Biology subjects

Meunier, P.

Publications and source records attributed to Meunier, P..

5 recordsLinked to original sources

bioRxiv2026

EFFICIENT EXPANSION OF NK-92 CELL LINE USING A NOVEL LOW-SHEAR STRESS BIOREACTOR

BackgroundThe use of autologous or allogeneic cell therapies has now entered to the clinical practice in several fields of medicine, especially in oncology and hematology. From this regard, 2D-cell manufacturing is complex and costly and bioreactors have attracted major interest for efficient and cost-effective mass production of cells. Bioreactors have several advantages such as homogeneous repartition of nutrients and gas, control of all culture parameters and increased yield. However, the important shear stress generated by those bioreactors is an important disadvantage as it can affect cell survival or cell quality. This important shear stress is the result of the mixing method using either blades (used in stirred-tanked bioreactors) or gas bubbles (used in airlift bioreactors). Another downside of the use of bioreactors is the difficulty to scale-up. As the volume increases, the shear stress generated by blades radically increases leading to cell death and a decrease of cell quality. DescriptionIn this study, we describe a bioreactor developed using a different mixing method effectively reducing the shear stress and facilitating scale-up. This bladeless method uses an inclination of the bioreactor as well as rotation to mix fluids in a container. Here we described different steps that led to the adaptation of this bioreactor, initially developed for fragile microalgae culture, for mammalian cell culture amplification. The bioreactor was tested to amplify a natural killer (NK) cell line NK92 which is an IL-2 dependent cell line used in clinical trials for cancer therapy. We have tested the influence of 1-The number of cells seeded; 2-The influence of the rotation speed on cell growth and viability; 3-The influence of the bioreactor angle on the above parameters; 4-The duration of the culture. ResultsCells were initially seeded at 2.5.105 / ml in a volume of 380 ml. According to the rotation speed of 15, 30, 45 and 60 rpm, we have observed an increase of cell numbers at day 3 (3-fold), day 5 (7-fold) and day 7 (10-fold) compared to seeding, the best expansion being obtained at day 7 with a rotation speed of 45 rpm. The optimal angle of rotation was found to be 3 degree, with an optimal amplification at day 7 versus day 3 (p < 0.01). The viability was also found to be optimal in the latter condition. ConclusionsThese preliminary results demonstrate that NK92 cells could be amplified using this bioreactor. In the best tested condition, neither cell viability nor cell growth was impacted. These results strongly suggest the potential use of this device in future clinically applicable conditions.

bioengineering↗

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↗

bioRxiv2025

Identification of CaVβ1 isoforms required for neuromuscular junction formation and maintenance

Voltage-gated Ca{superscript 2} channels (VGCCs) are regulated by four CaV{beta} subunits (CaV{beta}1-CaV{beta}4), each showing specific expression patterns in excitable cells. While primarily known for regulating VGCC function, CaV{beta} proteins also have channel-independent roles, including gene expression modulation. Among these, CaV{beta}1 is expressed in skeletal muscle as multiple isoforms. The adult isoform, CaV{beta}1D, localizes at the triad and modulates CaV1 activity during Excitation-Contraction Coupling (ECC). In this study, we investigated the lesser-known embryonic/perinatal CaV{beta}1 isoforms and their roles in neuromuscular junction (NMJ) formation, maturation, and maintenance. We found that CaV{beta}1 isoform expression is developmentally regulated through differential promoter activation. Specifically, CaV{beta}1A is expressed in embryonic muscle and reactivated in denervated adult muscle, alongside the known CaV{beta}1E isoform. Nerve injury in adult muscle triggers a shift in promoter usage, resulting in re-expression of embryonic/perinatal Cacnb1A and Cacnb1E transcripts. Functional analyses using aneural agrin-induced AChR clustering on primary myotubes demonstrated that these isoforms contribute to NMJ formation. Additionally, their expression during early postnatal development is essential for NMJ maturation and long-term maintenance. These findings reveal previously unrecognized roles of CaV{beta}1 isoforms beyond VGCC regulation, highlighting their significance in neuromuscular system development and homeostasis.

physiology↗

bioRxiv2025

Systemic Factors Affect Bone Health in SMA Type II Patients and a Mouse Model of SMA

Spinal muscular atrophy (SMA) is a rare developmental disorder affecting multiple tissues. Among the non-central nervous system tissues implicated in SMA is the skeletal system, including bone and cartilage. Low bone mineral density, increased numbers of fractures of the long bones and vertebra, hip pain, and scoliosis have been reported across the spectrum of SMA patients. While lack of ambulation likely contributes significantly to bone pathology, SMA patients have markedly lower bone density compared to other non-ambulatory patients with debilitating diseases such as Duchenne muscular dystrophy, suggesting that there is a cell-intrinsic contribution of SMN to bone homeostasis and function. Mouse models of SMA have also confirmed the presence of bone and cartilage phenotypes. These alterations frequently persist post-treatment. Recent advancements in therapeutic strategies, approved by both the FDA and the EMA, have represented a leap forward in the management of SMA. However, treatment gaps remain. Post-treatment, patients frequently face continued challenges with scoliosis, bone fractures, and persistent muscle weakness--conditions that underscore the urgent need for more comprehensive therapeutic strategies with combination therapies that can support skeletal health. To date, no molecular map exists of the changes that occur in SMA patient bone and cartilage, impeding the ability of finding targeted therapies. To address this clinical need, we profiled the transcriptome of the vertebral bone and cartilage in a cohort of 11 Type II SMA patients who were undergoing surgery for scoliosis correction and compared them to 7 idiopathic scoliosis and 2 DMD controls. Additionally, we characterized the skeletal health of a mouse model of type I SMA. We find that multisystemic factors including liver and muscle health affect the underlying SMA bone pathology. Specifically, we detect alterations in the balance between osteoclasts and osteoblasts, changes in PPAR{gamma} signaling, mitochondrial oxidative phosphorylation and fatty acid beta-oxidation, and alterations in the muscle-derived factor Irisin that play a role in overall SMA bone pathology.

genomics↗

bioRxiv2024

Early endosome disturbance and endolysosomal pathway dysfunction in Duchenne muscular dystrophy

Duchenne muscular dystrophy (DMD) is a lethal dystrophy characterized by the progressive loss of muscle fibers caused by mutations in DMD gene and absence of the dystrophin protein. While autophagy and lysosome biogenesis defects have been described in DMD muscles, the endosomal pathway has never been studied. Here, we showed that impaired lysosome formation is associated with altered acidification and reduced degradative function of the endolysosomal pathway in muscle cells derived from DMD patients. Our data demonstrated that early endosomes are increased in these cells as well as in muscle biopsies from DMD patients and two animal models of DMD, mdx mice and GRMD dogs. We determined that these abnormalities are due to the lack of dystrophin per se and could be correlated with disease progression and severity. We further identified an abnormal upregulation of the Rab5 GTPase protein, one key actor of early endosomal biogenesis and fusion, in the three DMD models which may underlie the endosomal defects. Finally, we demonstrated that Rab5 knock-down in human DMD muscle cells as well as dystrophin restoration in GRMD dogs, normalize Rab5 expression levels and rescue endosomal abnormalities. This study unveils a defect in a pathway essential for muscle homeostasis and for efficacy of adeno associated virus vectors and antisense oligonucleotides-mediated therapies.

pathology↗