Biomedical subjects

Stéphane König

Publications and source records attributed to Stéphane König.

5 recordsLinked to original sources

PubMed2006

Calcium sources used by post-natal human myoblasts during initial differentiation.

Increases in cytoplasmic Ca(2+) are crucial for inducing the initial steps of myoblast differentiation that ultimately lead to fusion; yet the mechanisms that produce this elevated Ca(2+) have not been fully resolved. For example, it is still unclear whether the increase comes exclusively from membrane Ca(2+) influx or also from Ca(2+) release from internal stores. To address this, we investigated early differentiation of myoblast clones each derived from single post-natal human satellite cells. Initial differentiation was assayed by immunostaining myonuclei for the transcription factor MEF2. When Ca(2+) influx was eliminated by using low external Ca(2+) media, we found that approximately half the clones could still differentiate. Of the clones that required influx of external Ca(2+), most clones used T-type Ca(2+) channels, but others used store-operated channels as influx-generating mechanisms. On the other hand, clones that differentiated in low external Ca(2+) relied on Ca(2+) release from internal stores through IP(3) receptors. Interestingly, by following clones over time, we observed that some switched their preferred Ca(2+) source: clones that initially used calcium release from internal stores to differentiate later required Ca(2+) influx and inversely. In conclusion, we show that human myoblasts can use three alternative mechanisms to increase cytoplasmic Ca(2+) at the onset of the differentiation process: influx through T-types Ca(2+) channels, influx through store operated channels and release from internal stores through IP(3) receptors. In addition, we suggest that, probably because Ca(2+) elevation is essential during initial differentiation, myoblasts may be able to select between these alternate Ca(2+) pathways.

Calcium↗

PubMed2004

Peptidyl-glycine alpha-amidating monooxygenase targeting and shaping of atrial secretory vesicles: inhibition by mutated N-terminal ProANP and PBA.

ANP (atrial natriuretic peptide) is widely recognized as an important vasorelaxant, diuretic, and cardioprotective hormone. Little is known, however, about how ANP-secretory vesicles form within the atrial myocytes. Secretory vesicles were visualized by fluorescence microscope imaging in live rat atrial myocytes expressing proANP-enhanced green fluorescent protein (EGFP), or N-terminal-mutated fusion proteins thought to suppress the calcium-dependent aggregation of proANP. Results showed the following: (1) aggregates of proANP and coexpressed proANP-EGFP recruited peptidylglycine alpha-amidating monooxygenase (PAM)-1, an abundant atrial integral vesicle membrane protein; (2) coexpressed N-terminal-mutated (Glu23,24-->Gln23,24) and N-terminal-deleted proANP-EGFP inhibited recruitment of PAM-1 by up to 60%; (3) 4-phenyl-3-butenoic acid (PBA) (10 mumol/L), a pharmacological inhibitor of the lumenal peptidylglycine alpha-hydroxylating monooxygenase domain of PAM proteins, inhibited recruitment of endogenous PAM-1 and of coexpressed pro-EGFP-PAM-1; (4) PBA had no effect on exocytosis of the potassium inward rectifier KIR2.1; (5) PBA induced a deformation of the secretory vesicles but did not inhibit docking. These findings suggest that recruitment of PAM-1 to secretory vesicles depends on intact N-terminal proANP and on the lumenal domain of PAM-1. Conversely, PAM-1 participates in shaping the proANP-secretory vesicles. The full text of this article is available online at http://circres.ahajournals.org.

Animals↗

PubMed2003

Lentivector-mediated transfer of Bmi-1 and telomerase in muscle satellite cells yields a duchenne myoblast cell line with long-term genotypic and phenotypic stability.

Conditionally immortalized human cells are valuable substrates for basic biologic studies, as well as for the production of specific proteins and for the creation of bioartificial organs. We previously demonstrated that the lentivector-mediated transduction of immortalizing genes into human primary cells is an efficient method for obtaining such cell lines. Here, we used human muscle satellite cells as model targets to examine the impact of the transduced genes on the genotypic and phenotypic characteristics of the immortalized cells. The most commonly used immortalizing gene, the SV40 large T antigen (T-Ag), was extremely efficient at inducing the continuous growth of primary myoblasts, but the resulting cells rapidly accumulated major chromosomal aberrations and exhibited profound phenotypic changes. In contrast, the constitutive expression of telomerase and Bmi-1 in satellite cells from a control individual and from a patient suffering from Duchenne's muscular dystrophy yielded cell lines that remained diploid and conserved their growth factor dependence for proliferation. However, despite the absence of detectable cytogenetic abnormalities, clones derived from satellite cells of a control individual exhibited a differentiation block in vitro. In contrast, a Duchenne-derived cell line exhibited all the phenotypic characteristics of its primary parent, including an ability to differentiate fully into myotubes when placed in proper culture conditions. This cell line should constitute a useful reagent for a wide range of studies aimed at this disease.

Adolescent↗

PubMed2003

Acceleration of human myoblast fusion by depolarization: graded Ca2+ signals involved.

We have previously shown that human myoblasts do not fuse when their voltage fails to reach the domain of a window T-type Ca(2+) current. We demonstrate, by changing the voltage in the window domain, that the Ca(2+) signal initiating fusion is not of the all-or-none type, but can be graded and is interpreted as such by the differentiation program. This was carried out by exploiting the properties of human ether-à-go-go related gene K(+) channels that we found to be expressed in human myoblasts. Methanesulfonanilide class III antiarrhythmic agents or antisense-RNA vectors were used to suppress completely ether-à-go-go related gene current. Both procedures induced a reproducible depolarization from -74 to -64 mV, precisely in the window domain where the T-type Ca(2+) current increases with voltage. This 10 mV depolarization raised the cytoplasmic free Ca(2+) concentration, and triggered a tenfold acceleration of myoblast fusion. Our results suggest that any mechanism able to modulate intracellular Ca(2+) concentration could affect the rate of myoblast fusion.

Calcium↗

PubMed2001

Vascular cell adhesion molecule-1 induced by human T-cell leukaemia virus type 1 Tax protein in T-cells stimulates proliferation of human T-lymphocytes.

Human T-cell leukaemia/lymphotropic virus type 1 (HTLV-1), aetiologically linked to lymphoproliferative as well as inflammatory diseases, infects and activates CD4(+) helper T-cells and thus alters immunoregulatory pathways. The viral regulatory Tax protein has been shown previously to induce the expression of vascular cell adhesion molecule-1 (VCAM-1) by T-cells. To determine the functional role of this adhesion molecule, Jurkat T-cells stably expressing either Tax or both Tax and Rex (another viral regulatory protein) were used in binding and coculture assays performed with either control Jurkat cells or primary human T-lymphocytes. Evidence was provided that VCAM-1 acting in synergy with leucocyte function-associated antigen-3 promotes T-cell-T-cell interactions and increases T-cell proliferation. Interestingly, Rex was found to modulate these events. These data establish that VCAM-1 induced by Tax on T-cells thus contributes to the immunopathological process triggered by HTLV-1 infection.

CD58 Antigens↗