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  • Author or Editor: Takao Sugiura x
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Objective—To determine the growth-related changes in metabolic and anatomic properties in equine muscle fiber type, including hybrid fibers identified with immunohistochemical analysis.

Animals—24 2-, 6-, 12-, and 24-month-old female Thoroughbreds.

Procedure—Samples were obtained from the gluteus medius muscle of all horses. Expression of myosin heavy chain (MHC) isoforms MHC-I, -IIa, -IIb, and -IIx in each muscle fiber was detected by use of 4 primary monoclonal antibodies: BA-D5, SC-71, BFF3, and BF-35, respectively. Five muscle fiber types (types I, I/IIA, IIA, IIA/IIX, and IIX) were immunohistochemically identified. The area and activity of succinic dehydrogenase (SDH) in each fiber type were determined by use of quantitative histochemical staining and image analysis.

Results—Although the proportion of type I and IIX fibers did not change with age, the proportion of type IIA and IIA/IIX fibers significantly increased and decreased, respectively, from 2 months to 24 months of age. The increase in proportion of type IIA fibers with growth may have been attributable to muscle fiber-type transition from type IIA/IIX fibers but not from type IIX fibers. Values for SDH activity and fiber area in hybrid fiber types were intermediate to those for their respective pure phenotypes.

Conclusions and Clinical Relevance—Hybrid fibers have an important role for determining the proportion of muscle fiber type in horses < 24 months old, and the metabolic and anatomic properties of the hybrid fibers are well coordinated, as in mature horses. (Am J Vet Res 2005;66:401–405)

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in American Journal of Veterinary Research



To determine histochemical and biochemical properties of muscle during adaptation to training on a flat or sloped track.


22 Thoroughbreds.


Samples were obtained from the middle gluteus muscle before and after training programs were conducted, using a needle-biopsy technique. Training programs consisted of horses running 1,600 m on a flat or sloped track for 16 weeks. Amplitude of middle gluteus muscle activity per burst was calculated. Muscle fiber composition and area were examined on serial cross sections processed by standard histochemical staining procedures (ATPase stain after prior incubation with an acid or base, followed by succinate dehydrogenase [SDH] stain). Furthermore, SDH and phosphofructokinase activities were determined biochemically, and composition of myosin heavy chain (MHC) isoforms was analyzed electrophoretically.


Training resulted in substantial adaptations. Relative contribution of muscle fiber with high SDH activity (type-I and -IIa fibers) to total cross-sectional area, SDH activity, and composition of MHC-IIa isoforms were increased. Gel electrophoresis revealed a large amount of MHC-IIx isoform and a small amount of MHC-IIb isoform in the muscle. Although root mean square of muscle activity for training on a sloped track was 7.6% higher than the value obtained while training on a flat track, muscle histochemical and biochemical properties did not differ significantly between groups training on flat and sloped tracks.


Training adaptations for contractile and metabolic properties of the middle gluteus muscle were evident for the 2 types of training. However, training adaptations did not differ significantly between groups trained on flat or sloped tracks. (Am J Vet Res 1999;60:1536–1539)

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in American Journal of Veterinary Research


Objective—To determine the effect of growth and training on metabolic properties in muscle fibers of the gluteus medius muscle in adolescent Thoroughbred horses.

Animals—Twenty 2-year-old Thoroughbreds.

Procedure—Horses were randomly assigned to 2 groups. Horses in the training group were trained for 16 weeks, and control horses were kept on pasture without training. Samples were obtained by use of a needle-biopsy technique from the middle gluteus muscle of each horse before and after the training period. Composition and oxidative enzyme (succinic dehydrogenase [SDH]) activity of each fiber type were determined by use of quantitative histochemical staining procedures. Whole-muscle activity of SDH and glycolytic enzyme (phosphofructokinase) as well as myosin heavy-chain isoforms were analyzed biochemically and electrophoretically, respectively.

Results—The SDH activity of type-I and -IIA fibers increased during growth, whereas whole-muscle activity was unchanged. Percentage of type-IIX/B muscle fibers decreased during training, whereas that of myosin heavy-chain IIa increased. The SDH activity of each fiber type as well as whole-muscle SDH activity increased during training. An especially noticeable increase in SDH activity was found in type-IIX/B fibers.

Conclusions and Clinical Relevance—Changes in muscle fibers of adolescent Thoroughbreds are caused by training and not by growth. The most noticeable change was for the SDH activity of type-IIX/B fibers. These changes in the gluteus medius muscle of adolescent Thoroughbreds were considered to be appropriate adaptations to running middle distances at high speeds. (Am J Vet Res 2002;63:1408–1412)

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in American Journal of Veterinary Research


Objective—To compare methods for harvesting canine bone marrow stromal cells (BMSCs) and determine the biological properties of canine BMSCs at successive passages in vitro.

Sample—BMSCs collected from the femurs of 9 Beagles.

Procedures—A fibroblast assay was performed to compare 2 methods for harvesting BMSCs: the aspiration and perfusion method. Flow cytometric analysis was performed to evaluate the cell surface markers. Changes in proliferative activity were analyzed by examining radioactivity of hydrogen 3-thymidine. Cell senescence was studied via senescence-associated β-galactosidase staining, and differentiation properties (osteogenesis and adipogenesis) were estimated in association with passage.

Results—The aspiration method yielded significantly more fibroblasts than the perfusion method. The cells harvested by both methods gave positive results for CD44 and CD90 and negative results for CD34 and CD45. After induction, the cells had osteogenic and adipogenic phenotypes. The biological properties of BMSCs harvested by the aspiration method were estimated in association with passage. With increasing number of passages, the proliferative activity was reduced and the proportion of cells with senescence-associated β-galactosidase staining was increased. The capacity of differentiation was reduced at passage 3.

Conclusions and Clinical Relevance—The aspiration method was superior for collection of BMSCs. In early passages, canine BMSCs had the proliferative activity and potential of osteogenic and adipogenic differentiation, but this decreased with increased number of passages. Consideration of passage will be important to the success of any strategy that seeks to regenerate tissue though the use of BMSCs.

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in American Journal of Veterinary Research