Biology subjects

Lafuente-Gomez, N.

Publications and source records attributed to Lafuente-Gomez, N..

4 recordsLinked to original sources

bioRxiv2026

Cholinergic modulation engages a CHRM1-IL-17 neuroimmune axis to promote diabetic wound repair

Chronic non-healing wounds are a major complication of diabetes, where persistent inflammation fails to transition into effective tissue repair (1-3). Here we identify cholinergic modulation as a mechanism that promotes productive inflammation and diabetic wound healing. Using a healing-associated fibroblast signature derived from single-cell profiling of human diabetic foot ulcers (3), we identified the acetylcholinesterase (AChE) inhibitor metrifonate as a candidate inducer of reparative wound programs. Topical delivery accelerated healing in diabetic mice and increased acetylcholine availability within the wound. Complementary receptor pharmacology identified muscarinic acetylcholine receptor 1 (CHRM1) as the principal receptor mediating the reparative response. Transcriptomic and cellular analyses revealed early induction of IL-17-associated inflammatory programs and enrichment of CHRM1NK1.1 innate lymphocytes. Depletion of NK1.1 cells or neutralization of IL-17A abolished the pro-healing effect, whereas spatial transcriptomics revealed treatment-associated organization of NK cells within vascular niches. Localized treatment further promoted repair in diabetic porcine wounds and human skin models. Together, these findings identify a CHRM1-IL-17 neuroimmune axis linking cholinergic modulation to diabetic wound repair and establish local AChE inhibition as a potential therapeutic strategy for chronic wounds.

immunology↗

bioRxiv2026

Carboxymethyl Dextran Coated Iron Oxide Magnetic Nanoparticles as Vehicles for CRISPR Ribonucleoproteins

The efficient and safe intracellular delivery of CRISPR ribonucleoproteins (RNPs) remains a major challenge for the development of genome editing strategies. Magnetic iron oxide nanoparticles (MNPs) offer an attractive and biocompatible delivery platform. However, their use for the delivery of functional CRISPR RNPs remains comparatively underexplored. Here, we developed carboxymethyl dextran (CMD)-coated MNPs as non-viral carriers for Cas9 and Cas12a RNPs. Nanocomplexes were formed through electrostatic interactions, without the need for chemical coupling reagents or additional surface functionalization. Both RNPs efficiently associated with MNPs, and the resulting nanocomplexes exhibited stable physicochemical properties and retained their cargo in serum-supplemented culture medium for up to 24 hours. Cellular uptake was confirmed by microscopy, while biocompatibility assays revealed no overt cytotoxicity. Functional genome editing was demonstrated at reporter and endogenous gene loci in HEK-293T cells, as well as in primary human dermal fibroblasts. Finally, we explored the therapeutic potential of this platform for recessive dystrophic epidermolysis bullosa (RDEB) by targeting COL7A1 exon 80, with the aim of removing the prevalent c.6527insC mutation while preserving the reading frame. Cas9 RNP nanocomplexes enabled exon 80 deletion. These findings establish CMD-coated MNPs as a versatile platform for the delivery of Cas9 and Cas12a RNPs and support further investigation of magnetic nanoparticle-mediated genome editing for inherited skin disorders.

molecular biology↗

bioRxiv2025

Hairpin-Functionalized Gold Nanoparticles as an Adaptable Platform for Detecting MicroRNA Signatures

Early, accurate, and fast diagnosis is essential to ensure positive health outcomes, through effective treatment interventions and disease control, as well as the study of physiological changes related to gene expression. Liquid biopsies and point-of-care (PoC) detection are valuable tools to achieve this goal, allowing the study and monitoring of a patients molecular profile in a timely and simple manner. MicroRNAs (or miRNAs) have been proposed as biomarkers for detection in liquid biopsies, as they are stable in body fluids and their dysregulation is associated with many diseases. In this work, a sensor based on gold nanoparticles (AuNPs) functionalized with oligonucleotides is reported, aiming for the detection of nucleic acids, in particular miRNAs. This sensor is based on the recognition of target sequences by hairpin-shaped oligonucleotide probes, which allows the modulation of the colloidal stability of the AuNPs, producing color changes detectable with the naked eye. The system is used to detect a panel of miRNAs, demonstrating its versatility for the detection of relevant nucleic acid signatures. The sensor detects single miRNAs with good sensitivity and selectivity and, what is more, it can be used to recognize several miRNAs simultaneously at picomolar concentrations. The system was further adapted to a lateral flow assay (LFA) format, producing a visible colored line on lateral flow test strips, and coupled with isothermal amplification to reach femtomolar detection levels. Its properties make it suitable for use in the point-of-care (PoC), contributing to fast and early detection of pathological and physiological molecular profiles.

molecular biology↗

bioRxiv2025

Human progenitor T-cell differentiation regulated by the mechanical resistance of thymus-mimetic extracellular matrices

Therapeutic T-cell engineering ex vivo from human hematopoietic stem cells (HSCs) focuses on recapitulating notch1-signaling and 4{beta}1-integrin-mediated adhesion within the thymic niche with supportive stromal cell feeder-layers or surface-immobilized recombinant protein-based engineered thymic niches (ETNs). The relevant Notch1-DLL-4 and 4{beta}1-integrin-VCAM-1 interactions are known to respond to mechanical forces that regulate their bond dissociation behaviors and downstream signal transduction, yet manipulating the mechanosensitive features of these key receptor-ligand interactions in thymopoiesis has been largely ignored in current ETN designs. Here, we demonstrate that human T-cell development from cord blood-derived CD34+ HSCs is regulated via molecular cooperativity in notch1 and integrin-mediated mechanotransduction. Mechanically confining interpenetrating network (IPN) hydrogel-based 3D cell culture comprised of collagen type I and alginate polysaccharides functionalized with DLL-4 and VCAM-1 is used as a model viscoelastic 3D ETN to manipulate human progenitor (pro)T-cell differentiation. This ETN enables orthogonal control of the mechanical and biomolecular features of the thymic niche, including thymopoietic ligand density, modulus, and viscoelastic properties (e.g., stress relaxation kinetics). We identify that soft, viscous matrices that enhance activation of the notch1-pathway, and subsequently notch1 intracellular domain (NICD) nuclear import sustain the T-cell development gene regulatory network during proT-cell differentiation. Conversely, stiff, elastic matrices inhibit HSC commitment to the T-lineage, and rather promotes Myeloid-cell differentiation. Our observations indicate mechanical reciprocity in signaling pathways indispensable to thymopoiesis, and highlights extracellular matrix mechanics as a variable in controlling hematopoietic stem cell fate decisions.

bioengineering↗