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F Wuytack

Publications and source records attributed to F Wuytack.

At least 19 recordsLinked to original sources
PubMed2006

New perspectives on the role of SERCA2's Ca2+ affinity in cardiac function.

Cardiomyocyte relaxation and contraction are tightly controlled by the activity of the cardiac sarco(endo)plasmic reticulum (SR) Ca2+ transport ATPase (SERCA2a). The SR Ca2+ -uptake activity not only determines the speed of Ca(2+) removal during relaxation, but also the SR Ca2+ content and therefore the amount of Ca2+ released for cardiomyocyte contraction. The Ca2+ affinity is the major determinant of the pump's activity in the physiological Ca2+ concentration range. In the heart, the affinity of the pump for Ca2+ needs to be controlled between narrow borders, since an imbalanced affinity may evoke hypertrophic cardiomyopathy. Several small proteins (phospholamban, sarcolipin) adjust the Ca2+ affinity of the pump to the physiological needs of the cardiomyocyte. It is generally accepted that a chronically reduced Ca2+ affinity of the pump contributes to depressed SR Ca2+ handling in heart failure. Moreover, a persistently lower Ca2+ affinity is sufficient to impair cardiomyocyte SR Ca2+ handling and contractility inducing dilated cardiomyopathy in mice and humans. Conversely, the expression of SERCA2a, a pump with a lower Ca2+ affinity than the housekeeping isoform SERCA2b, is crucial to maintain normal cardiac function and growth. Novel findings demonstrated that a chronically increased Ca2+ affinity also may trigger cardiac hypertrophy in mice and humans. In addition, recent studies suggest that some models of heart failure are marked by a higher affinity of the pump for Ca2+, and hence by improved cardiomyocyte relaxation and contraction. Depressed cardiomyocyte SR Ca2+ uptake activity may therefore not be a universal hallmark of heart failure.

Alternative Splicing↗
PubMed2005

Cytosolic Ca2+ signals depending on the functional state of the Golgi in HeLa cells.

The Golgi apparatus is, like the endoplasmic reticulum, an inositol-1,4,5-trisphosphate-sensitive Ca2+ store, but its role in setting up Ca2+ signals is not well understood. We have now measured histamine-induced Ca2+ signals in HeLa cells pretreated with brefeldin A, a fungal metabolite that leads to the fragmentation and subsequent disappearance of the Golgi apparatus by its reabsorption within the endoplasmic reticulum. Ca2+ responses in which the free cytoplasmic Ca2+ concentration returned to resting levels during the histamine stimulation (mainly baseline Ca2+ oscillations or a single Ca2+ peak) occurred more often in brefeldin A pretreated cells, resulting in a lower Ca2+ plateau in population measurements. The latencies before the onset of the Ca2+ signals were longer after brefeldin A pretreatment. These results suggest that the integrity of the Golgi apparatus contributes to the shaping of intracellular Ca2+ signals.

Brefeldin A↗
PubMed2004

Inositol trisphosphate producing agonists do not mobilize the thapsigargin-insensitive part of the endoplasmic-reticulum and Golgi Ca2+ store.

Non-mitochondrial intracellular Ca2+ stores contain both thapsigargin-sensitive sarco(endo)plasmic-reticulum Ca2+-ATPases (SERCA) and thapsigargin-insensitive secretory-pathway Ca2+-ATPases (SPCA1). We now have studied the Ca2+-release properties of the compartments associated with these pumps in intact, i.e. non-permeabilized, cells of different origin (HeLa, keratinocytes, 16HBE14o-, COS-1, A7r5) and with different approaches (45Ca2+ fluxes, Ca2+ imaging and measurements of the free luminal [Ca2+] in the endoplasmic-reticulum and the Golgi apparatus using targeted aequorin). Application of an extracellular agonist in the absence of thapsigargin induced in all cells a Ca2+ release from both the endoplasmic-reticulum and the Golgi apparatus. The agonists were not able to release Ca2+ in the presence of 10 microM thapsigargin, except in COS-1 cells overexpressing SPCA1, where this pump not only appeared in the Golgi compartment but also overflowed into the agonist-sensitive part of the endoplasmic-reticulum. We conclude that the subcompartments of the endoplasmic-reticulum and of the Golgi complex that endogenously express SPCA1 are insensitive to agonist stimulation.

Animals↗
PubMed2004

Calcium release from the Golgi apparatus and the endoplasmic reticulum in HeLa cells stably expressing targeted aequorin to these compartments.

Extracellular agonists mobilize Ca2+ from SERCA-comprising intracellular Ca2+ stores located in both the Golgi apparatus and the endoplasmic reticulum. Ca2+ release from both these compartments was studied in HeLa cells stably expressing the luminescent Ca2+ indicator aequorin specifically targeted to these compartments. Changes in lumenal [Ca2+] as detected by the aequorin measurements were correlated with parallel changes in total Ca2+ content of the stores. The latencies and initial rates of Ca2+ release from the Golgi apparatus and the endoplasmic reticulum were quite similar. However, maximal Ca2+ release measured with Golgi-targeted aequorin terminated faster than that from the endoplasmic reticulum. The rate and extent of Ca2+ depletion from both compartments correlated well with the peak amplitude of the cytosolic [Ca2+] rise. Time-course experiments further revealed that the peak of the cytosolic Ca2+ response occurred before the lumenal [Ca2+] reached its lowest level. We conclude that both the Golgi apparatus and the endoplasmic reticulum contribute to the rise in cytosolic [Ca2+] upon agonist stimulation, but the kinetics of the Ca2+ release are different.

Aequorin↗
PubMed2003

Similar Ca(2+)-signaling properties in keratinocytes and in COS-1 cells overexpressing the secretory-pathway Ca(2+)-ATPase SPCA1.

Mutations in the ubiquitously expressed secretory-pathway Ca(2+)-ATPase (SPCA1) Ca(2+) pump result in Hailey-Hailey disease, which almost exclusively affects the epidermal part of the skin. We have studied Ca(2+) signaling in human keratinocytes by measuring the free Ca(2+) concentration in the cytoplasm and in the lumen of both the Golgi apparatus and the endoplasmic reticulum. These signals were compared with those recorded in SPCA1-overexpressing and control COS-1 cells. Both the sarco(endo)plasmic-reticulum Ca(2+)-ATPase (SERCA) and SPCA1 can mediate Ca(2+) uptake into the Golgi stacks. Our results indicate that keratinocytes mainly used the SPCA1 Ca(2+) pump to load the Golgi complex with Ca(2+) whereas the SERCA Ca(2+) pump was mainly used in control COS-1 cells. Cytosolic Ca(2+) signals in keratinocytes induced by extracellular ATP or capacitative Ca(2+) entry were characterized by an unusually long latency reflecting extra Ca(2+) buffering by an SPCA1-containing Ca(2+) store, similarly as in SPCA1-overexpressing COS-1 cells. Removal of extracellular Ca(2+) elicited spontaneous cytosolic Ca(2+) transients in keratinocytes, similarly as in SPCA1-overexpressing COS-1 cells. With respect to Ca(2+) signaling keratinocytes and SPCA1-overexpressing COS-1 cells therefore behaved similarly but differed from control COS-1 cells. The relatively large contribution of the SPCA1 pumps for loading the Golgi stores with Ca(2+) in keratinocytes may, at least partially, explain why mutations in the SPCA1 gene preferentially affect the skin in Hailey-Hailey patients.

Animals↗
PubMed2002

Expression of a P-type Ca(2+)-transport ATPase in Bacillus subtilis during sporulation.

The open reading frame designated yloB in the genomic sequence of Bacillus subtilis encodes a putative protein that is most similar to the typically eukaryotic type IIA family of P-type ion-motive ATPases, including the endo(sarco)plasmic reticulum (SERCA) and PMR1 Ca(2+)-transporters, located respectively in the SERCA and the Golgi apparatus. The overall amino acid sequence is more similar to that of the Pmr1s than to the SERCAs, whereas the inverse is seen for the 10 amino acids that form the two Ca(2+)-binding sites in SERCA. Sporulating but not vegetative B. subtilis cells express the predicted protein, as shown by Western blotting and by the formation of a Ca(2+)-dependent phosphorylated intermediate. Half-maximal activation of phosphointermediate formation occurred at 2.5 microM Ca(2+). Insertion mutation of the yloB gene did not affect the growth of vegetative cells, did not prevent the formation of viable spores, and did not significantly affect 45Ca accumulation during sporulation. However, spores from knockouts were less resistant to heat and showed a slower rate of germination. It is concluded that the P-type Ca(2+)-transport ATPase from B. subtilis is not essential for survival, but assists in the formation of resistant spores. The evolutionary relationship of the transporter to the eukaryotic P-type Ca(2+)-transport ATPases is discussed.

Bacillus subtilis↗
PubMed2001

Replacement of the muscle-specific sarcoplasmic reticulum Ca(2+)-ATPase isoform SERCA2a by the nonmuscle SERCA2b homologue causes mild concentric hypertrophy and impairs contraction-relaxation of the heart.

The cardiac sarco(endo)plasmic reticulum Ca(2+)-ATPase gene (ATP2A2) encodes the following two different protein isoforms: SERCA2a (muscle-specific) and SERCA2b (ubiquitous). We have investigated whether this isoform specificity is required for normal cardiac function. Gene targeting in mice successfully disrupted the splicing mechanism responsible for generating the SERCA2a isoform. Homozygous SERCA2a(-/-) mice displayed a complete loss of SERCA2a mRNA and protein resulting in a switch to the SERCA2b isoform. The expression of SERCA2b mRNA and protein in hearts of SERCA2a(-/-) mice corresponded to only 50% of wild-type SERCA2 levels. Cardiac phospholamban mRNA levels were unaltered in SERCA2a(-/-) mice, but total phospholamban protein levels increased 2-fold. The transgenic phenotype was characterized by a approximately 20% increase in embryonic and neonatal mortality (early phenotype), with histopathologic evidence of major cardiac malformations. Adult SERCA2a(-/-) animals (adult phenotype) showed a reduced spontaneous nocturnal activity and developed a mild compensatory concentric cardiac hypertrophy with impaired cardiac contractility and relaxation, but preserved beta-adrenergic response. Ca(2+) uptake levels in SERCA2a(-/-) cardiac homogenates were reduced by approximately 50%. In isolated cells, relaxation and Ca(2+) removal by the SR were significantly reduced. Comparison of our data with those obtained in mice expressing similar cardiac levels of SERCA2a instead of SERCA2b indicate the importance of the muscle-specific SERCA2a isoform for normal cardiac development and for the cardiac contraction-relaxation cycle.

Alternative Splicing↗
PubMed2001

Ca2+ pools and cell growth. Evidence for sarcoendoplasmic Ca2+-ATPases 2B involvement in human prostate cancer cell growth control.

The present study demonstrates for the first time that intracellular calcium-ATPases and calcium pool content are closely associated with prostate cancer LNCaP cell growth. Cell growth was modulated by changing the amount of epidermal growth factor, serum, and androgene in culture media. Using the microspectrofluorimetric method with Fura-2 and Mag Fura-2 as probes, we show that in these cells, the growth rate is correlated with intracellular calcium pool content. Indeed, an increased growth rate is correlated with an increase in the calcium pool filling state, whereas growth-inhibited cells show a reduced calcium pool load. Using Western blotting and immunocytochemistry, we show that endoplasmic reticulum calcium pump expression is closely linked to LNCaP cell growth, and are a common target of physiological stimuli that control cell growth. Moreover, we clearly demonstrate that inhibition of these pumps, using thapsigargin, inhibits LNCaP cell growth and prevents growth factor from stimulating cell proliferation. Our results thus provide evidence for the essential role of functional endoplasmic reticulum calcium pumps and calcium pool in control of prostate cancer LNCaP cell growth, raising the prospect of new targets for the treatment of prostate cancer.

Blotting, Western↗
PubMed2001

The sarco-endoplasmic reticulum Ca2+ ATPase is required for development and muscle function in Caenorhabditis elegans.

The sarco-endoplasmic reticulum Ca(2+)-transport ATPase (SERCA) loads intracellular releasable Ca(2+) stores by transporting cytosolic Ca(2+) into the endoplasmic (ER) or sarcoplasmic reticulum (SR). We characterized the only SERCA homologue of the nematode Caenorhabditis elegans, which is encoded by the sca-1 gene. The sca-1 transcript is alternatively spliced in a similar mode as the vertebrate SERCA2 transcript, giving rise to two protein variants: CeSERCAa and CeSERCAb. These proteins showed structural and functional conservation to the vertebrate SERCA2a/b proteins. The CeSERCAs were primarily expressed in contractile tissues. Loss of CeSERCA through gene ablation or RNA interference resulted in contractile dysfunctioning and in early larval or embryonic lethality, respectively. Similar defects could be induced pharmacologically using the SERCA-specific inhibitor thapsigargin, which bound CeSERCA at a conserved site. The conservation of SERCA2 homologues in C. elegans will allow genetic and chemical suppressor analyses to identify promising drug targets and lead molecules for treatment of SERCA-related diseases such as heart disease.

Animals↗
PubMed2001

Baseline cytosolic Ca2+ oscillations derived from a non-endoplasmic reticulum Ca2+ store.

Cytosolic Ca(2+) oscillations can be due to cycles of release and re-uptake of internally stored Ca(2+). To investigate the nature of these Ca(2+) stores, we expressed the Pmr1 Ca(2+) pump of Caenorhabditis elegans in COS-1 cells and pretreated the cells with thapsigargin to prevent Ca(2+) uptake by the sarco(endo)plasmic reticulum Ca(2+)-ATPase. Pmr1 co-localized with the Golgi-specific 58K protein and was targeted to a Ca(2+) store that was less leaky for Ca(2+) than the endoplasmic reticulum and whose inositol trisphosphate receptors were less sensitive to inositol trisphosphate and ATP than those in the endoplasmic reticulum. ATP-stimulated Pmr1-overexpressing cells responded after a latency to extracellular Ca(2+) with a regenerative Ca(2+) signal, which could be prevented by caffeine. They also produced very stable ilimaquinone-sensitive baseline Ca(2+) spikes, even in the presence of thapsigargin. Such responses never occurred in non-transfected cells or in cells that overexpressed the type-1 sarco(endo)plasmic reticulum Ca(2+)-ATPase. Abortive Ca(2+) spikes also occurred in histamine-stimulated untransfected HeLa cells pretreated with thapsigargin, and they too were inhibited by ilimaquinone. We conclude that the Pmr1-induced Ca(2+) store, which probably corresponds to the Golgi compartment, can play a crucial role in setting up baseline Ca(2+) spiking.

Adenosine Triphosphate↗
PubMed2001

Regenerating soleus and extensor digitorum longus muscles of the rat show elevated levels of TNF-alpha and its receptors, TNFR-60 and TNFR-80.

We measured the mRNA and protein levels of tumor necrosis factor-alpha (TNF-alpha) and the transcript levels of its receptors (TNFR-60 and TNFR-80) in the rat soleus (slow twitch) and extensor digitorum longus (EDL; fast twitch) muscles regenerating from notexin-induced necrosis. On the first day after administration of the toxin, when most fibers were necrotic and invaded by inflammatory cells/macrophages, dramatic increases of transcript and protein levels of TNF-alpha and of the mRNA levels of its receptors were observed. The transcript levels of TNF-alpha and TNFR-60, but not of TNFR-80, showed a second but smaller increase at the time when newly formed muscle fibers became reinnervated. In situ hybridization showed that on day 1, during the phase of extensive necrosis, the transcript of TNF-alpha was abundantly present and on day 4 of regeneration it was most often seen in areas devoid of desmin. The mRNA level of TNF-alpha was not detectable in BC(3)H1- and C2C12-cultured myoblasts and it was low in freeze-injured muscle, corresponding to the relatively mild degree of inflammation elicited by freezing. Therefore, our results are most consistent with the view that inflammatory cells/macrophages are the main source of TNF-alpha.

Animals↗
PubMed2000

The Golgi PMR1 P-type ATPase of Caenorhabditis elegans. Identification of the gene and demonstration of calcium and manganese transport.

In recent years, it has been well established that the Ca(2+) concentration in the lumen of intracellular organelles is a key determinant of cell function. Despite the fact that essential functions of the Golgi apparatus depend on the Ca(2+) and Mn(2+) concentration in its lumen, little is known on the transport system responsible for ion accumulation. The Golgi ion pump PMR1 has been functionally studied only in yeast. In humans, mutations in the orthologous gene ATP2C1 cause Hailey-Hailey disease. We report here the identification of the PMR1 homologue in the model organism Caenorhabditis elegans and after ectopic expression the direct study of its ion transport in permeabilized COS-1 cells. The C. elegans genome is predicted to contain a single PMR1 orthologue on chromosome I. We found evidence for alternative splicing in the 5'-untranslated region, but no indication for the generation of different protein isoforms. C. elegans PMR1 overexpressed in COS-1 cells transports Ca(2+) and Mn(2+) with high affinity into the Golgi apparatus in a thapsigargin-insensitive manner. Part of the accumulated Ca(2+) can be released by inositol 1,4,5-trisphosphate, in agreement with the idea that the Golgi apparatus is an inositol 1,4,5-trisphosphate-sensitive Ca(2+) store.

Adenosine Triphosphatases↗
PubMed2000

Low temperature molecular adaptation of the skeletal muscle sarco(endo)plasmic reticulum Ca2+-ATPase 1 (SERCA 1) in the wood frog (Rana sylvatica).

We have compared the primary sequence and enzymatic properties of the sarcoplasmic reticulum Ca(2+)-ATPases from a cold-tolerant frog Rana sylvatica with those of a closely related cold-intolerant frog, Rana clamitans. Sarcoplasmic reticulum isolated from leg muscles of both species contains a major protein ( approximately 100 kDa) that reacts with a monoclonal antibody against sarco(endo)plasmic reticulum Ca(2+)-ATPase type 1 (SERCA1). The apparent molecular mass of R. sylvatica SERCA1 is 115 kDa, whereas that of R. clamitans is 105 kDa. However, the deduced amino acid sequences obtained from cDNAs do not indicate a difference in molecular weight, thus suggesting post-translational protein modification of R. sylvatica SERCA1. Comparison of the temperature dependence of both ATP hydrolysis and Ca(2+) transport indicates that R. sylvatica SERCA1 exhibits significantly lower activation energy below 20 degrees C and an approximately 2-fold greater Ca(2+)-ATPase activity near 0 degrees C. Furthermore, R. sylvatica SERCA1 exhibits simple Michaelis-Menten kinetics with ATP and Ca(2+) as opposed to the two-site ATP kinetics and positive cooperativity with Ca(2+) observed for R. clamitans and mammalian SERCA1s. Cooperativity has been linked to protein-protein interaction in SERCA1, and this property may be altered in R. sylvatica SERCA1. Primary sequence comparison shows that R. sylvatica SERCA1 exhibits seven unique amino acid substitutions, three of which are in the ATP binding domain. We also report for the first time the presence of alternative splicing in the frog, resulting in isoforms SERCA1a and SERCA1b. Thus, it appears that the low temperature muscle contractility of R. sylvatica can be explained partially by significant functional and structural differences in SERCA1.

Adaptation, Physiological↗
PubMed2000

Presence of functional sarcoplasmic reticulum in the developing heart and its confinement to chamber myocardium.

During development fast-contracting atrial and ventricular chambers develop from a peristaltic-contracting heart tube. This study addresses the question of whether chamber formation is paralleled by a matching expression of the sarcoplasmic reticulum (SR) Ca(2+) pump. We studied indo-1 Ca(2+) transients elicited by field stimulation of linear heart tube stages and of explants from atria and outflow tracts of the prototypical preseptational E13 rat heart. Ca(2+) transients of H/H 11+ chicken hearts, which constitute the prototypic linear heart tube stage, were sensitive to verapamil only, indicating a minor contribution of Ca(2+)-triggered SR Ca(2+) release. Outflow tract transients displayed sensitivity to the inhibitors similar to that of the linear heart tube stages. Atrial Ca(2+) transients disappeared upon addition of ryanodine, tetracaine, or verapamil, indicating the presence of Ca(2+)-triggered SR Ca(2+) release. Quantitative radioactive in situ hybridization on sections of E13 rat hearts showed approximately 10-fold higher SERCA2a mRNA levels in the atria compared to nonmyocardial tissue and approximately 5-fold higher expression in compact ventricular myocardium. The myocardium of atrioventricular canal, outflow tract, inner curvature, and ventricular trabecules displayed weak expression. Immunohistochemistry on sections of rat and human embryos showed a similar pattern. The significance of these findings is threefold. (i) A functional SR is present long before birth. (ii) SR development is concomitant with cardiac chamber development, explaining regional differences in cardiac function. (iii) The pattern of SERCA2a expression underscores a manner of chamber development by differentiation at the outer curvature, rather than by segmentation of the linear heart tube.

Animals↗
PubMed2000

Corticosteroids decrease mRNA levels of SERCA pumps, whereas they increase sarcolipin mRNA in the rat diaphragm.

1. In order to explore the potential role of the sarcoplasmic-endoplasmic reticulum Ca2+-ATPase (SERCA)-type pumps and of their modulators phospholamban (PLB) and sarcolipin (SLN) in the functional alterations of the diaphragm induced by corticosteroid treatment, expression of SERCA, PLB and SLN was assessed by RT-PCR in the diaphragm of rats treated daily for 5 days either with triamcinolone (80 mg kg-1, n = 8) or with saline (control; 0.6 ml, n = 8). 2. Triamcinolone treatment reduced the normalised overall amount of all SERCA mRNA in diaphragm by 70 % compared to controls (P < 0.05). This reduction was accounted for by a relatively larger decrease in the SERCA1 mRNA (-69 %, P < 0.05) whilst the decrease in SERCA2 mRNA (-49 %, P = 0.09) did not reach statistical significance. As a result the relative proportion of SERCA2 mRNA was increased from 43 +/- 7 % in control diaphragm to 52 +/- 4 % after triamcinolone treatment (P < 0.05). 3. Only the adult isoform of SERCA1 (i.e. SERCA1a) mRNA was found in the diaphragm of the 15-week-old control rats. Furthermore, triamcinolone treatment resulted in reduced levels of SERCA2a (-40 %, P < 0.05) and increased levels of SLN mRNA (+100 %, P < 0.05), while the decrease in PLB mRNA (-31 %, P = 0.277) did not reach statistical significance. SERCA1b, SERCA2b and SERCA3 mRNA levels fell below the detection limit in the diaphragm of both control and triamcinolone-treated rats. 4. Compared to control diaphragm, control rat heart showed a relatively high PLB/(SERCA1 + SERCA2) mRNA ratio of 7.88 while this ratio amounted only to 0.16 in control extensor digitorum longus (EDL) muscle. Remarkably, the SLN/(SERCA1 + SERCA2) mRNA ratio in normal cardiac muscle (0.96) was nearly the same as in diaphragm, but in EDL it amounted to only 0.05 that in diaphragm. This indicates the very low expression of SLN in rat EDL. 5. These data reveal that considerable alterations in SERCA mRNA levels accompany the functional changes seen in diaphragm after corticosteroid treatment. The relatively larger decrease in SERCA1 mRNA is in agreement with the selective type II fibre atrophy previously observed in the diaphragm of triamcinolone-treated rats, but the magnitude of SERCA alterations is more pronounced than expected on the basis of the structural changes in the diaphragm. The increase in SLN mRNA levels may represent a compensatory mechanism.

Adrenal Cortex Hormones↗
PubMed2000

The expression of SR calcium transport ATPase and the Na(+)/Ca(2+)Exchanger are antithetically regulated during mouse cardiac development and in Hypo/hyperthyroidism.

The mouse has been used extensively for generating transgenic animal models to study cardiovascular disease. Recently, a number of transgenic mouse models have been created to investigate the importance of sarcoplasmic reticulum (SR) Ca(2+)transport proteins in cardiac pathophysiology. However, the expression and regulation of cardiac SR Ca(2+)ATPase and other Ca(2+)transport proteins have not been studied in detail in the mouse. In this study, we used multiplex RNase mapping analysis to determine SERCA2, phospholamban (PLB), and Na(+)/Ca(2+)-exchanger (NCX-1) gene expression throughout mouse heart development and in hypo/hyperthyroid animals. Our results demonstrate that the expression of SERCA2 and PLB mRNA increase eight-fold from fetal to adult stages, indicating that SR function increases with heart development. In contrast, the expression of the Na(+)/Ca(2+)-exchanger gene is two-fold higher in fetal heart compared to adult. Our study also makes the important observation that in hypothyroidic hearts the NCX-1 mRNA and protein levels were upregulated, whereas the SERCA2 mRNA/protein levels were downregulated. In hyperthyroidic hearts, however, an opposite response was identified. These findings are important and point out that the expression of NCX-1 is regulated antithetically to that of SERCA2 during heart development and in response to alterations in thyroid hormone levels.

Animals↗
PubMed2000

Myostatin levels in regenerating rat muscles and in myogenic cell cultures.

Myostatin is a newly described member of the TGF-beta superfamily acting as a secreted negative regulator of skeletal muscle mass in several species, but whose mode of action remains largely unknown. In the present work, we followed the myostatin mRNA and protein levels in rat soleus and extensor digitorum longus (EDL) muscles regenerating in vivo from notexin-induced necrosis, and the myostatin transcript levels in two different in vitro myogenic differentiation models: i.e. in mouse BC3H1 and C2Cl2 cultured cells. The in vivo regenerating rat skeletal muscles showed a characteristic time-dependent expression of myostatin mRNA. After notexin injection, the transcript levels dropped below the detection limit on day 1 in soleus and close to the detection limit on day 3 in EDL, then increased to a maximum on day 7 in soleus and after 28 days finally reached the control values in both types of muscles. In contrast, the myostatin protein levels increased dramatically on the first days of regeneration in both muscles, i.e. at the time when its transcript level was low. Later on the myostatin protein level gradually declined to normal in soleus while in EDL it stayed high some days longer and decreased to normal on days 21-28. In vitro proliferating myoblasts produced low level of myostatin mRNA, which increased upon induction of differentiation suggesting that functional innervation is no prerequisite for myostatin expression. Myostatin production in vitro seems not to be dependent on myocyte fusion either, since it is observed in differentiated BC3H1 cells, which are defective in myofiber formation.

Animals↗