Miho Hirabayashi

Difference in tenderness and pH decline between water buffalo meat and beef during postmortem aging.

The objective of this research was to determine the difference in tenderness and some characteristics of water buffalo meat and beef during postmortem aging. Five female crossbred water-buffalo (Philippine Carabao×Bulgarian Murrah) and five female crossbred cattle (Brahman×Philippine Native), were finished on the same diet for 6 months and slaughtered at 30 months of age. The muscle pH was measured at 40min, 3h, 7h, 24h, and 48h postmortem. Longissimus thoracis (LT) and semimembranosus (SM) muscles were excised at 2d postmortem, and shear force was measured at 2, 4, 7, and 14d postmortem. Glycogen and lactate concentrations were determined from 0, 2, and 4d LT samples, and myosin heavy chain type of buffalo and cattle LT was determined by ELISA methods. Myofibrillar protein degradation was also observed by SDS-PAGE and Western blotting of fast-type troponin T. Results showed that the buffalo meat had significantly lower shear force values compared to beef for LT and SM muscles, which was supported by a difference in troponin T degradation. Postmortem pH decline of buffalo meat was significantly slower than that of beef, which was confirmed by lactic acid concentrations, but was not explained by glycogen content. In addition, there was no significant difference in the ratio of slow to fast type muscle fibers in buffalo and cattle, indicating that myosin heavy chain type was not responsible for the difference in pH decline and tenderness between the buffalo meat and beef. This study demonstrated that the tenderness of water buffalo meat was superior to that of Brahman beef, which may have been due to the difference in pH decline and the subsequent effect on muscle protease activity.

Alleviation of maternal hyperthermia-induced early embryonic death by administration of melatonin to mice

Abstract:  Maternal hyperthermia induces early embryonic death via increased oxidative stress to the embryo. In this study, we examined whether melatonin administered to heat‐stressed pregnant mice would reduce hyperthermia‐induced embryonic death. Mice were heat stressed (12 hr at 35°C, 60% relative humidity) on the day of mating and melatonin (3 mg/kg body weight) was injected subcutaneously every 2 hr during heat exposure. Thereafter, zygotes were collected, and in vitro developmental ability and intracellular glutathione (GSH) content were assessed. In addition, reactive oxygen species (ROS) levels and free radical scavenging activity (FRSA) in the oviduct as well as lipid peroxidation in the liver were measured. Melatonin administration was associated with a tendency for higher intracellular GSH content in zygotes (1.67 pmol/zygote) and a significantly higher percentage of embryos that developed to the morula or blastocyst stage (47.91%; P < 0.01) compared with the parameters in heat‐stressed mice that were administered a placebo (1.48 pmol GSH/zygote and 14.78% development). Lipid peroxidation levels in the liver and ROS levels in the oviduct were the same in melatonin‐treated stressed mice and the controls, while these parameters were significantly higher in heat‐stressed mice that were not treated with melatonin. Furthermore, FRSA in the oviduct was significantly (P < 0.05) higher in the melatonin‐treated mice than in the controls. These results suggest that administration of melatonin to heat‐stressed mice alleviates hyperthermia‐induced early embryonic death and that this is accomplished in part by maintaining a neutral redox status within the mother.

Redox Status of the Oviduct and Cdc2 Activity in 2-Cell Stage Embryos in Heat-Stressed Mice

Abstract Mammalian preimplantation embryos are vulnerable to heat stress. However, the mechanisms by which maternal heat stress compromises embryonic development are unclear. We hypothesized that the loss of developmental competence in maternally heat-stressed embryos results from enhanced oxidative stress in the oviducts. In experiment 1, oviducts and zygotes were collected from mice that were heat-stressed at 35°C and 60% relative humidity for 12 h on the day of pregnancy as well as from control mice. The zygotes were cultured for 84 h to assess their development, and the H2O2 level, glutathione concentration, and free radical scavenging activity (FRSA) were measured in the oviduct. In experiment 2, zygotes were cultured for 22 h to reach the late G2 phase in the 2-cell stage, and Cdc2 activity was assessed using immunoblotting. A high percentage (87.6%) of control embryos developed to morulae or blastocysts, whereas the majority (67.4%) of the heat-stressed group arrested at the 2-cell stage. Although heat stress did not alter the FRSA or glutathione concentration in the oviducts, the H2O2 level (P < 0.01) and its ratio to the FRSA (P < 0.05) significantly increased in the heat-stressed group. The Cdc2 activation at the 2-cell stage, as shown by the ratio of the dephosphorylated form to the phosphorylated form, was evident in control embryos but absent in heat-stressed embryos, and the level was similar to that in embryos blocked at the 2-cell stage (positive control). These results indicate that maternal heat stress enhances oxidative stress in the oviducts and that loss of developmental competence in maternally heat-stressed embryos correlates with a defect in Cdc2 activity at the 2-cell stage.

Protease activity higher in postmortem water buffalo meat than Brahman beef

We previously demonstrated that postmortem water buffalo meat had higher tenderness than Brahman beef. In order to explain this difference in tenderness, the objective of the current study was to investigate the protease activity in these two meats. Five female crossbred water buffalo (Philippine Carabao×Bulgarian Murrah) and five female crossbred cattle (Brahman×Philippine Native) were slaughtered at 30months of age, followed by immediate sampling of Longissimus thoracis muscle for measurement of protease activity. Results showed that buffalo meat had significantly higher protease activity compared to beef (P<0.05). Furthermore, calpain inhibitor 1, a specific inhibitor of calpains 1 and 2, was the most effective inhibitor of protease activity. There was no difference in calpastatin activity, and no major differences were observed in calpains 1, 2, and calpastatin expression by Western blotting. This study suggests that higher calpain activity in early postmortem buffalo meat was responsible for the increased tenderness of water buffalo meat compared to beef.

Possible roles of myostatin and PGC-1α in the increase of skeletal muscle and transformation of fiber type in cold-exposed chicks: Expression of myostatin and PGC-1α in chicks exposed to cold

This study examined the hypothesis that myostatin and PGC-1alpha are involved in the increase in skeletal muscle mass and transformation of fiber type in cold-exposed chicks. One-week-old chicks were exposed to acute (24h) or long-term (8d) cold at 4 degrees C or kept warm at 30 degrees C. Acute cold exposure induced a significant increase in the skeletal muscle weight and the ratio of slow- to fast-fiber specific troponin I expression (sTnI/fTnI), accompanied by a significant decrease in lactate dehydrogenase activity. Expression of myostatin mRNA in the muscle was significantly lower in cold-exposed chicks than in the controls, whereas PGC-1alpha mRNA expression was significantly enhanced. These changes in the gene expression rapidly returned to the levels of the control chicks after the end of cold exposure, whereas the changes in fiber type and enzymatic activity were not resumed within 24h after removal of cold exposure. On the other hand, long-term exposure to cold resulted in a remarkable increase in skeletal muscle weight, accompanied by a significant increase in the ratio of sTnI/fTnI and the enzymatic activities of cytochrome oxidase and lactate dehydrogenase. However, the expression level of myostatin mRNA in cold-exposed chicks was not different from that in their age-matched control chicks and that of PGC-1alpha mRNA was significantly lower than in the controls. These results indicate that myostatin and PGC-1alpha expression in the skeletal muscle rapidly change in response to acute cold, suggesting the possibility that these two genes could be involved in the increase in muscle mass and transformation of fiber type, respectively, at the initial stage of adaptation in cold-exposed chicks.

Possible roles of myostatin and PGC-1alpha in the increase of skeletal muscle and transformation of fiber type in cold-exposed chicks: expression of myostatin and PGC-1alpha in chicks exposed to cold.

This study examined the hypothesis that myostatin and PGC-1alpha are involved in the increase in skeletal muscle mass and transformation of fiber type in cold-exposed chicks. One-week-old chicks were exposed to acute (24h) or long-term (8d) cold at 4 degrees C or kept warm at 30 degrees C. Acute cold exposure induced a significant increase in the skeletal muscle weight and the ratio of slow- to fast-fiber specific troponin I expression (sTnI/fTnI), accompanied by a significant decrease in lactate dehydrogenase activity. Expression of myostatin mRNA in the muscle was significantly lower in cold-exposed chicks than in the controls, whereas PGC-1alpha mRNA expression was significantly enhanced. These changes in the gene expression rapidly returned to the levels of the control chicks after the end of cold exposure, whereas the changes in fiber type and enzymatic activity were not resumed within 24h after removal of cold exposure. On the other hand, long-term exposure to cold resulted in a remarkable increase in skeletal muscle weight, accompanied by a significant increase in the ratio of sTnI/fTnI and the enzymatic activities of cytochrome oxidase and lactate dehydrogenase. However, the expression level of myostatin mRNA in cold-exposed chicks was not different from that in their age-matched control chicks and that of PGC-1alpha mRNA was significantly lower than in the controls. These results indicate that myostatin and PGC-1alpha expression in the skeletal muscle rapidly change in response to acute cold, suggesting the possibility that these two genes could be involved in the increase in muscle mass and transformation of fiber type, respectively, at the initial stage of adaptation in cold-exposed chicks.

Increased Mass of Slow-Type Skeletal Muscles and Depressed Myostatin Gene Expression in Cold-Tolerant Chicks

Temperature is maintained in birds by skeletal muscle shivering as well as by non-shivering thermogenesis in a cold environment because they lack brown adipose tissue, which is a mammalian thermogenic organ. Chicks acquire cold tolerance after their skeletal muscles mature. Here, we found that muscle fibers transformed to the slow-twitch type with increasing gene expression of peroxisome proliferator-activated receptor-&ggr; coactivator-1&agr; (PGC-1&agr;), and that the mass increased with decreasing myostatin gene expression, in the leg muscles of 7-day-old and younger chicks within 24 h of cold exposure. Muscle fibers did not transform and the mass did not increase within 24 h of cold exposure in muscles from chicks older than 8 days of age. Myostatin mRNA expression remained depressed in cold-tolerant muscles for 24 h, whereas cold-enhanced growth of the muscle continued for 48 h. Myostatin expression was depressed and muscle mass was increased only in chick leg muscles that comprised both fast- and slow-twitch fibers. These results suggest that the acute regulation of PGC-1&agr;a and myostatin gene expression in leg muscles is required for chicks to acquire cold tolerance up to 7 days of age.

Role of prolactin-like protein (PRL-L) in cold-induced increase of muscle mass in chicks

This study examined the hypothesis that a novel prolactin-like protein gene (PRL-L) is involved in cold-induced growth of skeletal muscle in chicks. Six-day-old chicks (Gallus gallus domesticus) were exposed to cold at 4°C or kept warm at 30°C for 24h. Cold exposure induced significant increases in PRL-L expression that coincided with increases in the weight of the sartorius muscle, which comprises both fast- and slow-twitch fibers. Meanwhile, no induction of PRL-L mRNA was observed in the heart, liver, kidney, brain, or fat. Myoblast cells that expressed PRL-L mRNA grew faster than untransduced cells in media containing 2% serum. These results suggested that PRL-L might be involved in in controlling cold-induced muscle growth of chicks.

Relation of postmortem protease activity to tenderness in buffalo meat and Brahman beef

Abstract We previously showed that meat from crossbred water buffalo had significantly higher tenderness than beef from crossbred Brahman cattle of the same age, gender, and diet. Extensive studies on meat tenderness have indicated that proteases degrade muscle fibre proteins during postmortem storage, leading to weakening of the myofibrillar structure and an increase in tenderness. Thus, we investigated the difference in protease activity immediately postmortem, in order to explain the difference in tenderness between buffalo meat and beef. Five female crossbred water-buffalo (Philippine Carabao x Bulgarian Murrah) and five female crossbred cattle (Brahman x Philippine Native) were slaughtered at 30 months of age, and Longissimus thoracis muscle was sampled immediately post-slaughter. Protease activity at different pH levels and the effect of various inhibitors on protease activity were examined. Results showed that buffalo meat had significantly higher protease activity compared to beef, and calpain inhibitor 1 was the most effective inhibitor. As calpain inhibitor 1 is a specific inhibitor of calpain 1 and 2, the results suggest that higher calpain activity in buffalo meat was responsible for the higher tenderness of buffalo meat compared to Brahman beef.

Increased expression of NOR-1 mRNA in the skeletal muscles of cold-exposed neonatal chicks.

Nuclear receptor subfamily 4, group A (NR4A) subgroup orphan receptors are rapidly induced by various physiological stimuli and have been suggested to regulate oxidative metabolism and muscle mass in mammalian skeletal muscle. The results showed that the NR4A subgroup orphan receptor, NOR-1 (NR4A3), was acutely increased in skeletal muscles of neonatal chicks in response to short-term cold exposure. The increased NOR-1 gene expression was concomitant with cold-induced changes in gene expression of both myostatin and proliferator-activated receptor-gamma coactivator-1 (PGC-1α), and the increase in skeletal muscle mass. These observations suggest that NOR-1 might play a role in controlling skeletal muscle growth in cold-exposed neonatal chicks.