Biology subjects

Giordani, L.

Publications and source records attributed to Giordani, L..

10 recordsLinked to original sources

bioRxiv2026

Axonopathy in Duchenne Muscular Dystrophy limits microdystrophin gene therapy efficacy

Duchenne muscular dystrophy (DMD) is classically defined as a primary myopathy, and current AAV-mediated microdystrophin gene therapies are shown to successfully preserve muscle integrity. However, their efficacy in recovering functional outcomes remains to improve. We hypothesized that this limitation stems from an unaccounted vulnerability within the peripheral nerve. Here, we demonstrate that the mdx mouse model exhibits a peripheral axonopathy independently of muscle necrosis. Using single-nucleus RNA sequencing and structural analyses, we have identified an active denervation program and a profound failure of neural repair pathways. Importantly, we revealed that the full-length dystrophin isoform Dp427c is expressed in the healthy peripheral nerve, intimately following the cytoskeletal organization and accumulating at regions of high biomechanical stress, including Schmidt-Lanterman incisures and Nodes of Ranvier. In its absence, nerves of mdx mice loss an essential scaffolding support, leading to localized structural collapse. Furthermore, we showed that muscle-restricted microdystrophin gene therapy rescues sarcolemmal integrity but failed to restore nerve-muscle connectivity or resolved neurotransmission defects. These findings fundamentally redefine DMD as an integrated motor unit pathology, thereby underscoring the absolute necessity of implementing combined therapeutic strategies that target both the muscle and the peripheral nervous system.

physiology↗

bioRxiv2025

Spatial and Multiomic profiling of muscle regeneration dynamics in Duchenne Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is a pediatric degenerative myopathy caused by the absence of functional dystrophin. As a result, DMD muscles exhibit compromised myofiber integrity and increased susceptibility to mechanical damage. In early disease stages, muscles undergo repeated cycles of degeneration and regeneration; over time, however, this regenerative capacity declines, leading to the gradual replacement of muscle tissue with fat and fibrosis. While several signaling pathways have been identified as deregulated in dystrophic muscle, the cellular and molecular mechanisms underlying this regenerative exhaustion remain to be fully elucidated. To address this, we constructed a comprehensive cellular atlas of human dystrophic muscle using high-resolution spatial transcriptomics (Visium HD), capturing the cellular crosstalk within regenerative regions. Cell-to-cell communication analysis revealed activation of Notch signaling mediated by NOTCH3 in activated satellite cells. Immunostaining confirmed elevated NOTCH3 expression in both DMD patient samples and in the mdx mouse model at late disease stages. Silencing of NOTCH3 in primary myoblasts improved myogenic differentiation, pinpointing NOTCH3-mediated signaling as a contributor to regeneration impairment. To further dissect the dynamics of regeneration and infer the gene regulatory networks governing myogenic differentiation, we integrated paired snRNA-seq and snATAC-seq data from young, adult, and aged mdx mice. This analysis identified GLIS3 upregulation as an additional barrier to effective myogenesis. GLIS3 displayed an overall increase in dystrophic muscles, while silencing experiments enhanced differentiation in myoblasts. Together, our work reveals intrinsic defects in the dystrophic stem cell compartment that emerge during disease progression and hinder the execution of the myogenic program. These findings suggest NOTCH3 and GLIS3 as potential therapeutic targets to enhance regeneration and maintain muscle integrity in DMD. This study provides a high-resolution map of the dystrophic regenerative landscape and offers a valuable resource for future translational research.

cell biology↗

bioRxiv2025

GDF5 modulation of MuSC pool as a potential therapeutic benefit for DMD

Duchenne muscular dystrophy (DMD) is a fatal disease caused by dystrophin deficiency, leading to degeneration of the entire musculature. To improve muscle pathophysiology and gene therapy for DMD, we investigated the potential of growth differentiation factor 5 (GDF5) in the DMD mdx mouse model. We showed that the overexpression of GDF5 in the muscle improved its histology, reduced inflammation, modulated regeneration and induced the appearance of de novo fibers. We demonstrated that muscle satellite cells (MuSCs) are targeted by GDF5 which enhanced their proliferation and slowed down their myogenic commitment and finally their fusion. When combined with AAV-mediated microdystrophin gene therapy, the leading therapeutic strategy, GDF5 further increased the number of microdystrophin-positive fibers compared to gene therapy alone. These findings highlight GDF5 as a promising modulator of DMD pathology and provide the first evidence of a synergistic effect of the combination of GDF5-based intervention and AAV-microdystrophin treatment.

cell biology↗

bioRxiv2024

CD90 identifies distinct fractions of muscle stem cells with different modalities of activation and quiescence maintenance

Stem cell transition from quiescence to activation is crucial to guarantee productive tissue regeneration. Here we show that CD90 diversifies quiescent muscle stem cells (MuSCs) in murine and human muscle into two subpopulations differing in the kinetics of activation, CD90+ve MuSCs exhibiting a faster exit quiescence and predominating the initial phases of regeneration compared to CD90-ve MuSCs. In the absence of injury, the CD90+ve fraction is primed toward activation through an active CD90-AMPK axis but is maintained in quiescence through signals from the extracellular matrix. Our studies show that Collagen VI, which is preferentially expressed by CD90+ve MuSCs, binds to the Calcitonin receptor and plays a role in this context. Moreover, while the number of CD90+ve and CD90-ve subpopulations is similar in healthy muscles, the CD90-ve fraction predominates in the muscles of murine models of Duchenne and Ullrich congenital muscular dystrophies. These findings provide novel insights into the mechanistic determinants of MuSCs functional heterogeneity and have implications for understanding the stimulation of repair in dystrophic muscle.

cell biology↗

bioRxiv2023

Setdb1 safeguards genome integrity in muscle stem cells to allow for regenerative myogenesis and inflammation

Modulations in chromatin structure orchestrate gene expression and direct stem cell fate. More specifically, the histone 3 lysine 9 Methyltransferase Setdb1 controls transcriptional repression to regulate pluripotency and self-renewal. While Setdb1 functions have been extensively studied in embryonic stem cells and in cancer cells, less is known on its role in adult stem cells in vivo. Here, we show that Setdb1 expression by adult muscle stem cells (MuSCs) is required for muscle tissue regeneration following acute injury. We find that SETDB1 represses the expression of the endogenous retroviruses (ERVs) family of transposable elements in MuSCs. ERV de-expression in Setdb1-null MuSCs prevents their amplification following exit from quiescence and promotes cell death. Multi-omics profiling further shows that the absence of SETDB1 in MuSCs leads to the activation of the DNA-sensing cGAS-STING pathway, entailing increased cytokine expression. In vivo, conditional disruption of Setdb1 in MuSCs provokes aberrant infiltration of inflammatory cells including the appearance of a pathological macrophage lineage. The ensuing histiocytosis is accompanied by necrosis of the newly formed muscle fibers which, in addition with the progressive loss of MuSCs, completely abolish skeletal muscle tissue repair. In contrast, disruption of Setdb1 gene in another muscle-resident cell type, the fibro-adipogenic progenitors (FAPs), does not impact regenerative inflammation. In conclusion, the control of genome stability by SETDB1 in an adult somatic stem cell is necessary for both its regenerative potential and an adequate inflammation regulating tissue repair.

cell biology↗

bioRxiv2022

Spatially resolved transcriptomics reveals innervation-responsive functional clusters in skeletal muscle

Striated muscle is a highly organized structure composed by well-defined anatomical domains with integrated but distinct assignments. So far, the lack of a direct correlation between tissue architecture and gene expression has limited our understanding of how each unit responds to physio-pathologic contexts. Here, we show how the combined use of spatially resolved transcriptomics and immunofluorescence can bridge this gap by enabling the unbiased identification of such domains and the characterization of their response to external perturbations. Using a spatiotemporal analysis, we followed the changes in the transcriptomics profile of specific domains in muscle in a model of denervation. Furthermore, our approach allowed us to identify the spatial distribution and nerve dependence of atrophic signalling pathway and polyamine metabolism to glycolytic fibers. Indeed, we demonstrate a pronounced alteration of polyamine homeostasis upon denervation. Our dataset will serve as a resource for future studies of the mechanisms underlying skeletal muscle homeostasis and innervation. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=188 HEIGHT=200 SRC="FIGDIR/small/486563v1_ufig1.gif" ALT="Figure 1"> View larger version (59K): org.highwire.dtl.DTLVardef@6ee767org.highwire.dtl.DTLVardef@11a4eb4org.highwire.dtl.DTLVardef@25228eorg.highwire.dtl.DTLVardef@1ffa60a_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2020

GREM1 is epigenetically reprogrammed in muscle cells after exercise training and controls myogenesis and metabolism

Exercise training improves skeletal muscle function, notably through tissue regeneration by muscle stem cells. Here, we hypothesized that exercise training reprograms the epigenome of muscle cell, which could account for better muscle function. Genome-wide DNA methylation of myotube cultures established from middle-aged obese men before and after endurance exercise training identified a differentially methylated region (DMR) located downstream of Gremlin 1 (GREM1), which was associated with increased GREM1 expression. GREM1 expression was lower in muscle satellite cells from obese, compared to lean mice, and exercise training restored GREM1 levels to those of control animals. We show that GREM1 regulates muscle differentiation through the negative control of satellite cell self-renewal, and that GREM1 controls muscle lineage commitment and lipid oxidation through the AMPK pathway. Our study identifies novel functions of GREM1 and reveals an epigenetic mechanism by which exercise training reprograms muscle stem cells to improve skeletal muscle function.

cell biology↗

bioRxiv2020

Myofiber stretch induces tensile and shear deformation of muscle stem cells in their native niche

BackgroundMuscle stem cells (MuSCs) are requisite for skeletal muscle regeneration and homeostasis. Proper functioning of MuSCs, including activation, proliferation, and fate decision, is determined by an orchestrated series of events and communication between MuSCs and their niche consisting of the host myofiber and neighbouring cells. A multitude of biochemical stimuli are known to regulate fate and function of MuSCs. However, in addition to biochemical factors, it is conceivable that MuSCs residing between basal lamina and sarcolemma of myofibers are subjected to mechanical forces during muscle stretch-shortening cycles due to myofascial connections between MuSCs and myofibers. MuSCs have been shown to respond to mechanical forces in vitro but it remains to be proven whether physical forces are also exerted on MuSCs in their native niche and whether they contribute to the functioning and fate of MuSCs. MethodsMuSCs deformation in their native niche resulting from mechanical loading of ex vivo myofiber bundles were visualized utilizing mT/mG double-fluorescent Cre-reporter mouse and multiphoton microscopy. MuSCs were subjected to 1 hour pulsating fluid shear stress with a peak shear stress rate of 8.8 Pa/s. After treatment, nitric oxide and mRNA expression levels of genes involved in regulation of MuSC proliferation and differentiation were determined. ResultsEx vivo stretching of extensor digitorum longus and soleus myofiber bundles caused compression as well as tensile and shear deformation of MuSCs in their niche. MuSCs responded to pulsating fluid shear stress in vitro with increased nitric oxide production and an upward trend in iNOS mRNA levels, while nNOS expression was unaltered. Pulsating fluid shear stress enhanced gene expression of c-Fos, Cdk4, and IL-6, while expression of Wnt1, MyoD, Myog, Wnt5a, COX2, Rspo1, Vangl2, Wnt10b, and MGF remained unchanged. ConclusionsWe conclude that MuSCs in their native niche are subjected to force-induced deformations due to myofiber stretch-shortening. Moreover, MuSCs are mechanoresponsive as evident by pulsating fluid shear stress-mediated expression of factors by MuSCs known to promote proliferation.

cell biology↗

bioRxiv2019

TGFβ signaling curbs cell fusion and muscle regeneration

Fusion of muscle progenitor cells is necessary for skeletal muscle development and repair. Cell fusion is a multistep process involving cell migration, adhesion, membrane remodeling and actin-nucleation pathways to generate multinucleated myotubes. While the cellular and molecular mechanisms promoting muscle cell fusion have been intensely investigated in recent years, molecular brakes restraining cell-cell fusion events to control syncytia formation have remained elusive. Here, we show that transforming growth factor beta (TGF{beta}) signaling is active in adult muscle cells throughout the fusion process and reduce muscle cell fusion independently of the differentiation step. In contrast, inhibition of TGF{beta} signaling enhances cell fusion and promotes branching between myotubes. Pharmacological modulation of the pathway in vivo perturbs muscle regeneration after injury. Exogenous addition of TGF{beta} protein results in a loss of muscle function while inhibition of the TGF{beta} pathway induces the formation of giant myofibres. Transcriptome analyses and functional assays revealed that TGF{beta} acts on actin dynamics and reduce cell spreading through modulation of actin-based protrusions. Together our results reveal a signaling pathway that limits mammalian myoblast fusion and add a new level of understanding to the molecular regulation of myogenesis.

cell biology↗

bioRxiv2018

High-dimensional single-cell cartography reveals novel skeletal muscle resident cell populations

Adult tissue repair and regeneration require the activation of resident stem/progenitor cells that can self-renew and generate differentiated progeny. The regenerative capacity of skeletal muscle relies on muscle satellite cells (MuSCs) and their interplay with different cell types within the niche. Yet, our understanding of the cells that compose the skeletal muscle tissue is limited and molecular definitions of the principal cell types are lacking. Using a combined approach of single-cell RNA-sequencing and mass cytometry, we precisely mapped the different cell types in adult skeletal muscle tissue and highlighted previously overlooked populations. We identified known functional populations, characterized their gene signatures, and determined key markers. Among the ten main cell populations present in skeletal muscle, we found an unexpected complexity in the interstitial compartment and identified two new cell populations. One express the transcription factor Scleraxis and generate tenocyte-like cells. The second express smooth muscle and mesenchymal cell markers (SMMCs). While distinct from MuSCs, SMMCs are endowed with myogenic potential and promote MuSC engraftment following transplantation. Our high-dimensional single-cell atlas uncovers principles of an adult tissue composition and can be exploited to reveal unknown cellular sub-fractions that contribute to tissue regeneration.

cell biology↗