Cultivated meat, also known as cell-based or clean meat, utilizes mesenchymal stem cells to cultivate mature cell types like adipocytes, which are pivotal for imparting the desired taste and texture. The delivery of differentiated cells, crucial in cultivated meat production, is facilitated through extensive exploration of 3D culturing techniques mimicking physiological environments. In this study, we investigated the adipogenic differentiation potential of bovine umbilical cord stem cells (BUSCs), sourced from discarded birth tissue, and assessed the feasibility of delivering differentiated cells for cultivated meat using gelatin methacrylate (GelMA) as a carrier for adipose tissue. Various adipogenic inducers, previously reported to be effective for human mesenchymal stem cells (hMSCs), were evaluated individually or in combination for their efficacy in promoting the adipogenesis of BUSCs. Surprisingly, while the traditional adipogenic inducers, including insulin, dexamethasone, isobutylmethylxantine (IBMX), indomethacin, and rosiglitazone, showed no significant effect on the adipogenic differentiation of BUSCs, efficient differentiation was achieved in the presence of a fatty acid cocktail. Furthermore, we explored methods for the delivery of BUSCs. Differentiated cells were delivered either encapsulated in GelMA hydrogel or populated on the surface of GelMA microparticles (MPs) as the adipose component of cultivated meat. Our findings reveal that after adipogenic induction, the lipid production per cell was comparable when cultured either within hydrogel or on MPs. However, GelMA-MPs supported better cell growth compared to hydrogel encapsulation. Consequently, the overall lipid production is higher when BUSCs are delivered via GelMA-MPs rather than encapsulation. This study not only systematically evaluated the impact of common adipogenic inducers on BUSCs, but also identified GelMA-MPs as a promising carrier for delivering bovine adipocytes for cultivated meat production.

The demand for innovative therapeutic interventions to expedite wound healing, particularly in vulnerable populations such as aging and diabetic patients, has prompted the exploration of novel strategies. Mesenchymal stem cell (MSC)-based therapy emerges as a promising avenue for treating acute and chronic wounds. However, its clinical application faces persistent challenges, notably the low survivability and limited retention time of engraftment in wound environments. Addressing this, a strategy to sustain the viability and functionality of human MSCs (hMSCs) in a graft-able format has been identified as crucial for advanced wound care. Hydrogel microparticles (HMPs) emerge as promising entities in the field of wound healing, showcasing versatile capabilities in delivering both cells and bioactive molecules/drugs. In this study, gelatin HMPs (GelMPs) were synthesized via an optimized mild processing method. GelMPs with distinct diameter sizes were sorted and characterized. The growth of hMSCs on GelMPs with various sizes was evaluated. The release of wound healing promoting factors from hMSCs cultured on different GelMPs were assessed using scratch wound assays and gene expression analysis. GelMPs with a size smaller than 100 microns supported better cell growth and cell migration compared to larger sizes (100 microns or 200 microns). While encapsulation of hMSCs in hydrogels has been a common route for delivering viable hMSCs, we hypothesized that hMSCs cultured on GelMPs are more robust than those encapsulated in hydrogels. To test this hypothesis, hMSCs were cultured on GelMPs or in the cross-linked methacrylated gelatin hydrogel (GelMA). Comparative analysis of growth and wound healing effects revealed that hMSCs cultured on GelMPs exhibited higher viability and released more wound healing activities in vitro. This observation highlights the potential of GelMPs, especially those with a size smaller than 100 microns, as a promising carrier for delivering hMSCs in wound healing applications, providing valuable insights for the optimization of advanced therapeutic strategies.

Aim: To develop a novel stabilizing agent for silver nanoparticles (AgNPs) with the aim of enhancing its antibacterial efficacy against wound associated pathogens while mitigating their cytotoxic e...

Cultivated meat is a fast-growing research field and an industry with great potential to overcome the limitations of traditional meat production. Cultivated meat utilizes cell culture and tissue engineering technologies to culture a vast number of cells in vitro and grow/assemble them into structures mimicking the muscle tissues of livestock animals. Stem cells with self-renewal and lineage-specific differentiation abilities have been considered one of the key cell sources for cultivated meats. However, the extensive in vitro culturing/expansion of stem cells results in a reduction in their abilities to proliferate and differentiate. Extracellular matrix (ECM) has been used as a culturing substrate to support cell expansion for cell-based therapies in regenerative medicine due to its resemblance to the native microenvironment of cells. In this study, the effect of the ECM on the expansion of bovine umbilical cord stromal cells (BUSC) in vitro was evaluated and characterized. BUSCs with multi-lineage differentiation potentials were isolated from bovine placental tissue. Decellularized ECM prepared from a confluent monolayer of bovine fibroblasts (BF) is free of cellular components but contains major ECM proteins such as fibronectin and type I collagen and ECM-associated growth factors. Expansion of BUSC on ECM for three passages (around three weeks) resulted in about 500-fold amplification, while cells were amplified less than 10-fold when cultured on standard tissue culture plates (TCP). Moreover, the presence of ECM reduced the requirement for serum in the culture medium. Importantly, the cells amplified on ECM retained their differentiation abilities better than cells cultured on TCP. The results of our study support the notion that monolayer cell-derived ECM may be a strategy to expand bovine cells in vitro effectively and efficiently.

Cartilage damage is common in sports injuries and cartilage-related diseases, such as degenerative joint and rheumatic disorders. Autologous chondrocyte implantation (ACI) is a widely used cell-based therapy for repairing cartilage damage in clinical practice. In this procedure, a patient’s chondrocytes are isolated, cultured in vitro to expand the cell population, and then implanted into the damaged site. However, in vitro expansion of chondrocytes on standard 2D culture surfaces leads to dedifferentiation (loss of the chondrocyte phenotype), and the delivery of detached cells has proven to be ineffective. To overcome these limitations, the matrix-assisted ACI (MACI) procedure was developed. In MACI, matrices such as hydrogels and microspheres are used as cell carriers or scaffolds to deliver expanded chondrocytes, enhancing cell viability and precision delivery. To streamline the two key steps of MACI—cell expansion and delivery—this study aims to investigate various configurations of gelatin-based hydrogels for their potential to support both cell expansion and delivery as a single step. This study evaluated gelatin microspheres (Gel MS), micronized photo-crosslinked GelMA microparticles (GelMA MP), and bulky GelMA hydrogels containing cells (GelMA HG). Cell growth, maintenance of the chondrocyte phenotype, and cartilage extracellular matrix (ECM) production were assessed in pellet cultures for cells grown on/in these carriers, compared with cells cultured on tissue culture-treated polystyrene (TCP). Our results demonstrate that normal human knee articular chondrocytes exhibit robust growth on Gel MS and form aggregates enriched with glycosaminoglycan-rich ECM. Gel MS outperformed both GelMA MP and GelMA HG as a cell carrier by both supporting long-term cell growth with reduced dedifferentiation and precision delivery.

Volumetric muscle loss (VML) is a devastating injury in which extensive skeletal muscle loss overwhelms endogenous healing. Despite their therapeutic efficacy, acellular constructs fail to provide a comprehensive set of bioactive cues required for muscle development. Notably, very few constructs provide bioelectrical cues necessary for muscle development. To address that need, this study aimed to develop a myogenic platform that provides a comprehensive set of bioactive cues, including bioelectrical cues that can be delivered wirelessly, to promote skeletal muscle development. To achieve this, we engineered a microchanneled, piezoelectric, 3D-printed platform based on a unique interpenetrating hydrogel network of gelatin methacrylate and poly(acrylic acid) that incorporates barium titanate nanoparticles. Rheological studies showed stable gel-like viscoelasticity and shear-thinning behavior of the hydrogels, indicating their suitability for extrusion-3D printing. Physicochemical characterization demonstrated the platform’s structural fidelity, precise microchannel definition, pronounced flexibility, and self-adhesiveness. While the platform’s stiffness was comparable to that of skeletal muscle, it also generated a surface potential of ∼90 mV when deformed by ultrasound (US), highlighting its piezoelectric nature and ability to harness electromechanically triggered bioelectrical cues. FE-SEM and immunofluorescence analyses revealed that microchannels and US-induced piezoelectricity from the platform can synergistically help form dense, compact, and aligned myotubes. Notably, we demonstrate that the US-induced piezoelectric platforms can promote early-stage myotube development, enhance myogenic progression, and help form mature, elongated, thick myotubes over time. Finally, we observe that the piezoelectric platforms exhibit increased FAK and MAPK activation in adherent cells over time, suggesting that these pathways can contribute to transducing electromechanical cues that promote myotube maturation. Overall, this is the first-of-its-kind comprehensive study to engineer a platform that delivers a comprehensive set of bioactive cues, specifically wirelessly delivered bioelectrical cues, resulting in a spatiotemporally bioinstructive microenvironment for skeletal muscle development. We believe this unique platform could be a promising regenerative therapy for VML.