3D Bioprinting of Artificial Skin Substitute with Improved Mechanical Property and Regulated Cell Behavior through Integrating Patterned Nanofibrous Films.

Three-dimensional (3D) bioprinting has advantages for constructing artificial skin tissues in replicating the structures and functions of native skin. Although many studies have presented improved effect of printing skin substitutes in wound healing, using hydrogel inks to fabricate 3D bioprinting architectures with complicated structures, mimicking mechanical properties, and appropriate cellular environments is still challenging. Inspired by collagen nanofibers withstanding stress and regulating cell behavior, a patterned nanofibrous film was introduced to the printed hydrogel scaffold to fabricate a composite artificial skin substitute (CASS). The artificial dermis was printed using gelatin-hyaluronan hybrid hydrogels containing human dermal fibroblasts with gradient porosity and integrated with patterned nanofibrous films simultaneously, while the artificial epidermis was formed by seeding human keratinocytes upon the dermis. The collagen-mimicking nanofibrous film effectively improved the tensile strength and fracture resistance of the CASS, making it sewable for firm implantation into skin defects. Meanwhile, the patterned nanofibrous film also provided the biological cues to guide cell behavior. Consequently, CASS could effectively accelerate the regeneration of large-area skin defects in mouse and pig models by promoting re-epithelialization and collagen deposition. This research developed an effective strategy to prepare composite bioprinting architectures for enhancing mechanical property and regulating cell behavior, and CASS could be a promising skin substitute for treating large-area skin defects.

4D printed hydrogel scaffold with swelling-stiffening properties and programmable deformation for minimally invasive implantation.

The power of three-dimensional printing in designing personalized scaffolds with precise dimensions and properties is well-known. However, minimally invasive implantation of complex scaffolds is still challenging. Here, we develop amphiphilic dynamic thermoset polyurethanes catering for multi-material four-dimensional printing to fabricate supportive scaffolds with body temperature-triggered shape memory and water-triggered programmable deformation. Shape memory effect enables the two-dimensional printed pattern to be fixed into temporary one-dimensional shape, facilitating transcatheter delivery. Upon implantation, the body temperature triggers shape recovery of the one-dimensional shape to its original two-dimensional pattern. After swelling, the hydrated pattern undergoes programmable morphing into the desired three-dimensional structure because of swelling mismatch. The structure exhibits unusual soft-to-stiff transition due to the water-driven microphase separation formed between hydrophilic and hydrophobic chain segments. The integration of shape memory, programmable deformability, and swelling-stiffening properties makes the developed dynamic thermoset polyurethanes promising supportive void-filling scaffold materials for minimally invasive implantation.

miR-204 ameliorates osteoarthritis pain by inhibiting SP1-LRP1 signaling and blocking neuro-cartilage interaction.

Osteoarthritis (OA) is a painful degenerative joint disease and is the leading cause of chronic disability among elderly individuals. To improve the quality of life for patients with OA, the primary goal for OA treatment is to relieve the pain. During OA progression, nerve ingrowth was observed in synovial tissue and articular cartilage. These abnormal neonatal nerves act as nociceptors to detect OA pain signals. The molecular mechanisms for transmitting OA pain in the joint tissues to the central nerve system (CNS) is currently unknown. MicroRNA miR-204 has been demonstrated to maintain the homeostasis of joint tissues and have chondro-protective effect on OA pathogenesis. However, the role of miR-204 in OA pain has not been determined. In this study, we investigated interactions between chondrocytes and neural cells and evaluated the effect and mechanism of miR-204 delivered by exosome in the treatment of OA pain in an experimental OA mouse model. Our findings demonstrated that miR-204 could protect OA pain by inhibition of SP1- LDL Receptor Related Protein 1 (LRP1) signaling and blocking neuro-cartilage interaction in the joint. Our studies defined novel molecular targets for the treatment of OA pain.

Engineering stem cells to produce exosomes with enhanced bone regeneration effects: an alternative strategy for gene therapy.

BACKGROUND: Exosomes derived from stem cells have been widely studied for promoting regeneration and reconstruction of multiple tissues as "cell-free" therapies. However, the applications of exosomes have been hindered by limited sources and insufficient therapeutic potency. RESULTS: In this study, a stem cell-mediated gene therapy strategy is developed in which mediator mesenchymal stem cells are genetically engineered by bone morphogenetic protein-2 gene to produce exosomes (MSC-BMP2-Exo) with enhanced bone regeneration potency. This effect is attributed to the synergistic effect of the content derived from MSCs and the up-regulated BMP2 gene expression. The MSC-BMP2-Exo also present homing ability to the injured site. The toxic effect of genetical transfection vehicles is borne by mediator MSCs, while the produced exosomes exhibit excellent biocompatibility. In addition, by plasmid tracking, it is interesting to find a portion of plasmid DNA can be encapsulated by exosomes and delivered to recipient cells. CONCLUSIONS: In this strategy, engineered MSCs function as cellular factories, which effectively produce exosomes with designed and enhanced therapeutic effects. The accelerating effect in bone healing and the good biocompatibility suggest the potential clinical application of this strategy.

Microfluidic fabrication of inhalable large porous microspheres loaded with H2S-releasing aspirin derivative for pulmonary arterial hypertension therapy.

Hydrogen sulfide (H2S) has recently emerged as a novel gaseous mediator with protective actions in the treatment of pulmonary arterial hypertension (PAH). However, the therapeutic potential of H2S in PAH has been substantially hampered due to the lack of appropriate donors that could mimic the slow and continuous generation of H2S in vivo. Large porous microspheres (LPMs) have low density and large surface area leading to excellent absorption capabilities and aerodynamic properties. They are extensively studied as pulmonary delivery carriers for controlled and sustained release of drug molecules in the treatment of pulmonary disorders. Therefore, we hypothesized that LPMs containing H2S-releasing aspirin derivative (ACS14), a novel synthetic H2S donor may be a feasible option to facilitate the use of H2S in PAH treatment. LPMs were prepared with a biodegradable polymer, poly(lactic-co-glycolic acid) (PLGA) by a microfluidic technique. Surface morphology, lung deposition characteristics, safety and H2S release profiles of the formulation were evaluated. The resulting ACS14-containing LPMs (ACS14 MSs) displayed excellent aerodynamic properties (mass median aerodynamic diameter of 4.4 ± 0.4 µm), desirable drug loading and entrapment efficiency (25.8 ± 2.7% and 77.4 ± 6.9%, respectively) with slow and sustained H2S release for 24 h and negligible cytotoxicity (~95% cell viability). Daily intratracheally administered with ACS14 MSs elicited improvement in the severity of PAH in a rat model of monocrotaline-induced PAH, with comparable efficacy to oral administration with sildenafil, a conventional PAH treatment. It also inhibited the process of endothelial-to-mesenchymal transition (EndMT), an important process in vascular remodeling of PAH by suppressing the induction of NF-κB-Snail pathway. Moreover, ACS14 MSs dose-dependently inhibited TGF-ß1-induced EndMT and the activation of NF-κB-Snail pathway in human pulmonary artery endothelial cells. In conclusion, our findings demonstrated that the designed microfluidics-assisted ACS14-containing LPMs have shown great potential to be used as an inhalable and efficacious H2S donor in the treatment of PAH.

Modeling Endothelialized Hepatic Tumor Microtissues for Drug Screening.

Compared to various traditional 2D approaches, the scaffold-based 3D tumor models have emerged as an effective strategy to investigate the complex mechanisms behind cancer progression and responses to drug treatments, by providing biomimetic extracellular matrix and stromal-like microenvironments including the vascular elements. Herein, the development of a 3D endothelialized hepatic tumor microtissue model based on the fusion of multicellular aggregates of human hepatocellular carcinoma cells and human umbilical vein endothelial cells cocultured in poly(lactic-co-glycolic acid)-based porous microspheres (PLGA PMs) is reported. In contrast to the conventional 2D culture, the cells within the PLGA PMs exhibit significantly higher half-maximal inhibitory concentration values against anticancer drugs, including doxorubicin and cisplatin. Furthermore, the feasibility of coculturing other cell types, such as fibroblasts (L929) and HepG2 cells, is investigated. Together, the findings emphasize the significance of engineered 3D hepatic tumor microtissue models using PLGA PM-based multicellular aggregates for drug screening applications.

Supercritical Fluid-Assisted Porous Microspheres for Efficient Delivery of Insulin and Inhalation Therapy of Diabetes.

Pulmonary delivery of drugs has attracted increasing attention in healthcare, as the lungs are an easily accessible site for noninvasive systemic delivery of drugs. Although pulmonary inhalation of porous microparticles has been shown to sustain drug delivery, there are limited reports on efficient delivery of insulin and inhalation therapy of diabetes based on supercritical carbon dioxide (SC-CO2 ) technology. Herein, this study reports the fabrication of insulin-loaded poly-l-lactide porous microspheres (INS-PLLA PMs) by using the SC-CO2 technology, and their use as an inhalation delivery system potentially for diabetes therapy. Biocompatibility and delivery efficiency of the PLLA PMs in the lungs are investigated. The PLLA PMs show negligible toxicity to lung-derived cells, resulting in no significant reduction in cell viability, as well as levels of various inflammatory mediators such as interleukin (IL)-6, IL-8, and tumor necrosis factor-α, compared with the negative control group. INS-PLLA PMs are further efficiently deposited in the trachea and the bronchi of superior lobes of the lungs, which exhibit pronounced hypoglycemic activity in induced diabetic rats. Together, the results demonstrate that the INS-PLLA PMs have a strong potential as an effective strategy for inhalation treatment of diabetes.