Biomedical subjects

J F Hoh

Publications and source records attributed to J F Hoh.

At least 19 recordsLinked to original sources

PubMed1992

Indirect myosin immunocytochemistry for the identification of fibre types in equine skeletal muscle.

The histochemical ATPase method for muscle fibre typing was first described by Brooke and Kaiser in 1970. However, problems have been found with the subdivision of type II fibres using this technique. To determine whether indirect myosin immunocytochemistry using anti-slow (5-4D), anti-fast (1A10) and anti-fast red (5-2B) monoclonal antibodies with cross reactivity for type I, II and IIa fibres, respectively, in a number of species, could identify three fibre types in equine skeletal muscle, data on fibre type composition and fibre size obtained using the two different techniques were compared. Results indicate that different myosin heavy chains can coexist in single equine muscle fibres. Type I and type II fibres were identified by immunocytochemistry, but subdivision of type II fibres was not possible. Although the percentage of type I and type II fibres was not significantly different for the two techniques, a few fibres reacted with both the 1A10 and 5-4D antibodies.

Animals↗

PubMed1992

Muscle sarcomere lesions and thrombosis after spaceflight and suspension unloading.

Spaceflight (flight) and tail suspension-hindlimb unloading (unloaded) produced significant decreases in fiber cross-sectional areas of the adductor longus (AL), a slow-twitch antigravity muscle. However, the mean wet weight of the flight AL muscles was near normal, whereas that of the suspension unloaded AL muscles was significantly reduced. Interstitial edema within the flight AL, but not in the unloaded AL, appeared to account for this apparent disagreement. In both experimental conditions, the slow-twitch oxidative fibers atrophied more than the fast-twitch oxidative-glycolytic and fast-twitch glycolytic fibers. Immunostaining showed that slow-twitch oxidative fibers expressed fast myosin, producing hybrid fibers containing slow and fast myosin isoforms. Two-dimensional gel electrophoresis of flight AL muscles revealed increased content of fast myosin light chains and decreased amounts of slow myosin light chains and fatty acid-binding protein. In the flight AL, absolute mitochondrial content decreased, but the relatively greater breakdown of myofibrillar proteins maintained mitochondrial concentration near normal in the central intermyofibrillar regions of fibers. Subsarcolemmal mitochondria were preferentially lost and reduced below normal concentration. Elevated fiber immunostaining for ubiquitin conjugates was suggestive of ubiquitin-mediated breakdown of myofibrillar proteins. On return to weight bearing for 8-11 h, the weakened atrophic muscles exhibited eccentric contraction-like lesions (hyperextension of sarcomeres with A-band filaments pulled apart and fragmented), tearing of the supporting connective tissue, and thrombosis of the microcirculation. Segmental necrosis of muscle fibers, denervation of neuromuscular junctions, and extravasation of red blood cells were minimal. Lymphocyte antibody markers did not indicate a significant immune reaction. The flight AL exhibited threefold more eccentric-like lesions than the unloaded AL; the high reentry G forces experienced by the flight animals, but not the unloaded group, possibly accounted for this difference. Muscle atrophy appears to increase the susceptibility to form eccentric contraction-like lesions after reloading; this may reflect weakening of the myofibrils and extracellular matrix. Microcirculation was also compromised by spaceflight, such that there was increased formation of thrombi in the post-capillary venules and capillaries. This blockage led to edema by 8-11 h after resumption of weight bearing by the COSMOS 2044 rats. The present findings indicate that defective microcirculation most likely accounted for the extensive tissue necrosis and microhemorrhages observed for COSMOS 1887 rats killed 2 days after landing.

Animals↗

PubMed1992

Muscle fiber types and function.

A major recent advance in the field of muscle fiber types has been the discovery that myogenic factors regulate fiber phenotypic properties. Myogenic influences occur in limb and trunk (somitic) muscle, but are particularly strong in jaw-closing muscles and extraocular muscles that express some unique myofibrillar proteins. In somitic muscles, a variant of fast fiber has been discovered, making four types of fibers: I, IIA, IIX, and IIB. These fibers express different isoforms of myofibrillar and other proteins. The speed and power of the four types of fibers are distinct and are controlled principally by their myosin heavy chains, which modulate the two regulatory steps in the crossbridge cycle, one controlling the rate of development of force, the other controlling the maximal velocity of shortening. Fast fibers have a higher threshold for Ca(2+)-activated force and a steeper force-pCa relation than do slow fibers. This difference is largely attributable to the cooperativity in the attachment of crossbridges and to the difference in Ca2+ binding capacity of their troponin C. Ca2+ initiates force development in muscle by increasing the rate of attachment of crossbridges. The phosphorylation of myosin light chain enhances this action. This effect of phosphorylation underlies the phenomenon of posttetanic potentiation of the isometric twitch in fast fibers.

Animals↗

PubMed1991

Expression of superfast myosin in aneural regenerates of cat jaw muscle.

We investigated whether innervation is necessary for the expression of superfast myosin in regenerating cat jaw-closing muscle. Strips of jaw muscle were permitted to regenerate bilaterally in the beds of a fast limb muscle with innervation on one side being prevented surgically. Immunocytochemical analyses using anti-myosin heavy chain antibodies were done at various times postoperatively, the latest being after 78 days. We found little difference between innervated and uninnervated regenerates up to 27 days postoperatively. All regenerating myotubes expressed fetal myosin. In addition, some myotubes stained for superfast or slow myosin, while others stained for both superfast and slow myosins. Subsequently, uninnervated myotubes became atrophic but continued to express fetal, slow, and superfast myosins while innervated myofibers suppressed fetal and slow myosin expression. These results are consistent with the notion that satellite cells of jaw-closing muscles are committed to express superfast myosin during myogenesis, and that the expression of this program is independent of innervation.

Animals↗

PubMed1991

Basal lamina and superfast myosin expression in regenerating cat jaw muscle.

We investigated the possible role of extracellular matrix in specifying the expression of superfast myosin during cat jaw muscle regeneration. Equal proportions of muscle tissue from jaw and limb were minced together after killing cellular elements from one source. We allowed the mince to regenerate in the bed of a fast limb muscle. Regenerates were analyzed immunocytochemically at 71 to 294 days after operation. Fibers in control regenerates containing live cells from both sources expressed fast, superfast or slow myosins, or a mixture of these myosins. In regenerates containing only one type of live cells, we detected only myosins appropriate to the live cells. Our results suggest that during regeneration the original extracellular matrix of jaw-closing or limb muscle is unable to specify the expression of superfast or fast myosins, respectively; they point to the cellular elements, probably the satellite cells, as determinants of muscle specificity during regeneration.

Animals↗

PubMed1991

Effects of dibutyryl cyclic AMP, ouabain, and xanthine derivatives on crossbridge kinetics in rat cardiac muscle.

In a previous communication, we showed that beta-adrenergic stimulation of cardiac muscles was associated with an increase in the rate of cycling of crossbridges as measured by perturbation analysis in the frequency domain. In this analysis, the frequency at which dynamic stiffness is a minimum (fmin) is taken as a measure of the rate of crossbridge cycling. In this paper, we test the hypothesis that the beta-adrenergic receptor-induced increase in crossbridge cycling rate is mediated by elevation of the intracellular level of cyclic AMP. The approach taken is to compare the effects on fmin in rat papillary muscles during Ba(2+)-activated contractures of 1) an agonist of cyclic AMP that can easily penetrate the cell, namely, dibutyryl cyclic AMP, 2) agents that block cyclic AMP phosphodiesterase, namely, the xanthine derivatives isobutylmethylxanthine and caffeine, and 3) an inotropic agent that does not affect the intracellular level of cyclic AMP, namely, ouabain. Our results showed that dibutyryl cyclic AMP at a dose of 5 mM has the same actions as beta-adrenergic stimulation: it potentiated the isometric twitch force, reduced the time to peak tension and time to half relaxation, and shifted fmin by a factor of 1.8 +/- 0.1 (n = 5). Isobutylmethylxanthine at up to 1.1 mM also acted in the same manner, increasing fmin by a factor of 1.8 +/- 0.2 (n = 6), but ouabain, at a dose (0.03 mM) sufficient to potentiate twitch force by 40 +/- 2% (n = 4), was without effect on the time course of the twitch nor was fmin changed (n = 4). Our findings support the hypothesis that a beta-adrenergic receptor-mediated increase in crossbridge cycling rate is due to an increase in intracellular cyclic AMP level and illustrate the usefulness of the frequency domain analysis approach in the analysis of the mechanism of action of inotropic agents.

1-Methyl-3-isobutylxanthine↗

PubMed1990

Localization of human cardiac beta-myosin heavy chain gene (MYH7) to chromosome 14q12 by in situ hybridization.

The genes coding for each human cardiac myosin heavy chain (alpha-MHC and beta-MHC, MYH6 and MYH7, respectively) are tightly linked and the alpha-MHC gene has been assigned to chromosome 14. In order to provide a more precise regional localization, in situ hybridization experiments were carried out using a 3H-labeled probe derived from a beta-MHC genomic clone. The results demonstrated that the human cardiac MHC genes are located within the q12 band of chromosome 14.

Chromosome Banding↗

PubMed1989

Immunocytochemical analysis of the perinatal development of cat masseter muscle using anti-myosin antibodies.

The developmental changes in myosin gene expression in the masseter muscle of embryonic and juvenile kittens were examined immunocytochemically using anti-myosin heavy chain antibodies of various specificities. In the mature cat, this muscle contains only two phenotypes, the majority of fibres are superfast, the rest being slow fibres. In foetal tissues, the histological appearance of bundles of myotubes, comprising a large central myotube surrounded by a rosette of smaller myotubes, strongly suggest the existence in the jaw muscle of primary and secondary fibres during development. Immunocytochemical data are consistent with the hypothesis that there are four types of fibre; two types of primary fibre as well as two types of secondary fibre. (1) Slow primaries stain strongly with an anti-slow myosin antibody throughout the period under study. These fibres transiently express embryonic but not foetal myosin. (2) Superfast primaries stain for embryonic/foetal and slow myosins in the perinatal period but progressively replace these myosins with superfast myosin during postnatal development. (3) Superfast secondaries initially express embryonic/foetal myosins, but later, beginning around the time of birth progressively replace these myosins with superfast myosin. These fibres do not express slow myosin. (4) Slow secondaries, which initially also express embryonic/foetal myosins, but which postnatally express slow or slow and superfast myosins and express only slow myosin in the adult. These four types of fibres are homologous to the four isotypes of limb muscle fibres and may be derived from distinct lineages of myoblasts.

Animals↗

PubMed1989

Regeneration of cat posterior temporalis muscle in culture.

Cat posterior temporalis muscle has a rapid speed of contraction associated with a unique superfast myosin isoform. Superfast myosin expression appears to be an intrinsic property of the muscle fibres and satellite cells, though in culture they failed to express superfast myosin. We have, therefore, cultured this muscle in a system which had previously been shown to encourage the expression of an adult phenotype. The presence of nerve cells resulted in effective regeneration of cat posterior temporalis muscle and even the formation of functional neuromuscular junctions. However, superfast myosin was not found even in mature, contracting, innervated cultures. Thyroid hormone, a known regulator of myosin isoform expression, also failed to elicit superfast myosin expression. Different culture conditions may allow a different outcome, but under circumstances in which mouse muscle expresses an adult phenotype, cat posterior temporalis muscle fails to do so.

Animals↗

PubMed1988

Immunocytochemical and electrophoretic analyses of changes in myosin gene expression in cat limb fast and slow muscles during postnatal development.

Changes in myosin synthesis during the postnatal development of the fast extensor digitorum longus (EDL) and the slow soleus muscles of the kitten were examined using immunocytochemical techniques supplemented by pyrophosphate gel electrophoresis and gel electrophoresis-derived enzyme linked immunosorbent assay (GEDELISA) of myosin isoforms. The antibodies used were monoclonals against heavy chains of slow and fast myosins and a polyclonal against foetal/embryonic myosin. In both muscles in the newborn kitten, there was a population of more mature fibres which stained strongly for slow but weakly for foetal/embryonic myosin. These fibres were considered to be primary fibres. They formed 4.8% of EDL fibres and 26% of soleus fibres at birth, and continued to express slow myosin in adult muscles. The less mature secondary fibres stained strongly for foetal/embryonic myosin, and these could be divided into two subpopulations; fast secondaries in which foetal/embryonic myosin was replaced by fast myosin, and slow secondaries in which the myosin was replaced by slow myosin. At 50 days the EDL had a large population of fast secondaries (83% of total fibres) and a small population of slow secondaries which gradually transformed into fast fibres with maturity. The vast majority of secondary fibres in the soleus were slow secondaries, in which slow myosin synthesis persisted in adult life. There was a restricted zone of fast secondaries in the soleus, and these gradually transformed into slow fibres in adult life. It is proposed that the emergence of primary fibres and the two populations of secondary fibres is myogenically determined.

Animals↗

PubMed1988

Immunocytochemical and electrophoretic analyses of changes in myosin gene expression in cat posterior temporalis muscle during postnatal development.

Changes in myosin gene expression during the postnatal development of the homogeneously superfast kitten posterior temporalis muscle were examined using immunocytochemical techniques supplemented by pyrophosphate gel electrophoresis and gel electrophoresis-derived enzyme linked immunosorbent assay (GEDELISA) of myosin isoforms. The antibodies used were polyclonals directed against the heavy chains of superfast and foetal myosins and monoclonals against the heavy chains of slow and fast myosins. The fibres of the posterior temporalis in the newborn kitten stained almost uniformly with the anti-foetal myosin antibody and the largest of these fibres stained strongly for superfast myosin. A subpopulation of fibres staining for superfast myosin also stained lightly for slow myosin. These slow staining fibres were evenly distributed in the centres of muscle fibre bundles, reminiscent of primary fibres in limb fast muscle. During subsequent development, slow myosin staining disappeared and superfast myosin replaced foetal myosin so that by 50 days the muscle was virtually homogeneously superfast as in the adult. Fast myosin was never expressed at any stage. It is proposed that fibres staining transiently for slow myosin are superfast primary fibres which are homologous to fast primary fibres recently described in regions of limb muscles devoid of slow fibres in the matured animal. Other jaw-closing muscles have significant populations of slow fibres in the mature animal and it is postulated that there exists in these muscles a second class of jaw primary fibres, the slow primary fibres, in which slow myosin synthesis would be sustained in the adult. It is suggested that the myogenic cells of jaw-closing and limb muscles are of two distinct types preprogrammed to express different muscle genes.

Animals↗

PubMed1988

Myogenic and neurogenic regulation of myosin gene expression in cat jaw-closing muscles regenerating in fast and slow limb muscle beds.

Immunocytochemical techniques were used to study changes in myosin gene expression during the regeneration of the cat posterior temporalis muscle transplanted into the bed of either the fast extensor digitorum longus (EDL) or the slow soleus muscle. Strips of the posterior temporalis, a homogeneously superfast muscle, were treated with Marcaine and then transplanted into limb muscle beds which had been completely cleared of host muscle fibres. The regenerates were examined 6 to 224 days after surgery. Early regenerates in both muscle beds reacted with antibodies against the heavy chain of foetal, slow and superfast myosins, but not with antibodies against fast myosin. In the long-term, regenerates innervated by the EDL nerve expressed only superfast myosin whereas in the regenerates innervated by the soleus nerve most fibres expressed only slow myosin and only a few fibres reacted exclusively with the anti-superfast myosin antibody even after 210 days. In contrast, EDL and soleus muscles regenerating in their own beds expressed foetal, slow and fast myosin, but did not express superfast myosin. The isometric contraction times of the various types of regenerates reflected the types of myosin synthesized. It is concluded that jaw and limb muscle cells exist as two distinct allotypes, each having a distinct repertoire for the expression of adult isomyosins, and that within that repertoire isomyosin gene expression can be modulated by the nerve. Thus, myosin gene expression in skeletal muscle fibres is regulated by both myogenic and neurogenic mechanisms.

Animals↗

PubMed1988

Adrenaline increases the rate of cycling of crossbridges in rat cardiac muscle as measured by pseudo-random binary noise-modulated perturbation analysis.

The mechanism of action of adrenaline on cardiac contractility in rat papillary muscles containing V1 and V3 isomyosins was analyzed during barium-activated contractures at 25 degrees C by frequency domain analysis using pseudo-random binary noise-modulated perturbations. The analysis characterizes a frequency (fmin) at which dynamic stiffness of a muscle is a minimum, a parameter that reflects the rate of cycling of crossbridges. We have previously shown that fmin for V1- and V3-containing papillary muscles were 2.1 +/- 0.2 Hz (mean +/- SD) (n = 10) and 1.1 +/- 0.2 Hz (n = 8), respectively, and that these values were independent of the level of activation. The present study's goal was to determine whether the inotropic action of adrenaline was associated with an increased rate of crossbridge cycling. The results show that a saturating dose of adrenaline increased fmin in V1 hearts by 49 +/- 2% (n = 11). The action on V3 hearts was significantly less; the increase in fmin was 26 +/- 2% (n = 6). The increase in fmin for V1 hearts was shown to be sensitive to the beta-blocking agent propranolol. These results suggest that adrenaline significantly increases the rate of crossbridge cycling by a beta-receptor-mediated mechanism. We conclude that the increased contractility of the heart in the presence of adrenaline arises not only from more complete activation of the contractile proteins but also from the increased rate at which each crossbridge can transduce energy.

Animals↗

PubMed1986

Influence of V1 and V3 isomyosins on the mechanical behaviour of rat papillary muscle as studied by pseudo-random binary noise modulated length perturbations.

Experiments were done on four-week-old rats, containing biochemically verified V1 only, and thyroidectomized adult rats, treated with propylthiouracil, verified to contain V3 only. Contracture tension was induced in isolated papillary muscles either by high potassium solution or 0.5 mmol l-1 Ba2+. Small amplitude length perturbations with peak-to-peak value not exceeding 0.15% L0 were applied to the activated muscle. Both the applied length perturbations and the corresponding resulting force changes were analysed by computer for dynamic stiffness and phase values. In order to reduce data acquisition time, pseudo-random binary noise length changes, rather than the conventional sinusoidal length changes, were used. The plot of the dynamic stiffness against frequency displays a minimum, akin to a resonance phenomenon. The frequency, fmin, at which this resonance occurs, reflects crossbridge kinetics. It was found that the fmin values for the two types of papillary muscles differed by a factor of two. Experiments were also done on chemically skinned muscles containing V1 or V3 isomyosin activated by different concentrations of either barium or calcium ions. It was found that fmin values of skinned fibres were higher than those obtained from intact fibres. However, for each type of muscle the fmin was independent of the activator used as well as the level of activation. The ratio of fmin for V3 to that for V1 remained the same as for intact preparations. We conclude that the difference in mechanical parameters did not arise a possible difference in excitation-contraction coupling mechanism, but rather is a difference in the dynamic properties of the two types of crossbridges.

Animals↗

PubMed1979