Cell Loaded GelMA:HEMA IPN hydrogels for corneal stroma engineering.

Stroma is the main refractive element of the cornea and damage to it is one of the main causes of blindness. In this study, cell loaded hydrogels of methacrylated gelatin (GelMA) and poly(2-hydroxyethyl methacrylate) (pHEMA) (8:2) interpenetrating network (IPN) hydrogels were prepared as the corneal stroma substitute and tested in situ and in vitro. Compressive modulus of the GelMA hydrogels was significantly enhanced with the addition of pHEMA in the structure (6.53 vs 155.49 kPa, respectively). More than 90% of the stromal keratocytes were viable in the GelMA and GelMA-HEMA hydrogels as calculated by Live-Dead Assay and NIH Image-J program. Cells synthesized representative collagens and proteoglycans in the hydrogels indicating that they preserved their keratocyte functions. Transparency of the cell loaded GelMA-HEMA hydrogels was increased significantly up to 90% at 700 nm during three weeks of incubation and was comparable with the transparency of native cornea. Cell loaded GelMA-HEMA corneal stroma model is novel and reported for the first time in the literature in terms of introduction of cells during the preparation phase of the hydrogels. The appropriate mechanical strength and high transparency of the cell loaded constructs indicates a viable alternative to the current devices used in the treatment of corneal blindness.

Enhancing CAR Macrophage Efferocytosis Via Surface Engineered Lipid Nanoparticles Targeting LXR Signaling.

The removal of dying cells, or efferocytosis, is an indispensable part of resolving inflammation. However, the inflammatory microenvironment of the atherosclerotic plaque frequently affects the biology of both apoptotic cells and resident phagocytes, rendering efferocytosis dysfunctional. To overcome this problem, a chimeric antigen receptor (CAR) macrophage that can target and engulf phagocytosis-resistant apoptotic cells expressing CD47 is developed. In both normal and inflammatory circumstances, CAR macrophages exhibit activity equivalent to antibody blockage. The surface of CAR macrophages is modified with reactive oxygen species (ROS)-responsive therapeutic nanoparticles targeting the liver X receptor pathway to improve their cell effector activities. The combination of CAR and nanoparticle engineering activated lipid efflux pumps enhances cell debris clearance and reduces inflammation. It is further suggested that the undifferentiated CAR-Ms can transmigrate within a mico-fabricated vessel system. It is also shown that our CAR macrophage can act as a chimeric switch receptor (CSR) to withstand the immunosuppressive inflammatory environment. The developed platform has the potential to contribute to the advancement of next-generation cardiovascular disease therapies and further studies include in vivo experiments.

Self-Assembled Hydrogel Microparticle-Based Tooth-Germ Organoids.

Here, we describe the characterization of tooth-germ organoids, three-dimensional (3D) constructs cultured in vitro with the potential to develop into living teeth. To date, the methods used to successfully create tooth organoids capable of forming functional teeth have been quite limited. Recently, hydrogel microparticles (HMP) have demonstrated utility in tissue repair and regeneration based on their useful characteristics, including their scaffolding ability, effective cell and drug delivery, their ability to mimic the natural tissue extracellular matrix, and their injectability. These outstanding properties led us to investigate the utility of using HMPs (average diameter: 158 ± 32 µm) derived from methacrylated gelatin (GelMA) (degree of substitution: 100%) to create tooth organoids. The tooth organoids were created by seeding human dental pulp stem cells (hDPSCs) and porcine dental epithelial cells (pDE) onto the HMPs, which provided an extensive surface area for the cells to effectively attach and proliferate. Interestingly, the cell-seeded HMPs cultured on low-attachment tissue culture plates with gentle rocking self-assembled into organoids, within which the cells maintained their viability and morphology throughout the incubation period. The self-assembled organoids reached a volume of ~50 mm3 within two weeks of the in vitro tissue culture. The co-cultured hDPSC-HMP and pDE-HMP structures effectively attached to each other without any externally applied forces. The presence of polarized, differentiated dental cells in these composite tooth-bud organoids demonstrated the potential of self-assembled dental cell HMPs to form tooth-bud organoid-like structures for potential applications in tooth regeneration strategies.

Methacrylated gelatin hydrogels as corneal stroma substitutes: in vivo study.

Methacrylated gelatin (GelMA) hydrogels were prepared to serve as corneal stroma equivalents. They were highly transparent (ca. 95% at 700 nm), mechanically strong and withstood handling and had high human corneal keratocyte viability (98%) after 21 days of culture period. In order to test the in vivo performance of the cell free GelMA hydrogels a pilot in vivo study was carried out using eyes of two white New Zealand rabbits. Hydrogel was implanted in a mid-stromal pocket created and without suture fixation, and observed for 8 weeks under a slit lamp. No edema, ulcer formation, inflammation or infection was observed in both the control (sham) and hydrogel implanted corneas. Corneal vascularization on week 3 was treated with one dose of anti-VEGF application. Hematoxylin and Eosin staining showed that the hydrogel was integrated with the host tissue with only a minimal foreign body reaction. Results demonstrated some degradation in the construct within 8 weeks as evidenced by the decrease of the diameter of the hydrogel from 4 mm to 2.6 mm. High transparency, adequate mechanical strength, biocompatibility and well integration with the host tissue, indicates that this hydrogel is a viable alternative to the current methods for the treatment of corneal blindness and deserves testing on larger number of rabbits and more extensively using microscopy, histology and immune histochemistry.

A bilayer scaffold prepared from collagen and carboxymethyl cellulose for skin tissue engineering applications.

Treatment of chronic skin wound such as diabetic ulcers, burns, pressure wounds are challenging problems in the medical area. The aim of this study was to design a bilayer skin equivalent mimicking the natural one to be used as a tissue engineered skin graft for use in the treatments of problematic wounds, and also as a model to be used in research related to skin, such as determination of the efficacy of transdermal bioactive agents on skin cells and treatment of acute skin damages that require immediate response. In this study, the top two layers of the skin were mimicked by producing a multilayer construct combining two different porous polymeric scaffolds: as the dermis layer a sodium carboxymethyl cellulose (NaCMC) hydrogel on which fibroblasts were added, and as the epidermis layer collagen (Coll) or chondroitin sulfate-incorporated collagen (CollCS) on which keratinocytes were added. The bilayer construct was designed to allow cross-talk between the two cell populations in the subsequent layers and achieves paracrine signalling. It had interconnected porosity, high water content, appropriate stability and elastic moduli. Expression of vascular endothelial growth factor (VEGF), basic-fibroblast growth factor (bFGF) and Interleukin 8 (IL-8), and the production of collagen I, collagen III, laminin and transglutaminase supported the attachment and proliferation of cells on both layers of the construct. Attachment and proliferation of fibroblasts on NaCMC were lower compared to performance of keratinocyte on collagen where keratinocytes created a dense and a stratified layer similar to epidermis. The resulting constructs succesfully mimicked in vitro the natural skin tissue. They are promising as grafts for use in the treatment of deep wounds and also as models for the study of the efficacy of bioactive agents on the skin.