Biology subjects

Halgand, B.

Publications and source records attributed to Halgand, B..

3 recordsLinked to original sources

bioRxiv2025

Transcriptomic and functional comparison of cells isolated from healthy and degenerated ovine intervertebral discs

Structured abstractO_ST_ABSBackgroundC_ST_ABSIntervertebral disc degeneration (IVDD) is a leading cause of chronic low back pain and disability. Understanding the cellular and molecular mechanisms underlying disc degeneration is crucial for developing effective therapies. Sheep have emerged as a promising large-animal model for IVDD research due to their similarities with humans. They exhibit resembling spine anatomy and biomechanics, and they develop spontaneous age-associated degeneration of the disc. However, the specific cellular alterations occurring in annulus fibrosus (AF) and nucleus pulposus (NP) ovine cells during degeneration remain poorly characterized. In vitro, the benefits of using cells from aged sheep over young ones to mimic degenerative processes remain to be tested. MethodsAF and NP cells from young and aged sheep were analysed using bulk RNA sequencing, with a focus on two hallmarks of IVDD: cellular senescence and metabolic alterations. Functional assays completed this focus by assessing cells response under basal conditions and after pro-degenerative stimuli (IL-1{beta}, senescence induction). In addition, bulk transcriptomic data were deconvoluted using a reference single-cell RNA-seq dataset from healthy and degenerated human discs, and gene co-expression modules were compared across species. ResultsMRI and histological analyses revealed homogeneous mild degeneration across all lumbar discs in aged sheep, while lamb discs were uniformly healthy. Cells transcriptomic profiling identified robust age- and tissue-specific signatures, with aged NP and AF cells showing upregulation of inflammatory mediators, ECM-remodelling enzymes, and senescence-associated pathways. Cross-species analysis revealed shared transcriptional modules between aged sheep cells and human degenerated disc cells, supporting the translational relevance of the ovine model. Remarkably, young and aged cells shared a similar functional behaviour when exposed to stress-related stimuli. ConclusionsThis work confirms the compatibility of sheep cells with in vitro testing and their relevance to model human IVDD. Cross-validation with human single-cell data further highlights common pathogenic pathways, reinforcing the translational potential of the model. However, no added benefits were found in using older animals compared to younger ones as cell sources in functional assays. HighlightsO_LITranscriptomic profiling of AF and NP cells from young and aged sheep C_LIO_LIAged cells show inflammatory, ECM-remodelling and senescence signatures C_LIO_LIDeconvolution with human scRNA-seq links aged ovine and degenerated discs C_LIO_LISheep cells retain in vitro responsiveness to pro-degenerative stimuli C_LIO_LISupports the ovine model as a translational tool for IVDD research C_LI

cell biology↗

bioRxiv2025

BIOFABRICATION OF AN OVINE INTERVERTEBRAL DISC MODEL BY COMBINING A POLYCAPROLACTONE FRAME WITH A BIOPRINTED ALGINATE HYDROGEL

The intervertebral disc (IVD) primarily comprises an outer ring of collagen fibers (annulus fibrosus, AF), which encases a soft, gelatinous core (nucleus pulposus, NP). Existing in vitro models have failed to integrate these two tissues effectively or accurately replicate their intricate organization. By combining two biofabrication techniques, we developed a novel 3D in vitro model that closely mimics the organization of an ovine IVD. Our approach employs a polycaprolactone (PCL) frame produced via melt electrowriting to recreate the multilamellar architecture of the annulus fibrosus. Ovine primary cells, encapsulated in a photocrosslinkable alginate hydrogel, were precisely extruded within the multilamellar structure, thereby mimicking the native shape and size of an ovine disc. The bioink containing the NP cells was deposited at the center of the construct, while the bioink with the AF cells was strategically layered in between the lamellae of the PCL frame. Photocrosslinking was optimized to match the native stiffness of the disc. The constructs were maintained in culture for 28 days, during which we thoroughly assessed reproducibility, stability, and cell viability and phenotype. The results unequivocally demonstrated that the PCL frame effectively guided the alignment and proliferation of AF cells, while the alginate hydrogel preserved NP cell phenotype. This model successfully replicates the organization of the IVD, providing a promising platform for advancing our understanding of disc biology and driving the development of novel therapeutic strategies.

bioengineering↗

bioRxiv2023

Microencapsulation of mesenchymal stromal cells in covalent alginate hydrogels for cell therapy

Osteoarthritis (OA) is the most common inflammatory joint disease and currently lacks an effective curative treatment. Intra-articular injection of mesenchymal stromal cells (MSCs) has gained attention as a relevant therapeutic approach for OA treatment due to the MSCs ability to secrete anti-inflammatory and immunomodulatory factors. Given their limited viability post- intraarticular injection and the potential leakage of cells out of the injection site, encapsulating MSCs in hydrogels is considered a promising strategy to protect them and provide a suitable 3D microenvironment to support their biological activities. Calcium-cross-linked alginate hydrogels are commonly used for MSC encapsulation, but their long-term in vivo stability remains uncertain. On the other hand, alginate cross-linking by the strain-promoted azide- alkyne cycloaddition (SPAAC) reaction would create a network unaffected by an ionic environment. Hence, this study aimed to develop an alginate-based hydrogel cross-linked via stable and cytocompatible covalent bonds for cell encapsulation. We established for the first time the formation of covalent alginate hydrogels between two SPAAC precursors, namely alginate-BCN and alginate-N3. These hydrogels exhibited in vitro stability and enabled the diffusion of molecules of interest. We then generated alginate-based SPAAC microgels of 170 m in mean diameter, suitable for intra-articular injection. We next encapsulated human adipose MSCs (hASCs) in these alginate-based SPAAC microgels and confirmed their cytocompatibility, with over 90 % of cells remaining viable after 14 days in culture. Finally, the microencapsulated hASCs maintained their biological properties and were able to secrete anti-inflammatory factors (IDO, PGE2, and HGF) when exposed to pro-inflammatory cytokines (TNF- and IFN-{gamma}). In the end, human-activated lymphocytes were cultured in contact with microencapsulated hASCs, and CD3+ T cell proliferation was quantified by flow cytometry. We demonstrated that the encapsulation process did not impair the hASC immunomodulatory activity. Overall, our findings show the potential of alginate-based SPAAC hydrogels for microencapsulating hASCs for cell therapy.

bioengineering↗