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Emanuela Gussoni

Publications and source records attributed to Emanuela Gussoni.

10 recordsLinked to original sources
PubMed2006

C6ORF32 is upregulated during muscle cell differentiation and induces the formation of cellular filopodia.

We have identified a gene by microarray analysis that is located on chromosome 6 (c6orf32), whose expression is increased during human fetal myoblast differentiation. The protein encoded by c6orf32 is expressed both in myogenic and non-myogenic primary cells isolated from 18-week old human fetal skeletal muscle. Immunofluorescent staining indicated that C6ORF32 localizes to the cellular cytoskeleton and filopodia, and often displays polarized expression within the cell. mRNA knockdown experiments in the C2C12 murine myoblast cell line demonstrated that cells lacking c6orf32 exhibit a myogenic differentiation defect, characterized by a decrease in the expression of myogenin and myosin heavy chain (MHC) proteins, whereas MyoD1 was unaltered. In contrast, overexpression of c6orf32 in C2C12 or HEK293 cells (a non-muscle cell line) promoted formation of long membrane protrusions (filopodia). Analysis of serial deletion mutants demonstrated that amino acids 55-113 of C6ORF32 are likely involved in filopodia formation. These results indicate that C6ORF32 is a novel protein likely to play multiple functions, including promoting myogenic cell differentiation, cytoskeletal rearrangement and filopodia formation.

Base Sequence↗
PubMed2006

Regulation of myogenic progenitor proliferation in human fetal skeletal muscle by BMP4 and its antagonist Gremlin.

Skeletal muscle side population (SP) cells are thought to be "stem"-like cells. Despite reports confirming the ability of muscle SP cells to give rise to differentiated progeny in vitro and in vivo, the molecular mechanisms defining their phenotype remain unclear. In this study, gene expression analyses of human fetal skeletal muscle demonstrate that bone morphogenetic protein 4 (BMP4) is highly expressed in SP cells but not in main population (MP) mononuclear muscle-derived cells. Functional studies revealed that BMP4 specifically induces proliferation of BMP receptor 1a-positive MP cells but has no effect on SP cells, which are BMPR1a-negative. In contrast, the BMP4 antagonist Gremlin, specifically up-regulated in MP cells, counteracts the stimulatory effects of BMP4 and inhibits proliferation of BMPR1a-positive muscle cells. In vivo, BMP4-positive cells can be found in the proximity of BMPR1a-positive cells in the interstitial spaces between myofibers. Gremlin is expressed by mature myofibers and interstitial cells, which are separate from BMP4-expressing cells. Together, these studies propose that BMP4 and Gremlin, which are highly expressed by human fetal skeletal muscle SP and MP cells, respectively, are regulators of myogenic progenitor proliferation.

Bone Morphogenetic Protein 4↗
PubMed2006

Melanoma cell adhesion molecule is a novel marker for human fetal myogenic cells and affects myoblast fusion.

Myoblast fusion is a highly regulated process that is important during muscle development and myofiber repair and is also likely to play a key role in the incorporation of donor cells in myofibers for cell-based therapy. Although several proteins involved in muscle cell fusion in Drosophila are known, less information is available on the regulation of this process in vertebrates, including humans. To identify proteins that are regulated during fusion of human myoblasts, microarray studies were performed on samples obtained from human fetal skeletal muscle of seven individuals. Primary muscle cells were isolated, expanded, induced to fuse in vitro, and gene expression comparisons were performed between myoblasts and early or late myotubes. Among the regulated genes, melanoma cell adhesion molecule (M-CAM) was found to be significantly downregulated during human fetal muscle cell fusion. M-CAM expression was confirmed on activated myoblasts, both in vitro and in vivo, and on myoendothelial cells (M-CAM(+) CD31(+)), which were positive for the myogenic markers desmin and MyoD. Lastly, in vitro functional studies using M-CAM RNA knockdown demonstrated that inhibition of M-CAM expression enhances myoblast fusion. These studies identify M-CAM as a novel marker for myogenic progenitors in human fetal muscle and confirm that downregulation of this protein promotes myoblast fusion.

Adult↗
PubMed2005

Myogenic potential of muscle side and main population cells after intravenous injection into sub-lethally irradiated mdx mice.

Muscle side population (SP) cells have demonstrated hematopoietic and myogenic activities in vivo upon intravenous (IV) injection into lethally irradiated mdx mice. In contrast, muscle main population (MP) cells were unable to rescue the bone marrow of lethally irradiated mice and, consequently, their in vivo myogenic potential could not be assessed using this method. In the current study, muscle SP or MP cells derived from syngeneic wild-type male mice were delivered to sub-lethally irradiated mdx female mice by single or serial IV injections. Recipient mice were euthanized 12 weeks after transplantation at which time the quadriceps and diaphragm muscles were analyzed for the presence of donor-derived cells. Mice injected with 10(4) muscle SP cells or with 10(6) MP cells appeared to have similar numbers of dystrophin-positive myofibers containing fused donor nuclei. Analysis of the remaining tissue via real-time quantitative PCR indicated that mice injected with muscle SP cells had a higher percentage of donor-derived Y-DNA in the quadriceps than mice injected with MP cells, suggesting that muscle SP cells may be enriched for progenitors able to engraft dystrophic skeletal muscles from the circulation. Although the overall engraftment did not reach therapeutically significant levels, these results indicate that further optimization of cell delivery techniques may lead to improved efficacy of cell-mediated therapy using muscle SP cells.

Animals↗
PubMed2004

Demystifying SP cell purification: viability, yield, and phenotype are defined by isolation parameters.

Side population (SP) cells isolated from bone marrow, skeletal muscle, and skin have been shown to engraft in dystrophic muscle. However, there have been questions on the phenotypical heterogeneity, tissue of origin, and relationships among SP cell populations extracted from different tissues. Studies on bone marrow SP cells have followed a consistent protocol for their isolation and results obtained are concordant. In contrast, protocols for the isolation of muscle SP cells vary greatly, and consequently reports on their phenotype, differentiation potential and origin have been inconsistent. To address this controversy, we demonstrate that isolation parameters, such as tissue dissociation, cell counting, Hoechst concentration, and stringency in the selection of SP cells, have an effect on the yield, viability, and homogeneity of SP cells derived from bone marrow, skeletal muscle, and skin. In this paper, we demonstrate that SP cells isolated from the bone marrow are distinct from SP cells extracted from skeletal muscle and skin tissues. This study offers an explanation for the controversy surrounding muscle SP cells, provides a detailed standardized protocol for their isolation, and highlights basic guidelines for reproducible and reliable isolation of SP cells from any tissue.

Animals↗
PubMed2004

Role of bone marrow cell trafficking in replenishing skeletal muscle SP and MP cell populations.

The multipotent nature of skeletal muscle-derived side population cells is demonstrated by their myogenic and hematopoietic potential in vivo. However, whether muscle side population cells are derived from the bone marrow is unclear. To study the long-term contribution of the hematopoietic system to muscle side population, whole bone marrow cells from Ly5.1 males or from e-GFP transgenic male mice were transplanted into lethally irradiated Ly5.2 females. Long-term cell trafficking of donor bone marrow cells to muscle side population was monitored 17 times in a 34-week study. Fluorescence-activated cell sorter analyses were used to detect Ly5.1 and GFP(+) donor cells, which were confirmed by fluorescence in situ hybridization of the Y-chromosome. Analyses post-transplantation indicated that whereas cells of donor origin could be found in the muscle, donor bone marrow cells had contributed little to the muscle side population. Attempts to increase cell trafficking by induced muscle damage again confirmed that more than 90% of side population cells present in the muscle were derived from the host. These results demonstrate that muscle side population cells are not replenished by the bone marrow and suggest a non-hematopoietic origin for this cell population.

Animals↗
PubMed2004

Stem cell therapy for muscular dystrophy.

Muscular dystrophy is a heterogeneous group of neuromuscular disorders that manifests as progressive muscle weakness, muscle wasting and, in many cases, death. Although there has been enormous progress in the molecular understanding of muscular dystrophy, there is still no cure. There are, however, several different therapeutic options under investigation, including adult-derived stem cell transplantation. Encouraging and pioneering experiments in mouse models for Duchenne's muscular dystrophy (DMD) demonstrated that myoblasts could be transplanted into dystrophic muscle; these myoblasts repaired a small proportion of damaged myofibres. Subsequent work has been devoted to optimisation of this technique. In doing so, a number of adult-derived stem cells have been isolated, characterised and used in promising animal transplantation experiments. Further research is ongoing, and is clearly necessary to make this therapy a viable treatment option for patients with muscular dystrophy.

Animals↗
PubMed2002

Recent advances in and therapeutic potential of muscle-derived stem cells.

Over the past few years, issues related to the commitment and potential of reservoir precursor cells that reside in most tissues have been revisited. Many reports have documented either plasticity or de-differentiation of a number of precursor cells isolated from several tissues, including bone marrow, brain, and skeletal muscle. These findings have challenged the dogma that mononuclear cells derived from adult, post-mitotic tissues can differentiate and contribute only to the tissue from which they originate. Thus, much current research in stem cells is testing the therapeutic potential of these cells to deliver normal genes and their encoded proteins into damaged or injured tissues. This review will focus on muscle-derived precursor cells and their apparently heterogeneous nature and summarize some of the most recent findings and hypotheses on their characterization and practical use.

Animals↗
PubMed2002

Long-term persistence of donor nuclei in a Duchenne muscular dystrophy patient receiving bone marrow transplantation.

Duchenne muscular dystrophy (DMD) is a severe progressive muscle-wasting disorder caused by mutations in the dystrophin gene. Studies have shown that bone marrow cells transplanted into lethally irradiated mdx mice, the mouse model of DMD, can become part of skeletal muscle myofibers. Whether human marrow cells also have this ability is unknown. Here we report the analysis of muscle biopsies from a DMD patient (DMD-BMT1) who received bone marrow transplantation at age 1 year for X-linked severe combined immune deficiency and who was diagnosed with DMD at age 12 years. Analysis of muscle biopsies from DMD-BMT1 revealed the presence of donor nuclei within a small number of muscle myofibers (0.5-0.9%). The majority of the myofibers produce a truncated, in-frame isoform of dystrophin lacking exons 44 and 45 (not wild-type). The presence of bone marrow-derived donor nuclei in the muscle of this patient documents the ability of exogenous human bone marrow cells to fuse into skeletal muscle and persist up to 13 years after transplantation.

Adolescent↗