The sternocleidomastoid (SCM) muscle is one of the neck muscles responsible for head posture and control of head movement. It functions in rotation, inclination, protraction, extension and flexion of the head, whilst chewing and in exerting increased respiratory efforts. This study is the first one describing the myosin heavy chain (MyHC) isoform composition of the SCM muscleof presumably healthy young males for the purpose of better understanding the contractile properties of the muscle as well as to help in evaluation of pathologically altered structure of the muscle. Autopsy samples were processed immunohistochemically to reveal the MyHC isoform composition. The muscle fibres expressed MyHC-1 (31.5%), -2a (29.7%) and -2x (4.3%) or co-expressed MyHC-2a with MyHC-2x (26.8%), MyHC-1 with MyHC-2a (4.1%) and/or MyHC-1, -2a with -2x (1.1%). In addition to the MyHC isoforms, characteristic of adult limb muscles, a very low percentage of muscle fibres (0.2-2.7%) expressed MyHC-neo, which is normally not found in adult limb muscles. Only two samples exhibited MyHC-neo at a rather higher percentage (6.3% and 7.5%) of muscle fibres. The high share of hybrid fibres and the presence of MyHC-neoin the SCM muscle differ from that of adult limb muscles where hybrid fibres are rare and the expression of immature MyHC isoforms occurs only in pathological or experimental conditions. Since the SCM muscle shares the same embryogenic potential as limb muscles, its distinct MyHC expression appears tobe associated with twin innervation and with the intrinsic specialisation toperform multiple functions.
COBISS.SI-ID: 29853401
We developed a staining protocol that enables simultaneous visualization of myosin heavy chain (MyHC) pure and hybrid muscle fiber types in rat skeletal muscle. Up to eight different muscle fiber types can be visualized in a single section of the rat extensor digitorum longus, which contains all four adult MyHC isoforms and shows plasticity during the denervation-reinnervation process. Triple immunofluorescent staining of MyHC-1, MyHC-2a and MyHC-2b with primary antibodies BA-D5 (isotype IgG2b), SC-71 (isotype IgG1), BF-F3 (isotype IgM), and with three fluorophore-labeled isotype-specific secondary antibodies displays different muscle fiber types in a merged image of red, green and blue channels, each in its own color. Immunoperoxidase staining with primary antibodies 6H1 directed against MyHC-2x can be additionally applied on the same tissue section to facilitate the identification of muscle fibres containing MyHC-2x. Triple staining can also be used in combination with other staining procedures to derive more information about the number of capillaries or the oxidative potential of muscle fiber types. Simultaneous visualization of multiple fiber types in a single merged image enables economical use of muscle samples and provides simple and rapid identification of all fiber types that are present in rat limb muscles.
COBISS.SI-ID: 30372825
It is well documented that, besides reducing blood LDL lipoproteins, HMG-CoA reductase inhibitors (statins) also suppress inflammatory markers and improve survival in sepsis. These beneficial effects can be at least partly explained by their capacity to inhibit the release of IL-6, which is generally regarded as a proinflammatory cytokine, although a variety of other actions including anti-inflammatory have been reported for this cytokine under various circumstances. In quantitative terms, IL-6 release is a major response of the skeletal muscle to various environmental stimuli and since muscle represents 40% of the body weight it can substantially contribute to the IL-6 blood level. The aim of our study was to provide more detailed insight into the effects of statins on the IL-6 release from the human skeletal muscle. Studying time and concentration dependency of the constitutive and lipopolysaccharide (LPS)-stimulated IL-6 release from the cultured human myotubes we found that 48 h pre-treatment with atorvastatin (AT) significantlyinhibits constitutive IL-6 secretion at high (1 ?M) and supra (10?M and 100 ?M) therapeutic concentrations. At these AT concentrations, LPS-stimulated IL-6 secretion was also significantly reduced by 48 h AT co- treatment or pre-treatment, but not by post-treatment; therapeutic (0.1 ?M) ATconcentration was efficient only in pre-treatment but not in co- treatment or post-treatment LPS protocols. This information is an important clue for theinvestigations of the molecular mechanisms underlying AT effects and its therapeutic applications.
COBISS.SI-ID: 29489625
Contraction stimulates Na+,K+-ATPase and AMP-activated protein kinase (AMPK) activity in skeletal muscle. Whether AMPK activation affects Na+,K+-ATPase activity in skeletal muscle remains to be determined. Short-term stimulation of rat L6 myotubes with the AMPK activator AICAR, activates AMPK and promotes translocation of the Na+,K+-ATPase alpha1-subunit to the plasma membrane and increases Na+,K+-ATPase activity as assessed by ouabain-sensitive 86Rb+-uptake. Cyanide-induced artificial anoxia, as well as a direct AMPK activator (A-769662) also increases AMPK phosphorylation and Na+,K+-ATPase activity. Thus, different stimuli that target AMPK concomitantly increase Na+,K+-ATPase activity. The effect of AICAR on Na+,K+-ATPase in L6 myotubes was attenuated by Compound C, an AMPK inhibitor, as well as siRNA-mediated AMPK silencing. The effects of AICAR on Na+,K+-ATPase were completely abolished in cultured primary mouse muscle cells lacking AMPK alpha-subunits. AMPK stimulation leads to Na+,K+-ATPase alpha1-subunit dephosphorylation at Ser18, which may prevent endocytosis of the sodium pump. AICAR stimulation leads to methylation and dephosphorylation of the catalytic subunit of the protein phosphatase (PP) 2A in L6 myotubes. Moreover, AICAR-triggered dephosphorylation of the Na+,K+-ATPase was prevented in L6 myotubes deficient in PP2A-specific protein phosphatase methylesterase-1 (PME-1), indicating a role for the PP2A/PME-1 complex in AMPK-mediated regulation of Na+,K+-ATPase. Thus contrary to the common paradigm, we report AMPK-dependent activation of an energy-consuming ion pumping process. This activation may be a potential mechanism by which exercise and metabolic stress activate the sodium pump in skeletal muscle.
COBISS.SI-ID: 29990617
Acetylcholinesterase (AChE) and agrin play unique functional roles in the neuromuscular junction (NMJ). AChE is a cholinergic and agrin a synaptogenetic component. In spite of their different functions, they share several common features: their targeting is determined by alternative splicing; unlike most other NMJ components they are expressed in both, muscle and motor neuron and both reside on the synaptic basal lamina of the NMJ. Also, both were reported to play various nonjunctional roles. However, while the origin of basal lamina bound agrin is undoubtedly neural, the neural origin of AChE, which is anchored to the basal lamina with collagenic tail ColQ, is elusive. Hypothesizing that motor neuron proteins targeted to the NMJbasal lamina share common temporal pattern of expression, which is coordinated with the formation of basal lamina, we compared expression of agrin isoforms with the expression of AChE-T and ColQ in the developing rat spinal cord at the stages before and after the formation of NMJ basal lamina. Cellular origin of AChE-T and agrin was determined by in situ hybridization and their quantitative levels by RT PCR. We found parallel increase in expression of the synaptogenetic (agrin 8) isoform of agrin and ColQ after theformation of basal lamina supporting the view that ColQ bound AChE and agrin 8 isoform are destined to the basal lamina. Catalytic AChE-T subunit and agrin isoforms 19 and 0 followed different expression patterns. In accordance with the reports of other authors, our investigations also revealed various alternative functions for AChE and agrin. We have already demonstrated participation of AChE in myoblast apoptosis; here we present the evidence that agrin promotes the maturation of heavy myosin chains and the excitation-contraction coupling. These results show that common features of AChE and agrin extend to their capacity to play multiple roles in muscle development.
COBISS.SI-ID: 30270681