Sequential expression of myogenic regulatory factors in bovine skeletal muscle and the satellite cell culture

Myogenic regulatory factors (MRFs) are important in the control of skeletal muscle development. To understand myogenic regulation by MRFs in bovine adult muscle cells, their expressions, namely that of Myf5, MyoD, myogenin, and MRF4 in the biceps femoris muscle (BF) and in the satellite cell culture, were analyzed by RT-PCR. In the BF, all four MRFs were expressed and in particular, myogenin and MRF4 were strongly expressed, whereas Myf5 was faintly expressed. The satellite cells prepared from the BF expressed Myf5, but only a trace of MyoD, at day 9 of culture. During the growth of the cells to day 14, the MyoD and myogenin expressions gradually increased, and that of MyoD expression reached its maximum at the confluence of the culture. After induction of myogenic differentiation by a serum-free medium at day 14, Myf5 expression gradually decreased, and the up-regulated expression of MyoD was suppressed, whereas myogenin expression continued to increase sharply. Following the myogenin expression, MRF4 also drastically increased toward the myotube formation of the cells. When huge myotubes were formed at day 18, Myf5 was expressed at a low level, whereas the MyoD expression remained at a moderate level.     

Cellularity of developing subcutaneous adipose tissue in Landrace and Meishan pigs: Adipocyte size differences between two breeds

Experiments were designed to compare the adipocyte cellularity of subcutaneous adipose tissue between growing Landrace (low backfat) and Meishan (high backfat) pigs at 1 week, 3 weeks, 6 weeks, 3 months and 5 months of age. As pigs aged, body weight and backfat thickness of both breeds significantly increased. When compared at equal ages, backfat thickness adjusted to equal body weight was greater for Meishan pigs. The mean diameter of fat cell size also increased with age, and by 6 weeks adipocytes from both outer and inner layers of subcutaneous adipose tissue were larger in Meishan pigs. At 5 months, approximately 80% of the adipose tissue mass in Meishan pigs was attributable to adipocytes measuring 95–165 µm in diameter, whereas adipocytes of 75–145 µm comprised most of the tissue mass in the Landrace. Although the contribution of smaller adipocytes (25–45 µm) to the tissue volume was negligible, both breeds showed a biphasic diameter distribution at all ages, suggesting that adipocyte hyperplasia is still active. Our results demonstrate that cellularity differences exist between the subcutaneous adipose tissues of Landrace and Meishan pigs, and adipocyte hypertrophy is the most overwhelming contributor to the greater backfat deposition for Meishan pigs.     

Comparative sequence analysis of four myosin heavy chain isoforms expressed in porcine skeletal muscles: Sequencing and characterization of the porcine myosin heavy chain slow isoform

A nucleotide sequence including the full coding region for the myosin heavy chain (MyHC) slow isoform was determined from the longissimus skeletal muscle. The deduced amino acid sequence was 1935 residues, which was the same length as the human and rat MyHC-slow isoforms. The porcine MyHC-slow isoform showed 97.6% and 97.4% amino acid identities to the human and rat isoforms on their entire regions, respectively. The functional regions were also highly conserved among mammalian MyHC-slow isoforms. Amino acid substitutions between the porcine MyHC-slow and MyHC-fast isoforms were concentrated on the functional regions. Loop 1, the controlling region of nucleotide binding and release, was conserved among the fast isoforms, but not between the slow and fast isoforms. Loop 2, a part of the actin binding region, was not conserved among any of the isoform types, and the most substitutions in this region were found in the slow isoform. The myosin essential light chain binding region was conserved among the fast isoforms, except for some substitutions in the 2b isoform, but was clearly different in the slow isoform. The myosin regulatory light chain binding region was conserved among the fast isoforms, but not between the slow and fast isoforms. These results indicate that the functional difference between the porcine MyHC-slow and -fast isoforms are controlled by the sequence diversity at the four functional regions compared.     

Population density and range use of sika deer,Cervus nippon, on Mt. Ohdaigahara, central Japan

We investigated the population density and range use of sika deer,Cervus nippon, on Mt. Ohdaigahara, central Japan, from July 1996 to July 1997. The deer population density estimated by the block count was high at 17.5–30.9 deer/km². Although the deer remained this area from spring to fall, they moved out in winter, probably because of deep snow. A spotlight count showed that the deer prefer the eastern area as spring-summer range, where the forest consists of coniferous trees over a floor cover ofSasa nipponica, andS. nipponica grassland. The deer also use dense coniferous forest for cover, especially in summer and fall. Part of this study was presented at the 108th Annual Meeting of the Japanese Forestry Society (1997). This study was financially supported by the Environment Agency, Japan.     

Pathological and phylogenetic features of prevalent canine distemper viruses in wild masked palm civets in Japan

Ten wild masked palm civets infected with canine distemper virus (CDV), captured in Japan from 2005 to 2007, were histopathologically and phylogenetically analyzed. Phylogenetic analysis based on the amino acid sequences of the H protein of two CDV isolates from masked palm civets revealed that the two isolates were classified into the clade of recent isolates in Japan. Histopathologically marked lesions of virus encephalitis were present in the brain, whereas gastrointestinal lesions were absent or at a mild degree. The distribution of the lesions resembles that of recent CDV cases in dogs. Therefore, recent CDV infections in masked palm civets could be caused by recently prevalent CDV in dogs. The possibility of the masked palm civet as a spreader of CDV among wildlife is also discussed.     

Effects of muscle type on beef taste‐traits assessed by an electric sensing system

To assess the role of muscle fiber type in beef taste‐traits, we analyzed cooked meats from bovine masseter, diaphragm, psoas major, longissimus thoracis, and semitendinosus muscles with an electric taste sensing system (INSENT SA402B). The system is composed of five taste sensors of polymer membranes fixing different lipids. The sensors, CT0, CA0, AAE, C00 and AE1 are designed to respond to the individual tastes of salty, sour, umami, bitter and astringent, respectively. The system found significant differences in the converted outputs of CA0 (cvCA0), C00 (cvC00) and AE1 (cvAE1) among the bovine muscles. The slow‐type muscles (masseter and diaphragm) showed lower cvCA0, higher cvC00, and higher cvAE1 than did the fast‐type muscles (psoas major, longissimus thoracis, and semitendinosus). Lactic acid content was different among muscle types and was highly related to the cvCA0 output and pH. carbonyl compounds and free fatty acids were higher in the slow‐type muscles. Free fatty acids were major components causing the difference in the C00 output among the muscle types. Iron content was also different among the muscle types and related to the cvC00 and cvAE1 outputs. These results suggested that the muscle fiber type affects the beef taste characteristics.     

Stimulatory effect of aluminum on the growth of tea pollen tubes

It is well known that when inorganic elements such as boron (B), calcium, cobalt, and manganese are added to media in combination or alone, they stimulate the germination and/or tube growth of various kinds of pollen (Schmücker 1933; Loo and Hwang 1944; Yamada 1958; Brewbaker and Kwack 1963). Among these elements, the stimulatory effect ofB is the most effective and generally recognized. Aluminum (Al) belongs along with B to Group IIIa in the periodic table, and Fähnrich and Ultlich (1964) reported that Al inhibited the germination and growth of the pollen tube of Petunia hybrida and Antirrhinum majus. However, stimulatory effects of Al on pollen germination or pollen tube growth had not been previously reported.     

Peroxisome proliferator-activated receptor-γ coactivator 1 α (PGC-1 α) expression and the formation of slow-twitch muscle fibers in porcine and bovine skeletal muscles

The peroxisome proliferator‐activated receptor‐γ coactivator‐1 α (PGC‐1 α) induces mitochondria biogenesis in skeletal muscles. To determine the relationships between PGC‐1 α and the muscle fiber types, the expression levels of PGC‐1 α were analyzed in porcine and bovine skeletal muscles. As a first step, the nucleotide sequences of the porcine and bovine PGC‐1 α were determined. The porcine and bovine PGC‐1 α cDNA encoded 796 amino acid sequences and showed 95.1% identity between the two species. The expression levels of the PGC‐1 α mRNA were analyzed in the same 10 skeletal muscles from four pigs and three cattle. The contents of porcine and bovine PGC‐1 α were higher in the tongue, masseter and diaphragm, and lower in the Biceps femoris, semimembranosus, Longissimus thoracis and semitendinosus muscles. The contents of myosin heavy chain slow‐type protein (MyHC‐slow) were also determined in the same muscles by ELISA. The analysis of MyHC‐slow showed results similar to those for the PGC‐1 α contents in all of the muscles except for the tongue. The content of MyHC‐slow in the tongue was the lowest among the porcine muscles, and moderate among the bovine muscles. The results suggest that PGC‐1 α relates to the development of oxidative muscle fibers, but is not the principal factor in determining type I fiber content.     

Proteome analysis of whole and water‐soluble proteins in masseter and semitendinosus muscles of Holstein cows

To assess both quantitative and qualitative differences between the slow‐ and fast‐type muscles, masseter (slow) and semitendinosus (fast) from four Holstein cows were analyzed by two‐dimensional difference gel electrophoresis (2D DIGE) and mass spectrometry. The proteome analysis identified 27 spots as 20 proteins in the whole protein fraction extracted with 8 mol/L urea solution, and 16 spots were identified as 11 proteins in the water‐soluble protein fraction. Two slow‐type myofibrillar proteins (myosin light chain‐1 slow‐b and myosin light chain‐2 slow), and aconitase‐2 mitochondria were present at higher levels in the masseter muscle (P < 0.05). Four fast‐type myofibrillar proteins (myosin light chain‐1 fast, myosin light chain‐2 fast, myosin light chain‐3 fast and tropomyosin‐1), and three enzymes of glycolytic pathway (enolase‐3, aldolase‐A and triosephosphate isomerase), were present at higher levels in the semitendinosus muscle (P < 0.05). Our proteome analysis showed that the composition of sarcoplasmic proteins as well as myofibrillar proteins was clearly different between slow‐ and fast‐type muscles.     

Relationships between tropomyosin and myosin heavy chain isoforms in bovine skeletal muscle

The composition of tropomyosin (TPM) and myosin heavy chain (MyHC) isoforms was analyzed in 10 physiologically different bovine muscles ( masseter , diaphragm, tongue, semispinalis, pectoralis profundus , biceps femoris, psoas major , semimembranosus, longissimus thoracis and semitendinosus ) to clarify the relationships between TPM and MyHC isoforms in different muscle fiber types. The content of TPM1 and TPM3 was different in muscles according to their function in muscle contraction, although the content of TPM2 was constantly about 50% of the total TPM in all muscles. The content of TPM1 was higher in semimembranosus , longissimus thoracis and semitendinosus, while that of TPM3 was higher in masseter and diaphragm. The high positive correlation between MyHC-slow content and TPM3 content ( r  = 0.92) suggested a coexpression of TPM3 and MyHC-slow isoforms in a muscle fiber. MyHC-slow and TPM3 were expressed at the same level in masseter and diaphragm, whereas there was more TPM3 than MyHC-slow in tongue and semispinalis , so it appears that the excess TPM3 in tongue and semispinalis is expressed with other MyHC isoforms. MyHC-2a was the only fast type isoform expressed in tongue and semispinalis . Therefore, the excess TPM3 was composed of myofibrils with MyHC-2a. The results suggested that a fiber expressing MyHC-2a would be regulated delicately by changing the TPM isoform types.