Effects of thallium(I) and thallium(III) on liposome membrane physical properties

The hypothesis that thallium (Tl) interaction with membrane phospholipids could result in the alteration of membrane physical properties was investigated. Working with liposomes composed of brain phosphatidylcholine and phosphatidylserine, we found that Tl(+), Tl(3+), and Tl(OH)(3) (0.5-25 microM): (a) increased membrane surface potential, (b) decreased the fluidity of the anionic regions of the membrane, in association with an increased fluidity in the cationic regions, and (c) promoted the rearrangement of lipids through lateral phase separation. The magnitude of these effects followed the order Tl(3+), Tl(OH)(3)>Tl(+). In addition, Tl(3+) also decreased the hydration of phospholipid polar headgroups and induced membrane permeabilization. The present results show that Tl interacts with membranes inducing major alterations in the rheology of the bilayer, which could be partially responsible for the neurotoxic effects of this metal.     

The localization of the different forms of troponin I in skeletal and cardiac muscle cells

1. 1. Antibodies raised against troponin I isolated from human cardiac and rabbit fast and slow skeletal muscles have been shown to be specific for the polymorphic forms of troponin I against which they were raised, i.e. they are tissue specific.

2. 2. These antibodies reacted with the polymorphic forms of troponin I, against which they were raised, that are present in tissues of other species such as the rhesus monkey, hamster and rat, i.e. they were species non-specific.

3. 3. Using the immunoperoxidase staining technique it has been shown that the fast and slow forms of troponin I are located in different cells in virtually all adult normal muscles examined.

4. 4. By comparison of the ATPase staining of skeletal muscle sections at pH 9.4 and 4.2 it is concluded that the fast form of troponin I is located in type II fibres and the slow form in type I fibres.

5. 5. It is suggested that immunoperoxidase staining with the antibodies to the fast and slow forms of troponin I provides an unambiguous new method of muscle fibre typing.

     

Proteomic responses of skeletal and cardiac muscle to exercise

Regular exercise is effective in the prevention of chronic diseases and confers a lower risk of death in individuals displaying risk factors such as hypertension and dyslipidemia. Thus, knowledge of the molecular responses to exercise provides a valuable contrast for interpreting investigations of disease and can highlight novel therapeutic targets. While exercise is an everyday experience and can be conceptualized in simple terms, it is also a complex physiological phenomenon and investigation of exercise responses requires sophisticated analytical techniques and careful standardization of the exercise stimulus. Proteomic investigation of exercise is in its infancy but the ability to link changes in function with comprehensive changes in protein expression and post-translational modification holds great promise for advancing physiology. This article highlights recent pioneering work investigating the effects of exercise in skeletal and cardiac muscle that has uncovered novel mechanisms underlying the benefits of physical activity.     

The calcium-ryanodine receptor complex of skeletal and cardiac muscle

[3H]Ryanodine binds with high affinity to saturable and Ca2+-dependent sites in heavy sarcoplasmic reticulum (SR) preparations from rabbit skeletal and cardiac muscle. Ruthenium red, known to interfere with Ca2+-induced Ca2+ release from SR vesicles, inhibits [3H]ryanodine specific binding in both skeletal and cardiac preparations whereas Mg2+, Ba2+, Cd2+ and La3+ selectively inhibit the skeletal preparation. The toxicological relevance of the [3H]ryanodine binding site is established by the correlation of binding inhibition with toxicity for seven ryanoids including two botanical insecticides. These findings provide direct evidence for Ca2+-ryanodine receptor complexes that may play a role in excitation-contraction coupling.     

Characterization of recombinant skeletal muscle (Ser-2843) and cardiac muscle (Ser-2809) ryanodine receptor phosphorylation mutants

Phosphorylation of the skeletal muscle (RyR1) and cardiac muscle (RyR2) ryanodine receptors has been reported to modulate channel activity. Abnormally high phosphorylation levels (hyperphosphorylation) at Ser-2843 in RyR1 and Ser-2809 in RyR2 and dissociation of FK506-binding proteins from the receptors have been implicated as one of the causes of altered calcium homeostasis observed during human heart failure. Using site-directed mutagenesis, we prepared recombinant RyR1 and RyR2 mutant receptors mimicking constitutively phosphorylated and dephosphorylated channels carrying a Ser/Asp (RyR1-S2843D and RyR2-S2809D) and Ser/Ala (RyR1-S2843A and RyR2-S2809A) substitution, respectively. Following transient expression in human embryonic kidney 293 cells, the effects of Ca2+, Mg2+, and ATP on channel function were determined using single channel and [3H]ryanodine binding measurements. In both assays, neither the skeletal nor cardiac mutants showed significant differences compared with wild type. Similarly essentially identical caffeine responses were observed in Ca2+ imaging measurements. Co-immunoprecipitation and Western blot analysis showed comparable binding of FK506-binding proteins to wild type and mutant receptors. Finally metabolic labeling experiments showed that the cardiac ryanodine receptor was phosphorylated at additional sites. Taken together, the results did not support the view that phosphorylation of a single site (RyR1-Ser-2843 and RyR2-Ser-2809) substantially changes RyR1 and RyR2 channel function.     

The role of super-relaxed myosin in skeletal and cardiac muscle

The super-relaxed (SRX) state of myosin was only recently reported in striated muscle. It is characterised by a sub-population of myosin heads with a highly inhibited rate of ATP turnover. Myosin heads in the SRX state are bound to each other along the thick filament core producing a highly ordered arrangement. Upon activation, these heads project into the interfilament space where they can bind to the actin filaments. Thus far, the population and lifetimes of myosin heads in the SRX state have been characterised in rabbit cardiac, and fast and slow skeletal muscle, as well as in the skeletal muscle of the tarantula. These studies suggest that the role of SRX in cardiac and skeletal muscle regulation is tailored to their specific functions. In skeletal muscle, the SRX modulates the resting metabolic rate. Cardiac SRX represents a “reserve” of inactive myosin heads that may protect the heart during times of stress, e.g. hypoxia and ischaemia. These heads may also be called up when there is a sustained demand for increased power. The SRX in cardiac muscle provides a potential target for novel therapies.     

The localization of VAMP5 in skeletal and cardiac muscle

Vesicle-associated membrane protein 5 (VAMP5) is a member of the SNARE protein family, which is generally thought to regulate the docking and fusion of vesicles with their target membranes. This study investigated the expression and localization of the VAMP5 protein. Immunoblotting analyses detected the VAMP5 protein in skeletal muscle, heart, spleen, lung, liver, and kidney tissue, but not in brain or small intestine tissue. Through the immunofluorescence microscopy of skeletal muscle, we found that the expression level of VAMP5 varies among fibers. Most of the fibers with high expression levels of VAMP5 were categorized as type IIa fibers on the basis of their myosin heavy chain subtypes. In addition, the expression patterns of VAMP5 and glucose transporter 4 (GLUT4) were similar. In cardiac muscle, we determined that VAMP5 was localized to the vicinity of intercalated discs. These results suggest that VAMP5 plays local roles in membrane trafficking in skeletal and cardiac muscle.     

MicroRNAs in skeletal and cardiac muscle development

MicroRNAs (miRNAs) are a recently discovered class of small non-coding RNAs, which are approximately 22 nucleotides in length. miRNAs negatively regulate gene expression by translational repression and target mRNA degradation. It has become clear that miRNAs are involved in many biological processes, including development, differentiation, proliferation, and apoptosis. Interestingly, many miRNAs are expressed in a tissue-specific manner and several miRNAs are specifically expressed in cardiac and skeletal muscles. In this review, we focus on those miRNAs that have been shown to be involved in muscle development. Compelling evidences have demonstrated that muscle miRNAs play an important role in the regulation of muscle proliferation and differentiation processes. However, it appears that miRNAs are not essential for early myogenesis and muscle specification. Importantly, dysregulation of miRNAs has been linked to muscle-related diseases, such as cardiac hypertrophy. A mutation resulting in a gain-of-function miRNA target site in the myostatin gene leads to down regulation of the targeted protein in Texel sheep. miRNAs therefore are a new class of regulators of muscle biology and they might become novel therapeutic targets in muscle-related human diseases.     

Biochemical adaptation of cardiac and skeletal muscle to physical activity.

1. Female Wistar rats were randomly assigned to control (C) or exercising (T) groups and subsequently portioned into 1, 3, 5 and 10 day T and C groups. The T groups completed a progressive endurance running program. Biochemical indices of adaptation were measured in cardiac muscle and in plantaris and soleus muscles of C and T animals after their last exercise bout. 2. In cardiac muscle, myofibrillar ATPase activity was significantly elevated in the 3T (0.241 +/- 0.031) and 5T (0.242 +/- 0.013) groups (P less than or equal to 0.05) compared to their respective controls (3C = 0.187 +/- 0.015 and 5C = 0.190 +/- 0.007). 3. After 10 days of training cardiac myofibrillar ATPase activity was elevated by 17% but this was not significant (P greater than or equal to 0.05). 4. No changes in myofibrillar ATPase activity were seen in skeletal muscle (P greater than or equal to 0.05), however, hexokinase activity progressively increased and was significantly elevated in the 3T, 5T and 10T soleus and plantaris muscles of rats over controls (P less than or equal to 0.05). 5. Minimal nonsignificant changes were noted in the hexokinase activity of the hearts of all T groups (P greater than or equal to 0.05). 6. These results indicate that metabolic adaptation of the heart and skeletal muscles takes place after as little as three training sessions. 7. Although the adaptation of the skeletal muscles continually progresses, the adaptation of the heart appears to be transitory.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ganglioside-binding proteins in skeletal and cardiac muscle

Several ganglioside-binding proteins have been identified inguinea pig skeletal and cardiac muscle. In the cytosolic fractionsof both tissues, a 130-kD protein was found to have the highestpropensity to bind lucifer yellow CH-labelled G_M1. This bindingcould be abolished by prior incubation of the protein with G_M2.Polysialogangliosides including G_D1a, G_D1b, G_T1b, and G_Q1b wereless effective. The 130-kD protein migrated as a doublet withapparent isoelectric points (pI) of 6.3 and 6.5, respectively,in isoelectric focusing gel, but as a single species with anapparent M_r of 43000 in SDS-polyacrylamide gel. Both the ganglioside-bindingand the immunological properties of the 43-kD subunit proteinwere different from those of rabbit skeletal muscle actin. Cardiacmuscle extract also contained a 77-kD minor ganglioside-bindingprotein that was absent in skeletal muscle. This protein hadan apparent pI of 5.4 and migrated as a 39-kD species in SDSgels. By contrast, only the particulate fraction of skeletalmuscle was found to contain a 180-kD major ganglioside-bindingprotein. Binding of fluorescent G_M1 to this protein was blockedby pre-incubation of the protein with G_M1 or G_M2. The 180-kDprotein migrated as a 98-kD species in SDS gels. However, itspropensity to bind lucifer yellow CH-G_M1 was at least 10 timesgreater than that of rabbit skeletal muscle phosphorylase b(M_r = 97400). The apparent pI (6.5) of the 180-kD protein alsowas slightly higher than that of rabbit phosphorylase. Tissuedistribution studies revealed that both the 130-kD and the 180-kDmajor ganglioside-binding proteins were muscle specific. Itis, therefore, possible that these two proteins may play someunique roles in ganglioside-related functions in muscle tissues. gangliosides ganglioside-binding proteins muscle     

Ca2+ binding to skeletal muscle troponin C in skeletal and cardiac myofibrils

Ca2+ binding to skeletal muscle troponin C in skeletal or cardiac myofibrils was measured by the centrifugation method using 45Ca. The specific Ca2+ binding to troponin C was obtained by subtracting the amount of Ca2+ bound to the CDTA-treated myofibrils (troponin C-depleted myofibrils) from that to the myofibrils reconstituted with troponin C. Results of Ca2+ binding measurement at various Ca2+ concentrations showed that skeletal troponin C had two classes of binding sites with different affinity for Ca2+. The Ca2+ binding of low-affinity sites in cardiac myofibrils was about eight times lower than that in skeletal myofibrils, while the high-affinity sites of troponin C in skeletal or cardiac myofibrils showed almost the same affinity for Ca2+. The Ca2+ sensitivity of the ATPase activity of skeletal troponin C-reconstituted cardiac myofibrils was also about eight times lower than that of skeletal myofibrils reconstituted with troponin C. These findings indicated that the difference in the sensitivity to Ca2+ of the ATPase activity between skeletal and cardiac CDTA-treated myofibrils reconstituted with skeletal troponin C was mostly due to the change in the affinity for Ca2+ of the low-affinity sites on the troponin C molecule.     

Localization of calcium in skeletal and cardiac muscle

Summary The requirement of calcium (Ca²⁺) in the excitation-contraction coupling of both skeletal and cardiac muscle is well established. However, the exact location of the intracellular storage sites of Ca²⁺ is not firmly established. We report here on the ultrastructural distribution of Ca²⁺ in white and red skeletal muscle and in cardiac muscle of the rat using combined phosphate-pyroantimonate (PPA) and oxalate-pyroantimonate (OPA) procedures. The methods are based on (a) stabilization and/or trapping of Ca²⁺ during the primary fixation step in glutaraldehyde by potassium phosphate or oxalate; (b) subsequent wash-out of all non-trapped cations such as Na⁺ and Mg²⁺ in potassium phosphate or oxalate; (c) conversion of the complexed or trapped Ca²⁺ into an electron-dense calcium pyroantimonate salt in 100 m-thick tissue sections; and (d) wash-out of the excess potassium pyroantimonate at alkaline pH.With the OPA procedure, mitochondria of all muscle types showed little precipitate. The junctional sarcoplasmic reticulum was stongly reactive in relaxed white skeletal muscle, negative in contracted white fibres and negative in red skeletal and cardiac muscle, independent of the state of relaxation-contraction. Other organelles were essentially free of deposits.With the PPA method, the precipitate was almost exclusively confined to the sarcolemma and its T-tubular invaginations in cardiac and slow skeletal muscle, and was absent in fast skeletal muscle. Apart from occasional deposits in mitochondria, all other organelles were free of precipitate. The sarcolemma-associated deposits were clearly confined to the inner leaflet of the lipid bilayer. The amount of precipitate varied within the contraction cycle, relaxed cells possessing the highest density.Exposure of the tissue to La³⁺ resulted in the complete absence of sarcolemma-bound precipitate suggesting that the Ca²⁺ is exchangeable. Furthermore, these cytological data suggest a basic difference in Ca²⁺ storage between white skeletal muscle on the one hand, and red skeletal and cardiac muscle on the other.     

GATA elements control repression of cardiac troponin I promoter activity in skeletal muscle cells

Background  

We reported previously that the cardiac troponin I (cTnI) promoter drives cardiac-specific expression of reporter genes in cardiac muscle cells and in transgenic mice, and that disruption of GATA elements inactivates the cTnI promoter in cultured cardiomyocytes. We have now examined the role of cTnI promoter GATA elements in skeletal muscle cells.     

Supramolecularly linked linear polymers of thallium(I) dithiocarbamates: Steric influence on the supramolecular interactions of methyl and ethylcyclohexyl dithiocarbamates of thallium(I)

Spectral and single crystal X-ray structural studies on [Tl(mchdtc)]₂ (1) and [Tl(echdtc)]₂ (2) (where mchdtc = methylcyclohexyldithiocarbamate and echdtc = ethylcyclohexyldithiocarbamate) were carried out. Both the synthesized complexes were characterized by UV-Vis, fluorescence, IR, ¹H and ¹³C NMR spectra. IR spectra of the complexes show the contribution of the thioureide form to the structures. Both compounds show weak fluorescence. The bond valence sums calculated for the complexes support the highly covalent nature of the Tl-S interactions. A Tl····H short interaction observed in the methyl analogue is totally absent in (2) because of the change in conformation of the cyclohexyl ring due to the introduction of ethyl group. Though the neighbouring non-bonded groups are flexible, thallium adjusts its thallophilic contacts to retain a hemisphere free for its pair of ‘s’ electrons in the presence of a sterically demanding ethyl group.     

The role of stem cells in skeletal and cardiac muscle repair.

In postnatal muscle, skeletal muscle precursors (myoblasts) can be derived from satellite cells (reserve cells located on the surface of mature myofibers) or from cells lying beyond the myofiber, e.g., interstitial connective tissue or bone marrow. Both of these classes of cells may have stem cell properties. In addition, the heretical idea that post-mitotic myonuclei lying within mature myofibers might be able to re-form myoblasts or stem cells is examined and related to recent observations for similar post-mitotic cardiomyocytes. In adult hearts (which previously were not considered capable of repair), the role of replicating endogenous cardiomyocytes and the recruitment of other (stem) cells into cardiomyocytes for new cardiac muscle formation has recently attracted much attention. The relative contribution of these various sources of precursor cells in postnatal muscles and the factors that may enhance stem cell participation in the formation of new skeletal and cardiac muscle in vivo are the focus of this review. We concluded that, although many endogenous cell types can be converted to skeletal muscle, the contribution of non-myogenic cells to the formation of new postnatal skeletal muscle in vivo appears to be negligible. Whether the recruitment of such cells to the myogenic lineage can be significantly enhanced by specific inducers and the appropriate microenvironment is a current topic of intense interest. However, dermal fibroblasts appear promising as a realistic alternative source of exogenous myoblasts for transplantation purposes. For heart muscle, experiments showing the participation of bone marrow-derived stem cells and endothelial cells in the repair of damaged cardiac muscle are encouraging.     

The interaction of bovine pancreatic deoxyribonuclease I and skeletal muscle actin

The rate of exchange of actin-bound nucleotide is decreased by a factor of about 20 when actin is complexed with DNAase I without affecting the binding constant of calcium for actin. Binding constants of DNAase I to monomeric and filamentous actin were determined to be 5 X 10(8) M-1 and 1.2 X 10(4) M-1 respectively. The depolymerisation of F-actin by DNAase I appears to be due to a shift in the G-F equilibrium of actin by DNAase I. Inhibition of the DNA-degrading activity of DNAase I by G-actin is of the partially competitive type.