Project description:In humans, chronic glucocorticoid use is associated with side effects like muscle wasting, obesity, and metabolic syndrome. Intermittent steroid dosing has been proposed in Duchenne Muscular Dystrophy patients to mitigate the side effects seen with daily steroid intake. We evaluated biomarkers from Duchenne Muscular Dystrophy patients, finding that, compared with chronic daily steroid use, weekend steroid use was associated with reduced serum insulin, free fatty acids, and branched chain amino acids, as well as reduction in fat mass despite having similar BMIs. We reasoned that intermittent prednisone administration in dystrophic mice would alter muscle epigenomic signatures, and we identified the coordinated action of the glucocorticoid receptor, KLF15 and MEF2C as mediators of a gene expression program driving metabolic reprogramming and enhanced nutrient utilization. Muscle lacking Klf15 failed to respond to intermittent steroids. Furthermore, coadministration of the histone acetyltransferase inhibitor anacardic acid with steroids in mdx mice eliminated steroid-specific epigenetic marks and abrogated the steroid response. Together, these findings indicate that intermittent, repeated exposure to glucocorticoids promotes performance in dystrophic muscle through an epigenetic program that enhances nutrient utilization.
Project description:The muscular dystrophies (MDs) are genetic muscle diseases that result in progressive muscle degeneration followed by the fibrotic replacement of affected muscles as regenerative processes fail. Therapeutics that specifically address the fibrosis and failed regeneration associated with MDs represent a major unmet clinical need for MD patients, particularly those with advanced-stage disease progression. The current study investigated targeting NAD(P)H oxidase 4 (NOX4) as a potential strategy to reduce fibrosis and promote regeneration in disease-burdened muscle that models Duchenne muscular dystrophy (DMD). NOX4 was elevated in the muscles of dystrophic mice and DMD patients, localizing primarily to interstitial cells located between muscle fibers. Genetic and pharmacological targeting of NOX4 significantly reduced fibrosis in dystrophic respiratory and limb muscles. Mechanistically, NOX4 targeting decreased the number of fibrosis-depositing cells (myofibroblasts) and restored the number of muscle-specific stem cells (satellite cells) localized to their physiological niche, thereby rejuvenating muscle regeneration. Furthermore, acute inhibition of NOX4 was sufficient to induce apoptotic clearing of myofibroblasts within dystrophic muscle. These data indicate that targeting NOX4 is an effective strategy to promote the beneficial remodeling of disease-burdened muscle representative of DMD and, potentially, other MDs and muscle pathologies.
Project description:ObjectivesPreferential damage to fast, glycolytic myofibers is common in many muscle-wasting diseases, including Duchenne muscular dystrophy (DMD). Promoting an oxidative phenotype could protect muscles from damage and ameliorate the dystrophic pathology with therapeutic relevance, but developing efficacious strategies requires understanding currently unknown biological roles for dystrophin and utrophin in dystrophic muscle adaptation and plasticity.MethodsCombining whole transcriptome RNA sequencing and mitochondrial proteomics with assessments of metabolic and contractile function, we investigated the roles of dystrophin and utrophin in fast-to-slow muscle remodeling with low-frequency electrical stimulation (LFS, 10 Hz, 12 h/d, 7 d/wk, 28 d) in mdx (dystrophin null) and dko (dystrophin/utrophin null) mice, two established preclinical models of DMD.ResultsNovel biological roles in adaptation were demonstrated by impaired transcriptional activation of estrogen-related receptor alpha-responsive genes supporting oxidative phosphorylation in dystrophic muscles. Further, utrophin expression in dystrophic muscles was required for LFS-induced remodeling of mitochondrial respiratory chain complexes, enhanced fiber respiration, and conferred protection from eccentric contraction-mediated damage.ConclusionsThese findings reveal novel roles for dystrophin and utrophin during LFS-induced metabolic remodeling of dystrophic muscle and highlight the therapeutic potential of LFS to ameliorate the dystrophic pathology and protect from contraction-induced injury with important implications for DMD and related muscle disorders.
Project description:Exosomes are promising therapeutics for tissue repair and regeneration to induce and guide appropriate immune responses in dystrophic pathologies. However, manipulating exosomes to control their biodistribution and targeting them in vivo to achieve adequate therapeutic benefits still poses a major challenge. Here we overcome this limitation by developing an externally controlled delivery system for primed annexin A1 myo-exosomes (Exomyo). Effective nanocarriers are realized by immobilizing the Exomyo onto ferromagnetic nanotubes to achieve controlled delivery and localization of Exomyo to skeletal muscles by systemic injection using an external magnetic field. Quantitative muscle-level analyses revealed that macrophages dominate the uptake of Exomyo from these ferromagnetic nanotubes in vivo to synergistically promote beneficial muscle responses in a murine animal model of Duchenne muscular dystrophy. Our findings provide insights into the development of exosome-based therapies for muscle diseases and, in general, highlight the formulation of effective functional nanocarriers aimed at optimizing exosome biodistribution.
Project description:Evidence from numerous studies has revealed the synchronous progression of aging in bone and muscle; however, little is known about the underlying mechanisms. To this end, human muscles and bones are harvested and the aging-associated transcriptional dynamics of two tissues in parallel using single-cell RNA sequencing are surveyed. A subset of lipid-associated macrophages (triggering receptor expressed on myeloid cells 2, TREM2+ Macs) is identified in both aged muscle and bone. Genes responsible for muscle dystrophy and bone loss, such as secreted phosphoprotein 1 (SPP1), are also highly expressed in TREM2+ Macs, suggesting its conserved role in aging-related features. A common transition toward pro-inflammatory phenotypes in aged CD4+ T cells across tissues is also observed, activated by the nuclear factor kappa B subunit 1 (NFKB1). CD4+ T cells in aged muscle experience Th1-like differentiation, whereas, in bone, a skewing toward Th17 cells is observed. Furthermore, these results highlight that degenerated myocytes produce BAG6-containing exosomes that can communicate with Th17 cells in the bone through its receptor natural cytotoxicity triggering receptor 3 (NCR3). This communication upregulates CD6 expression in Th17 cells, which then interact with TREM2+ Macs through CD6-ALCAM signaling, ultimately stimulating the transcription of SPP1 in TREM2+ Macs. The negative correlation between serum exosomal BCL2-associated athanogene 6 (BAG6) levels and bone mineral density further supports its role in mediating muscle and bone synchronization with aging.
Project description:Duchenne muscular dystrophy (DMD), the most prevalent lethal genetic disorder in children, is caused by mutations in the 2.2-MB dystrophin gene. Absence of dystrophin and the dystrophin-glycoprotein complex (DGC) from the sarcolemma leads to severe muscle wasting and eventual respiratory and/or cardiac failure. There is presently no effective therapy for DMD. Several lines of evidence have suggested that methods to increase expression of utrophin, a dystrophin paralog, show promise as a treatment for DMD. Adeno-associated viral (AAV) vectors are a promising vehicle for gene transfer to muscle, but microutrophin transgenes small enough to be carried by AAV have not been tested for function. In this study, we intravenously administered recombinant AAV (rAAV2/6) harboring a murine codon-optimized microutrophin (DeltaR4-R21/DeltaCT) transgene to adult dystrophin(-/-)/utrophin(-/-) (mdx:utrn(-/-)) double-knockout mice. Five-month-old mice demonstrated localization of microutrophin to the sarcolemma in all the muscles tested. These muscles displayed restoration of the DGC, increased myofiber size, and a considerable improvement in physiological performance when compared with untreated mdx:utrn(-/-) mice. Overall, microutrophin delivery alleviated most of the pathophysiological abnormalities associated with muscular dystrophy in the mdx:utrn(-/-) mouse model. This approach may hold promise as a treatment option for DMD because it avoids the potential immune responses that are associated with the delivery of exogenous dystrophin.
Project description:T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy derived from early T cell progenitors. Since relapsed T-ALL is associated with a poor prognosis improving initial treatment of patients is essential to avoid resistant selection of T-ALL. During initiation, development, metastasis and even in response to chemotherapy, tumor cells face strong metabolic challenges. In this study, we identify mitochondrial UnCoupling Protein 2 (UCP2) as a tricarboxylic acid (TCA) cycle metabolite transporter controlling glutamine metabolism associated with T-ALL cell proliferation. In T-ALL cell lines, we show that UCP2 expression is controlled by glutamine metabolism and is essential for their proliferation. Our data show that T-ALL cell lines differ in their substrate dependency and their energetic metabolism (glycolysis and oxidative). Thus, while UCP2 silencing decreases cell proliferation in all leukemia cells, it also alters mitochondrial respiration of T-ALL cells relying on glutamine-dependent oxidative metabolism by rewiring their cellular metabolism to glycolysis. In this context, the function of UCP2 in the metabolite export of malate enables appropriate TCA cycle to provide building blocks such as lipids for cell growth and mitochondrial respiration. Therefore, interfering with UCP2 function can be considered as an interesting strategy to decrease metabolic efficiency and proliferation rate of leukemia cells.
Project description:Mitochondria are the powerhouse of eukaryotic cells, which regulate cell metabolism and differentiation. Recently, mitochondrial transfer between cells has been shown to direct recipient cell fate. However, it is unclear whether mitochondria can translocate to stem cells and whether this transfer alters stem cell fate. Here, mesenchymal stem cell (MSC) regulation is examined by macrophages in the bone marrow environment. It is found that macrophages promote osteogenic differentiation of MSCs by delivering mitochondria to MSCs. However, under osteoporotic conditions, macrophages with altered phenotypes, and metabolic statuses release oxidatively damaged mitochondria. Increased mitochondrial transfer of M1-like macrophages to MSCs triggers a reactive oxygen species burst, which leads to metabolic remodeling. It is showed that abnormal metabolism in MSCs is caused by the abnormal succinate accumulation, which is a key factor in abnormal osteogenic differentiation. These results reveal that mitochondrial transfer from macrophages to MSCs allows metabolic crosstalk to regulate bone homeostasis. This mechanism identifies a potential target for the treatment of osteoporosis.
Project description:Muscle wasting is a major pathological feature observed in Duchenne muscular dystrophy (DMD) and is the result of the concerted effects of inflammation, oxidative stress and cell senescence. The inducible form of proteasome, or immunoproteasome (IP), is involved in all the above mentioned processes, regulating antigen presentation, cytokine production and immune cell response. IP inhibition has been previously shown to dampen the altered molecular, histological and functional features of 3-month-old mdx mice, the animal model for DMD. In this study, we described the role of ONX-0914, a selective inhibitor of the PSMB8 subunit of immunoproteasome, in ameliorating the pathological traits that could promote muscle wasting progression in older, 9-month-old mdx mice. ONX-0914 reduces the number of macrophages and effector memory T cells in muscle and spleen, while increasing the number of regulatory T cells. It modulates inflammatory markers both in skeletal and cardiac muscle, possibly counteracting heart remodeling and hypertrophy. Moreover, it buffers oxidative stress by improving mitochondrial efficiency. These changes ultimately lead to a marked decrease of fibrosis and, potentially, to more controlled myofiber degeneration/regeneration cycles. Therefore, ONX-0914 is a promising molecule that may slow down muscle mass loss, with relatively low side effects, in dystrophic patients with moderate to advanced disease.
Project description:In skeletal muscle, the cytolinker plectin is prominently expressed at Z-disks and the sarcolemma. Alternative splicing of plectin transcripts gives rise to more than eight protein isoforms differing only in small N-terminal sequences (5-180 residues), four of which (plectins 1, 1b, 1d, and 1f) are found at substantial levels in muscle tissue. Using plectin isoform-specific antibodies and isoform expression constructs, we show the differential regulation of plectin isoforms during myotube differentiation and their localization to different compartments of muscle fibers, identifying plectins 1 and 1f as sarcolemma-associated isoforms, whereas plectin 1d localizes exclusively to Z-disks. Coimmunoprecipitation and in vitro binding assays using recombinant protein fragments revealed the direct binding of plectin to dystrophin (utrophin) and beta-dystroglycan, the key components of the dystrophin-glycoprotein complex. We propose a model in which plectin acts as a universal mediator of desmin intermediate filament anchorage at the sarcolemma and Z-disks. It also explains the plectin phenotype observed in dystrophic skeletal muscle of mdx mice and Duchenne muscular dystrophy patients.