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Abstract CRISPR gene editing offers unprecedented genomic and transcriptomic control for precise regulation of cell function and phenotype. However, delivering the necessary CRISPR components to therapeutically relevant cell types without cytotoxicity or unexpected side effects remains challenging. The RALA cell penetrating peptide is an amphiphilic peptide that self‐assembles into nanoparticles through electrostatic interactions with anionic molecules and delivers them across the cell membrane. Given the low cytotoxicity, versatility, and competitive transfection rates of RALA, we aimed to establish this peptide as a new CRISPR delivery system in a wide range of molecular formats across different editing modalities. We report that RALA effectively encapsulated and delivered CRISPR DNAs, RNAs, and ribonucleic proteins (RNPs) to primary mesenchymal stem cells (MSCs), outperforming commercially available reagents. We then used the RALA peptide for the knock‐in and knock‐out of reporter genes into primary MSCs and the transcriptional activation of therapeutically relevant genes. Finally, we demonstrate in vivo gene editing using RALA to knock‐out luciferase and GFP in a reporter mouse model. In summary, we establish RALA as a powerful tool for safer and more effective delivery of CRISPR machinery in multiple cargo formats for a wide range of ex vivo and in vivo gene editing strategies. 

Osteoarthritis (OA) is the most prevalent musculoskeletal disorder and a leading cause of disability globally. Although many efforts have been made to treat this condition, current tissue engineering (TE) and regenerative medicine strategies fail to address the inflammatory tissue environment that leads to the rapid progression of the disease and prevents cartilage tissue formation. First, this review addresses in detail the current antiinflammatory therapies for OA with a special emphasis on pharmacological approaches, gene therapy, and mesenchymal stromal cell (MSC) intra-articular administration, and discusses the reasons behind the limited clinical success of these approaches at enabling cartilage regeneration. Then, we analyze the state-of-the-art TE strategies and how they can be improved by incorporating immunomodulatory capabilities such as the optimization of biomaterial composition, porosity and geometry, and the loading of anti-inflammatory molecules within an engineered structure. Finally, the review discusses the future directions for the new generation of TE strategies for OA treatment, specifically focusing on the spatiotemporal modulation of anti-inflammatory agent presentation to allow for tailored patient-specific therapies. 

Abstract The biochemical and physical properties of a scaffold can be tailored to elicit specific cellular responses. However, it is challenging to decouple their individual effects on cell‐material interactions. Here, we solvent‐cast 3D printed different ratios of high and low molecular weight (MW) poly(caprolactone) (PCL) to fabricate scaffolds with significantly different stiffnesses without affecting other properties. Ink viscosity was used to match processing conditions between inks and generate scaffolds with the same surface chemistry, crystallinity, filament diameter, and architecture. Increasing the ratio of low MW PCL resulted in a significant decrease in modulus. Scaffold modulus did not affect human mesenchymal stromal cell (hMSC) differentiation under osteogenic conditions. However, hMSC response was significantly affected by scaffold stiffness in chondrogenic media. Low stiffness promoted more stable chondrogenesis whereas high stiffness drove hMSC progression toward hypertrophy. These data illustrate how this versatile platform can be used to independently modify biochemical and physical cues in a single scaffold to synergistically enhance desired cellular response.