in the sarcoplasmic fraction and a higher abundance of the 97, 58, 34, 31, 15 and 11 kDa bands in the myofibrillar fraction. Intensity of the MHC band (200 kDa) was lower in PSE zone samples. By 2-D electrophoresis, it was shown that troponin T, MLC 1 and alpha-crystallin were less proteolysed in defective muscles, while creatine kinase fragments were more represented. One form of HSP 27 was absent from PSE zone samples. Overall, meat from PSE-zones and fast pH fall-PSE meat show numerous histological and biochemical similarities, particularly in their protein characteristics.

Game meat is commonly consumed in Europe but few studies have examined the quality related parameters. In this study we examined the changes in ultrastructure at four times postmortem in M. longissimus from moose ( Alces alces ) and reindeer ( Rangifer tarandus tarandus ). The moose were slaughtered during a hunt and reindeer by Swedish standard practices for this semi-domestic animal. Ultrastructural changes occurring in all animals included separation of the sarcolemma from myofibrils, I band breaks, and cytoskeleton breaks; however both I band breaks and cytoskeletal breaks were less common in moose and reindeer than values reported for sheep and beef. Fiber area in the longissimus thoracis muscle was approximately 3270 μm 2 for moose and 1170 μm 2 for reindeer indicating that the tenderness of reindeer meat may be largely determined by fiber size.

International audience; A cytosolic pH fall is associated with the ATP depletion during hypoxia. Increase in cytosolic free Ca 2 + has also been hypothesized as an accelerating factor in the death of cells under hypoxia. To assess to what extent pH and Ca 2 + are involved in the cell death, we investigated the changes of these two parameters in muscle cells when applying a c hemical hypoxia . The cellular response to hypoxia beeing variable from cell to cell, measurements were performed at a single cell level. The myogenic precursors were satellite cells obtained from masseter muscle of normal adult pigs or from halothane susceptible (HS) pigs. These latter are known to carry a genetic mutation of the ryanodine receptor, the calcium channel of the sarcoplasmic reticulum, leading to abnormal cytosolic calcium regulation.Cells were grown until differentiation stage where spontaneous contractions occur. Hypoxia was induced by adding sodium cyanide and iodoacetate to a medium containing 1.8mM Ca 2 + or Ca 2 + -free. Fluorescence of the probes loaded in the cells, BCECF (for pHi) and Fura2 (for [Ca 2 + ]i), were recorded under dual wavelength excitation, every 2 min during 30 min at 37 o C.In all conditions, pHi fell abruptely within the first 10 min of hypoxia and then stabilises at a value of 6.5. This fall seemed to be more pronounced in normal cells. There was a transient and early increase of [Ca 2 + ]i in normal cells while [Ca 2 + ]i increased slowly and progressively in HS cells. Ultimate values of [Ca 2 + ]i were about the same for both type of cells. Blebs and subsequent cell death occured more frequently in normal cells than in HS. These results suggest that cell death may not be directly related to [Ca 2 + ]i increase. pH fall may be a determinant factor in cell injury leading to cell death.

Cow muscles ( Longissimus dorsi, Tensor fasciae latae, Psoas major, Diaphragma medialis ), with different colour stabilities, were used to measure ‘Metmyoglobin reductase activity’ in different conditions. The effects of pH and temperature on in vitro metmyoglobin reduction were analysed. The highest metmyoglobin reductase activity was localized in microsomes and in more or less intact mitochondria by means of differential centrifugations. The most unstable muscles, from the colour point of view, presented the highest reducing activities and no differences were noted between activities measured in aerobic or anaerobic conditions.

To explain the different rates of metmyoglobin accumulation in bovine muscles, biochemical factors such as oxygen consumption rate, cytochrome a(+a3) content, myoglobin autoxidation and enzymic ferrimyoglobin reduction were studied. The measurements involved five cows and five bulls. Only six cows were used for Metmyoglobin Reducing Activity (MRA) measurements. Three muscles with different colour stability (Tensor fasciae latae, Psoas major and Diaphragma medialis) were chosen. Meat colour stability, as well as the oxido-reduction potential of myoglobin, were highly related to muscle type. The animal effect was low or non-significant. Muscles with the poorest colour stability, such as Psoas major and Diaphragma medialis had the highest oxidative activities (oxygen consumption rate) and the highest myoglobin autoxidation rates. Enzymic ferrimyoglobin reduction, estimated either by spectrophotometric measurements from muscle homogenates in aerobic conditions (‘Metmyoglobin Reductase Activity’) or by reflectance measurements in anaerobic conditions (MRA), does not explain the differences observed in muscle colour stability.

Trials have been carried out to examine the effects of a food additive supply (Maturex) and/or of anabolic implant (Revalor) on carcass characteristics and on veal meat qualities. Twelve Holstein × Francaise Frisonne Pie Noire calves acted as controls, twelve calves were implanted with Revalor, approximately 60 days prior to slaughter, twelve others were fed with Maturex, whilst the remaining twelve animals were fed with Maturex and implanted with Revalor. All animals were slaughtered at 4 months. Revalor implantation had a significant effect on carcass weight and conformation. Feeding with Maturex had no significant effect on different carcass characteristics. Neither anabolic treatment nor Maturex supplementation significantly affected pH value, haematocrit, pigment content, FOP, colour characteristics, collagen content and solubility, or dry matter. Although Revalor had a negative effect on weight losses during cooking, Maturex showed no significant effect. While Maturex showed a positive effect on tenderness and no effect on juiciness, Revalor showed a negative effect on tenderness and juiciness. For all sensory characteristics, no effect was noted when both Maturex and Revalor were used together.

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