Author Correction: Cell-mediated exon skipping normalizes dystrophin expression and muscle function in a new mouse model of Duchenne Muscular Dystrophy.

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Cell-mediated exon skipping normalizes dystrophin expression and muscle function in a new mouse model of Duchenne Muscular Dystrophy.

Cell therapy for muscular dystrophy has met with limited success, mainly due to the poor engraftment of donor cells, especially in fibrotic muscle at an advanced stage of the disease. We developed a cell-mediated exon skipping that exploits the multinucleated nature of myofibers to achieve cross-correction of resident, dystrophic nuclei by the U7 small nuclear RNA engineered to skip exon 51 of the dystrophin gene. We observed that co-culture of genetically corrected human DMD myogenic cells (but not of WT cells) with their dystrophic counterparts at a ratio of either 1:10 or 1:30 leads to dystrophin production at a level several folds higher than what predicted by simple dilution.

Mesoangioblasts at 20: From the embryonic aorta to the patient bed.

In 2002 we published an article describing a population of vessel-associated progenitors that we termed mesoangioblasts (MABs). During the past decade evidence had accumulated that during muscle development and regeneration things may be more complex than a simple sequence of binary choices (e.g.

Alteration of the late endocytic pathway in Charcot-Marie-Tooth type 2B disease.

The small GTPase RAB7A regulates late stages of the endocytic pathway and plays specific roles in neurons, controlling neurotrophins trafficking and signaling, neurite outgrowth and neuronal migration. Mutations in the RAB7A gene cause the autosomal dominant Charcot-Marie-Tooth type 2B (CMT2B) disease, an axonal peripheral neuropathy. As several neurodegenerative diseases are caused by alterations of endocytosis, we investigated whether CMT2B-causing mutations correlate with changes in this process.

Reversible immortalisation enables genetic correction of human muscle progenitors and engineering of next-generation human artificial chromosomes for Duchenne muscular dystrophy.

Transferring large or multiple genes into primary human stem/progenitor cells is challenged by restrictions in vector capacity, and this hurdle limits the success of gene therapy. A paradigm is Duchenne muscular dystrophy (DMD), an incurable disorder caused by mutations in the largest human gene: dystrophin. The combination of large-capacity vectors, such as human artificial chromosomes (HACs), with stem/progenitor cells may overcome this limitation.

Isolation and Characterization of Vessel-Associated Stem/Progenitor Cells from Skeletal Muscle.

More than 10 years ago, we isolated from mouse embryonic dorsal aorta a population of vessel-associated stem/progenitor cells, originally named mesoangioblasts (MABs ) , capable to differentiate in all mesodermal-derived tissues, including skeletal muscle. Similar though not identical cells have been later isolated and characterized from small vessels of adult mouse and human skeletal muscles. When delivered through the arterial circulation, MABs cross the blood vessel wall and participate in skeletal muscle regeneration , leading to an amelioration of muscular dystrophies in different preclinical animal models.

Intra-arterial transplantation of HLA-matched donor mesoangioblasts in Duchenne muscular dystrophy.

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Intra-arterial transplantation of HLA-matched donor mesoangioblasts in Duchenne muscular dystrophy.

Intra-arterial transplantation of mesoangioblasts proved safe and partially efficacious in preclinical models of muscular dystrophy. We now report the first-in-human, exploratory, non-randomized open-label phase I-IIa clinical trial of intra-arterial HLA-matched donor cell transplantation in 5 Duchenne patients. We administered escalating doses of donor-derived mesoangioblasts in limb arteries under immunosuppressive therapy (tacrolimus).

Mesoangioblast delivery of miniagrin ameliorates murine model of merosin-deficient congenital muscular dystrophy type 1A.

Background: Merosin-deficient congenital muscular dystrophy type-1A (MDC1A) is characterized by progressive muscular dystrophy and dysmyelinating neuropathy caused by mutations of the α2 chain of laminin-211, the predominant laminin isoform of muscles and nerves. MDC1A has no available treatment so far, although preclinical studies showed amelioration of the disease by the overexpression of miniagrin (MAG). MAG reconnects orphan laminin-211 receptors to other laminin isoforms available in the extracellular matrix of MDC1A mice.

PW1/Peg3 expression regulates key properties that determine mesoangioblast stem cell competence.

Mesoangioblasts are vessel-associated progenitor cells that show therapeutic promise for the treatment of muscular dystrophy. Mesoangioblasts have the ability to undergo skeletal muscle differentiation and cross the blood vessel wall regardless of the developmental stage at which they are isolated. Here we show that PW1/Peg3 is expressed at high levels in mesoangioblasts obtained from mouse, dog and human tissues and its level of expression correlates with their myogenic competence.

Inflammation converts human mesoangioblasts into targets of alloreactive immune responses: implications for allogeneic cell therapy of DMD.

Stem cell therapy is a promising approach to regenerate healthy tissues starting from a limited amount of self-renewing cells. Immunological rejection of cell therapy products might represent a major limitation. In this study, we investigated the immunological functional profile of mesoangioblasts, vessel-associated myogenic stem cells, currently tested in a phase 1-2a trial, active in our Institute, for the treatment of Duchenne muscular dystrophy.

Human iPSC-derived mesoangioblasts, like their tissue-derived counterparts, suppress T cell proliferation through IDO- and PGE-2-dependent pathways.

Human mesoangioblasts are currently in a phase I/II clinical trial for the treatment of patients with Duchenne muscular dystrophy. However, limitations associated with the finite life span of these cells combined with the significant numbers of mesoangioblasts required to treat all of the skeletal muscles in these patients restricts their therapeutic potential. Induced pluripotent stem cell (iPSC)-derived mesoangioblasts may provide the solution to this problem.

Mesoangioblasts suppress T cell proliferation through IDO and PGE-2-dependent pathways.

Human mesoangioblasts are vessel-associated stem cells that are currently in phase I/II clinical trials for the treatment of patients with Duchenne muscular dystrophy. To date, little is known about the effect of mesoangioblasts on human immune cells and vice versa. We hypothesized that mesoangioblasts could modulate the function of immune cells in a similar manner to mesenchymal stromal cells.

The increase of pericyte population in human neuromuscular disorders supports their role in muscle regeneration in vivo.

Pericytes are periendothelial cells that have been involved in many different functions including a possible role as mesodermal stem/progenitor cells. In the present study we demonstrate that alkaline phosphatase (AP) expression is specific for human muscular pericytes and can be used as a marker to identify them in skeletal muscle biopsies. We studied the pericyte population in skeletal muscle biopsies from controls, myopathic and neuropathic patients.

Transplantation of genetically corrected human iPSC-derived progenitors in mice with limb-girdle muscular dystrophy.

Mesoangioblasts are stem/progenitor cells derived from a subset of pericytes found in muscle that express alkaline phosphatase. They have been shown to ameliorate the disease phenotypes of different animal models of muscular dystrophy and are now undergoing clinical testing in children affected by Duchenne's muscular dystrophy. Here, we show that patients with a related disease, limb-girdle muscular dystrophy 2D (LGMD2D), which is caused by mutations in the gene encoding α-sarcoglycan, have reduced numbers of this pericyte subset and thus produce too few mesoangioblasts for use in autologous cell therapy.

Necdin enhances muscle reconstitution of dystrophic muscle by vessel-associated progenitors, by promoting cell survival and myogenic differentiation.

Improving stem cell therapy is a major goal for the treatment of muscle diseases, where physiological muscle regeneration is progressively exhausted. Vessel-associated stem cells, such as mesoangioblasts (MABs), appear to be the most promising cell type for the cell therapy for muscular dystrophies and have been shown to significantly contribute to restoration of muscle structure and function in different muscular dystrophy models. Here, we report that melanoma antigen-encoding gene (MAGE) protein necdin enhances muscle differentiation and regeneration by MABs.

Stem cell-mediated transfer of a human artificial chromosome ameliorates muscular dystrophy.

In contrast to conventional gene therapy vectors, human artificial chromosomes (HACs) are episomal vectors that can carry large regions of the genome containing regulatory elements. So far, HACs have not been used as vectors in gene therapy for treating genetic disorders. Here, we report the amelioration of the dystrophic phenotype in the mdx mouse model of Duchenne muscular dystrophy (DMD) using a combination of HAC-mediated gene replacement and transplantation with blood vessel-associated stem cells (mesoangioblasts).

Partial dysferlin reconstitution by adult murine mesoangioblasts is sufficient for full functional recovery in a murine model of dysferlinopathy.

Dysferlin deficiency leads to a peculiar form of muscular dystrophy due to a defect in sarcolemma repair and currently lacks a therapy. We developed a cell therapy protocol with wild-type adult murine mesoangioblasts. These cells differentiate with high efficiency into skeletal muscle in vitro but differ from satellite cells because they do not express Pax7.

Inflammatory and alternatively activated human macrophages attract vessel-associated stem cells, relying on separate HMGB1- and MMP-9-dependent pathways.

Inflammatory macrophages recruited at the site of damaged muscles progressively acquire an alternative activation profile. Inflammatory (M1) and alternatively activated (M2) macrophages exert various and even opposite functions. M1 cells amplify tissue damage, and M2 cells dispose of necrotic fibers and deliver survival signals to myogenic precursors, finally supporting healing.

Skeletal muscle differentiation of embryonic mesoangioblasts requires pax3 activity.

Mesoangioblasts have been characterized as a population of vessel-associated stem cells able to differentiate into several mesodermal cell types, including skeletal muscle. Here, we report that the paired box transcription factor Pax3 plays a crucial role in directing mouse mesoangioblasts toward skeletal myogenesis in vitro and in vivo. Mesoangioblasts isolated from the aorta of Pax3 null embryos are severely impaired in skeletal muscle differentiation, whereas most other differentiation programs are not affected by the absence of Pax3.

Isolation and characterization of mesoangioblasts from mouse, dog, and human tissues.

Mesoangioblasts are recently identified stem/progenitor cells, associated with small vessels of the mesoderm in mammals. Originally described in the mouse embryonic dorsal aorta, similar though not identical cells have been later identified and characterized from postnatal small vessels of skeletal muscle and heart (not described in this unit). They have in common the anatomical location, the expression of endothelial and/or pericyte markers, the ability to proliferate in culture, and the ability to undergo differentiation into various types of mesoderm cells upon proper culture conditions.

Autologous transplantation of muscle-derived CD133+ stem cells in Duchenne muscle patients.

Duchenne muscular dystrophy (DMD) is a lethal X-linked recessive muscle disease due to defect on the gene encoding dystrophin. The lack of a functional dystrophin in muscles results in the fragility of the muscle fiber membrane with progressive muscle weakness and premature death. There is no cure for DMD and current treatment options focus primarily on respiratory assistance, comfort care, and delaying the loss of ambulation.

Pericytes of human skeletal muscle are myogenic precursors distinct from satellite cells.

Cells derived from blood vessels of human skeletal muscle can regenerate skeletal muscle, similarly to embryonic mesoangioblasts. However, adult cells do not express endothelial markers, but instead express markers of pericytes, such as NG2 proteoglycan and alkaline phosphatase (ALP), and can be prospectively isolated from freshly dissociated ALP(+) cells. Unlike canonical myogenic precursors (satellite cells), pericyte-derived cells express myogenic markers only in differentiated myotubes, which they form spontaneously with high efficiency.

Mesoangioblast stem cells ameliorate muscle function in dystrophic dogs.

Duchenne muscular dystrophy remains an untreatable genetic disease that severely limits motility and life expectancy in affected children. The only animal model specifically reproducing the alterations in the dystrophin gene and the full spectrum of human pathology is the golden retriever dog model. Affected animals present a single mutation in intron 6, resulting in complete absence of the dystrophin protein, and early and severe muscle degeneration with nearly complete loss of motility and walking ability.