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J P Tremblay

Publications and source records attributed to J P Tremblay.

At least 19 recordsLinked to original sources
PubMed1992

Dystrophin-like immunoreactivity in monkey and human brain areas involved in learning and motor functions.

Two antidystrophin antibodies against different fragments of dystrophin were used to detect this polypeptide in monkey and human brains. Dystrophin was revealed by immunoperoxidase amplified with the biotin/avidin system and by immunoblotting. A dystrophin-like immunoreactivity was uniformly expressed in several brain regions implicated in learning and motor functions. Dystrophin function is not clear but our results raise the possibility that this protein may be involved in the cognitive impairment observed in several Duchenne muscular dystrophy (DMD) patients.

Animals↗
PubMed1992

Localization of dystrophin in the Purkinje cells of normal mice.

A monoclonal antibody that reacts with a mid rod fragment of dystrophin was used to localize this protein in the central nervous system (CNS). Due to a low abundance of dystrophin in the CNS, an immunoperoxidase reaction amplified with a biotin-avidin system was used. All Purkinje cells in normal mice were dystrophin positive while the mdx mouse cerebellum was completely devoid of reaction. Dystrophin staining was present in the soma and dendrites of Purkinje cells but not in their axons. This uniform dystrophin labelling in the normal mouse Purkinje cells indicates that this protein is not only localized in synaptic contact regions of the CNS.

Animals↗
PubMed1992

Human myoblast transplantation: preliminary results of 4 cases.

Myoblasts from immunocompatible donors have been transplanted into the muscles (tibialis anterior, biceps brachii, and/or extensor carpi radialis longus) of 4 Duchenne patients in the advanced stages of the disease. Although no immunosuppressive treatment was used, none of the patients showed any clinical signs of rejection such as fever, redness, and inflammation. One patient transiently produced antibodies against the donor myoblasts as determined by cytofluorometric analysis. This patient and 2 others were shown to form antibodies against their donor's myotubes. Muscle biopsies of the injected tibialis anterior of 4 patients revealed that 80%, 75%, 25%, and 0% of the muscle fibers, respectively, showed some degree of dystrophin immunostaining. The contralateral noninjected muscles of the latter 3 patients did not contain any dystrophin positive fibers, while that of the first patient showed dystrophin expression in 16% of the fibers examined. Myoblasts were also injected into the extensor carpi radialis longus or the biceps brachii of these patients. A few months subsequent to injection, one patient was shown to have a 143% increase of strength during static wrist extension. This result must be interpreted with caution because a double-blind strength-measuring protocol was not used. Furthermore, we have noted that this change slowly decayed over time. The strength of 2 other patients was increased less remarkably (41% and 51%), while the strength of the fourth patient was unchanged.

Adolescent↗
PubMed1992

A light and electron microscopic study of dystrophin localization at the mouse neuromuscular junction.

Duchenne muscular dystrophy (DMD) is characterized by a lack of dystrophin expression. Dystrophin is a 420 Kd protein localized in the muscle sarcolemma that most likely provides stability to the muscle plasma membrane. Neuromuscular junctions (NMJs) were localized by revealing either the acetylcholine receptors (AChRs) with alpha-bungarotoxin coupled with cascade blue or by revealing desmin, a protein found in higher concentration at the NMJs using immunochemistry. An accumulation of dystrophin was observed in normal mice by immunoperoxidase labelling at NMJs identified with these markers. Dystrophin was pinpointed on the postjunctional folds of NMJs by electron microscopy and was more abundant on the postjunctional membrane than on the remaining muscle membrane. Our observations are similar to previous observations suggesting that dystrophin may interact with the AChRs.

Animals↗
PubMed1992

Mosaic expression of dystrophin in the cerebellum of heterozygote dystrophic (mdx) mice.

The monoclonal NCLDys1 revealed the presence of dystrophin in the Purkinje cells of normal mice but not of mdx mice and a mosaic staining in Purkinje cells of heterozygote mdx mice. Dystrophin was present in the soma and the dendrites of the dystrophin positive Purkinje cells and was absent in both regions of the dystrophin negative Purkinje cells. However, the polyclonal antibody d10 produced a uniform labeling of all Purkinje cells not only in the normal mice but also in mdx and heterozygote mdx mice. This staining was attributed to a reaction of this antibody not only with dystrophin but also with a different isoform of dystrophin or with a dystrophin related protein present even in mdx mice.

Animals↗
PubMed1991

Dystrophin expression in myotubes formed by the fusion of normal and dystrophic myoblasts.

Mdx mouse dystrophy is characterized by the absence in the muscle cytoplasmic membrane of a high molecular weight protein called dystrophin. A possible avenue for treatment of muscular dystrophies is to inject normal myoblasts in a dystrophic muscle to form hybrid muscle fibers. Hybrid myotubes were formed in vitro by the fusion of normal rat and dystrophic mouse (mdx) myoblasts. Staining with Hoechst dye 33258 permitted the clear distinction of mouse and rat nuclei. Immunostaining demonstrated that dystrophin was present over the entire membrane of all hybrid myotubes even when nuclei ratio normal/dystrophic was low.

Animals↗
PubMed1991

Is dystrophin present in the nerve terminal at the neuromuscular junction? An immunohistochemical study of the heterozygote dystrophic (mdx) mouse.

Neuromuscular junctions (NMJs) were identified by revealing the presence of cholinergic receptors (AChR) with alpha-bungarotoxin coupled to the fluorescent dye cascade blue in 9- and 60-day-old normal and heterozygote mdx mice. Dystrophin was detected by an immunoperoxidase technique. All the muscle fibers of the normal animals observed in cross sections were immunoreactive for dystrophin and an accumulation of dystrophin was observed at all NMJs identified by alpha-bungarotoxin. In the 9-day-old mdx heterozygote animals, dystrophin positive, negative, and partially positive muscle cross sections were observed. Four different observations were made in these heterozygote animals on the coexistence of AChR and dystrophin. First, alpha-bungarotoxin sites (i.e., NMJs) were observed on dystrophin positive muscle fiber cross sections with an accumulation of dystrophin at these sites. Second, alpha-bungarotoxin sites were observed on dystrophin positive fibers without a dystrophin accumulation at NMJs. Third, there was a coexistence of alpha-bungarotoxin and dystrophin labelling at NMJs of muscle fibers with perimeters labelling negative for dystrophin. Fourth, NMJs, identified by alpha-bungarotoxin, were observed on muscle fibers negative for dystrophin even at the NMJ. These observations suggest that dystrophin is present not only in the muscle membrane but also in the presynaptic nerve terminals.

Acetylcholinesterase↗
PubMed1991

A monoclonal antibody to conotoxin reveals the distribution of a subset of calcium channels in the rat cerebellar cortex.

Voltage-sensitive calcium channels (VSCC) are a family of ionophores having different electrical and pharmacological properties. The omega-conotoxin GVIA (omega-CgTX) is a specific blocker of one subset of VSCCs. Because of the specificity of this toxin, a monoclonal anti-omega-CgTX antibody was generated against a omega-CgTX-key hole limpet hemocyanin conjugate and used as a specific marker to study VSCC distributions. This mab was shown to recognize omega-CgTX on Western blots and to display omega-CgTX-dependent immunoperoxidase staining of rat cerebellum. Incubation of fresh, unfixed sections of adult rat cerebellum in omega-CgTX followed by light fixation and peroxidase immunocytochemistry with mab anti-omega-CgTX revealed a specific pattern of labelling. All principal classes of cerebellar neurons were immunoreactive, but in general glial cells were not stained. Most interestingly, strong focal immunoreactivity was encountered at branching points of Purkinje cell dendrites. This characteristic staining pattern implies that a subset of VSCC is specifically concentrated in these regions and suggests that these channels may play a role in the functional integration of dendritic signals.

Animals↗
PubMed1991

Non-uniform responses to Ca2+ along the frog neuromuscular junction: effects on the probability of spontaneous and evoked transmitter release.

Spontaneous and evoked transmitter release activity was studied during selective application of Ca2+ in proximal (near the first contact of the axon on the muscle fiber) and distal regions of the frog neuromuscular junction. A new technique called "Microperfusion" was developed, which allowed us to apply a 30-microns-wide Ca2+ stream from an external pipette. The spread of this Ca2+ stream was monitored by adding Blue Dextran (40 mg/ml) to the Ca2+ solution. Microperfusion with a Ca2(+)-free Ringer containing Blue Dextran did not affect the miniature endplate potential frequency or amplitude. Changes of spontaneous transmitter release were studied either during microperfusion of Ringer containing 5 mM Ca2+ or during microperfusion of 2 mM Ca2+ simultaneously with the stimulation of the motor nerve. This second procedure also permitted study of the characteristics of evoked release. Microperfusion of Ca2+ induced a larger and more rapid increase in the miniature endplate potential frequency in proximal than in distal regions. The time required for the miniature endplate potential frequency to return to the control value after Ca2+ microperfusion was longer than the time needed to increase the frequency and this decay period was longer in the proximal region than in the distal one. Moreover, miniature endplate potentials produced in proximal regions, were typically larger and more variable than those produced in distal regions. In five experiments, the endplate potentials produced by 100-200 pulse pairs (interval of 15 ms at every 2 s) were recorded intracellularly during the microperfusion. The quantal content of the first endplate potential of the pair (EPP1) was systematically smaller in distal regions than in proximal regions. The percentage of failures and the coefficients of variation were higher in distal than in proximal regions, indicating a larger variability of quantal content. The frequency facilitation was not different between the two regions, but, however, the second stimuli of the pair usually produced a net increase of transmitter release which was greater in proximal than distal regions. Our experiments demonstrate that both the spontaneous and the evoked release are more responsive to Ca2+ application in the proximal than in the distal regions of the frog neuromuscular junction.

Animals↗
PubMed1991

Human myoblast transplantation: a simple assay for tumorigenicity.

A simple assay for tumorigenicity of myoblasts to be transplanted to Duchenne patients has been developed. The assay is based on culture in a soft agar medium for 2-3 weeks. The tumor cell line forms large cell clusters while the normal myoblasts do not proliferate and remain isolated.

Cell Transformation, Neoplastic↗
PubMed1991

A new technique to identify hybrid myotubes in vitro without culture fixation.

Fluorescent latex microspheres (FLMs) were used to label myoblasts and to permit the observation of hybrid myotubes before culture fixation. This type of labeling did not affect survival, development, or fusion of these cells. The FLMs were retained for several weeks. Labeled mouse myoblasts were co-cultured with unlabeled rat myoblasts to verify whether the marker was released and spread from labeled to unlabeled cells. The nuclear stain Hoechst 33258 was used to distinguish the myoblasts from both species and permitted the demonstration that there was virtually no re-uptake. Hybrid myotubes were also obtained by co-culturing mouse myoblasts containing rhodamine FLMs and rat myoblasts containing green FLMs. These mixed cultures were observed repeatedly with a fluorescent microscope without any cytotoxic effect. Several myotubes were observed before fixation of the cultures to contain both types of fluorescent labels. Subsequent fixation and staining with Hoechst dye confirmed that these myotubes were hybrids.

Animals↗
PubMed1991

In vitro bromodeoxyuridine labeling of nuclei: application to myotube hybridization.

Rat myoblast nuclei were labeled with various concentrations of bromodeoxyuridine (BrdU), an analogue of thymidine, for 24 or 48 hr. Almost every myoblast was labeled with BrdU at concentrations between 10(-7) M and 10(-5) M. When the cells were labeled with 0.5 microM or more, the percentage of labeled cells remained over 90% and 80% at 2 and 5 days, respectively. However, when the cells were labeled with BrdU concentration lower than 10(-7) M the percentage of labeled nuclei decreased more rapidly with time. The BrdU-labeled cells were mixed with an unlabeled population to determine whether their capacity to fuse was reduced. At a BrdU concentration of 0.5 x 10(-6) M, labeled myoblasts fused to a similar extent as unlabeled myoblasts, and a high percentage of marked cells were still perceptively labeled after 5 days. In contrast, the fusion capacity of myoblasts incubated with more than 10(-6) M BrdU was inhibited after only few rounds of DNA synthesis. These myoblasts were eventually able to fuse, however, when the BrdU diminished in the DNA due to cell division. These results indicate that labeling with BrdU at a concentration of 0.5 x 10(-6) M and an incorporation time of 48 hr is optimal to obtain perceptible immunocytochemical staining without affecting myoblast fusion. Such BrdU immunolabeling could be used as a nuclear marker for hybridization studies.

Animals↗