Here, we developed a combinatorial delivery platform for chronic wound healing applications. A microfluidic system was utilized to form a series of biopolymer-based microparticles with enhanced affinity to encapsulate and deliver vascular endothelial growth factor (VEGF). Presence of heparin into the structure can significantly increase the encapsulation efficiency up to 95% and lower the release rate of encapsulated VEGF. Our in vitro results demonstrated that sustained release of VEGF from microparticles can promote capillary network formation and sprouting of endothelial cells in 2D and 3D microenvironments. These engineered microparticles can also encapsulate antibiotic-loaded nanoparticles to offer a dual delivery system able to fight bacterial infection while promoting angiogenesis. We believe this highly tunable drug delivery platform can be used alone or in combination with other wound care products to improve the wound healing process and promote tissue regeneration.

Infection by SARS-CoV-2 virus often induces the dysregulation of immune responses, tissue damage, and blood clotting. Engineered biomaterials from the nano- to the macroscale can provide targeted drug delivery, controlled drug release, local immunomodulation, enhanced immunity, and other desirable functions to coordinate appropriate immune responses and to repair tissues. Based on the understanding of COVID-19 disease progression and immune responses to SARS-CoV-2, we discuss possible immunotherapeutic strategies and highlight biomaterial approaches from the perspectives of preventive immunization, therapeutic immunomodulation, and tissue healing and regeneration. Successful development of biomaterial platforms for immunization and immunomodulation will not only benefit COVID-19 patients, but also have broad applications for a variety of infectious diseases.

Periodontal disease begins as an inflammatory response to a bacterial biofilm deposited around the teeth, which over time leads to the destruction of tooth-supporting structures and consequently tooth loss. Conventional treatment strategies show limited efficacy in promoting regeneration of damaged periodontal tissues. Here, a delivery platform is developed for small extracellular vesicles (sEVs) derived from gingival mesenchymal stem cells (GMSCs) to treat periodontitis. EVs can achieve comparable therapeutic effects to their cells of origin. However, the short half-lives of EVs after their administration along with their rapid diffusion away from the delivery site necessitate frequent administration to achieve therapeutic benefits. To address these issues, "dual delivery" microparticles are engineered enabling microenvironment-sensitive release of EVs by metalloproteinases at the affected site along with antibiotics to suppress bacterial biofilm growth. GMSC sEVs are able to decrease the secretion of pro-inflammatory cytokines by monocytes/macrophages and T cells, suppress T-cell activation, and induce the formation of T regulatory cells (Tregs) in vitro and in a rat model of periodontal disease. One-time administration of immunomodulatory GMSC sEV-decorated microparticles leads to a significant improvement in regeneration of the damaged periodontal tissue. This approach will have potential clinical applications in the regeneration of a variety of tissues.

Mesenchymal stem cells (MSCs) derived from dental and orofacial tissues provide an alternative therapeutic option for craniofacial bone tissue regeneration. However, there is still a need to improve stem cell delivery vehicles to regulate the fate of the encapsulated MSCs for high quality tissue regeneration. Matrix elasticity plays a vital role in MSC fate determination. Here, we have prepared various hydrogel formulations based on alginate and gelatin methacryloyl (GelMA) and have encapsulated gingival mesenchymal stem cells (GMSCs) and human bone marrow MSCs (hBMMSCs) within these fabricated hydrogels. We demonstrate that addition of the GelMA to alginate hydrogel reduces the elasticity of the hydrogel mixture. While presence of GelMA in an alginate-based scaffold significantly increased the viability of encapsulated MSCs, increasing the concentration of GelMA downregulated the osteogenic differentiation of encapsulated MSCs in vitro due to decrease in the stiffness of the hydrogel matrix. The osteogenic suppression was rescued by addition of a potent osteogenic growth factor such as rh-BMP-2. In contrast, MSCs encapsulated in alginate hydrogel without GelMA were successfully osteo-differentiated without the aid of additional growth factors, as confirmed by expression of osteogenic markers (Runx2 and OCN), as well as positive staining using Xylenol orange. Interestingly, after two weeks of osteo-differentiation, hBMMSCs and GMSCs encapsulated in alginate/GelMA hydrogels still expressed CD146, an MSC surface marker, while MSCs encapsulated in alginate hydrogel failed to express any positive staining. Altogether, our findings suggest that it is possible to control the fate of encapsulated MSCs within hydrogels by tuning the mechanical properties of the matrix. We also reconfirmed the important role of the presence of inductive signals in guiding MSC differentiation. These findings may enable the design of new multifunctional scaffolds for spatial and temporal control over the fate and function of stem cells even post-transplantation. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2957-2967, 2017.

Activation of T cells by antigen presenting cells (APCs) initiates their proliferation, cytokine production, and killing of infected or cancerous cells. We and others have shown that T-cell receptors require mechanical forces for triggering, and these forces arise during the interaction of T cells with APCs. Efficient activation of T cells in vitro is necessary for clinical applications. In this paper, we studied the impact of combining mechanical, oscillatory movements provided by an orbital shaker with soft, biocompatible, artificial APCs (aAPCs) of various sizes and amounts of antigen. We showed that these aAPCs allow for testing the strength of signal delivered to T cells, and enabled us to confirm that that absolute amounts of antigen engaged by the T cell are more important for activation than the density of antigen. We also found that when our aAPCs interact with T cells in the context of an oscillatory mechanoenvironment, they roughly double antigenic signal strength, compared to conventional, static culture. Combining these effects, our aAPCs significantly outperformed the commonly used Dynabeads. We finally demonstrated that tuning the signal strength down to a submaximal "sweet spot" allows for robust expansion of induced regulatory T cells. In conclusion, augmenting engineered aAPCs with mechanical forces offers a novel approach for tuning of T-cell activation and differentiation.

Cell clusters that collectively migrate from primary tumors appear to be far more potent in forming distant metastases than single cancer cells. A better understanding of the collective cell migration phenomenon and the involvement of various cell types during this process is needed. Here, an in vitro platform based on inverted-pyramidal microwells to follow and quantify the collective migration of hundreds of tumor cell clusters at once is developed. These results indicate that mesenchymal stromal cells (MSCs) or cancer-associated fibroblasts (CAFs) in the heterotypic tumor cell clusters may facilitate metastatic dissemination by transporting low-motile cancer cells in a Rac-dependent manner and that extracellular vesicles secreted by mesenchymal cells only play a minor role in this process. Furthermore, in vivo studies show that cancer cell spheroids containing MSCs or CAFs have faster spreading rates. These findings highlight the active role of co-traveling stromal cells in the collective migration of tumor cell clusters and may help in developing better-targeted therapies.

The host immune system (T-lymphocytes and their pro-inflammatory cytokines) has been shown to compromise bone regeneration ability of mesenchymal stem cells (MSCs). We have recently shown that hydrogel, used as an encapsulating biomaterial affects the cross-talk among host immune cells and MSCs. However, the role of hydrogel elasticity and porosity in regulation of cross-talk between dental-derived MSCs and immune cells is unclear. In this study, we demonstrate that the modulus of elasticity and porosity of the scaffold influence T-lymphocyte-dental MSC interplay by regulating the penetration of inflammatory T cells and their cytokines. Moreover, we demonstrated that alginate hydrogels with different elasticity and microporous structure can regulate the viability and determine the fate of the encapsulated MSCs through modulation of NF-kB pathway. Our in vivo data show that alginate hydrogels with smaller pores and higher elasticity could prevent pro-inflammatory cytokine-induced MSC apoptosis by down-regulating the Caspase-3- and 8- associated proapoptotic cascades, leading to higher amounts of ectopic bone regeneration. Additionally, dental-derived MSCs encapsulated in hydrogel with higher elasticity exhibited lower expression levels of NF-kB p65 and Cox-2 in vivo. Taken together, our findings demonstrate that the mechanical characteristics and microarchitecture of the microenvironment encapsulating MSCs, in addition to presence of T-lymphocytes and their pro-inflammatory cytokines, affect the fate of encapsulated dental-derived MSCs.

Statement of significance

In this study, we demonstrate that alginate hydrogel regulates the viability and the fate of the encapsulated dental-derived MSCs through modulation of NF-kB pathway. Alginate hydrogels with smaller pores and higher elasticity prevent pro-inflammatory cytokine-induced MSC apoptosis by down-regulating the Caspase-3- and 8- associated proapoptotic cascade, leading to higher amounts of ectopic bone regeneration. MSCs encapsulated in hydrogel with higher elasticity exhibited lower expression levels of NF-kB p65 and Cox-2 in vivo. These findings confirm that the fate of encapsulated MSCs are affected by the stiffness and microarchitecture of the encapsulating hydrogel biomaterial, as well as presence of T-lymphocytes/pro-inflammatory cytokines providing evidence concerning material science, stem cell biology, the molecular mechanism of dental-derived MSC-associated therapies, and the potential clinical therapeutic impact of MSCs.

T cells recognize mechanical forces through a variety of cellular pathways, including mechanical triggering of both the T-cell receptor (TCR) and integrin LFA-1. Here we show that T cells can recognize forces arising from the mechanical rigidity of the microenvironment. We fabricated 3D scaffold matrices with mechanical stiffness tuned to the range 4-40 kPa and engineered them to be microporous, independently of stiffness. We cultured T cells and antigen presenting cells within the matrices and studied T-cell activation by flow cytometry and live-cell imaging. We found that there was an augmentation of T-cell activation, proliferation, and migration speed in the context of mechanically stiffer 3D matrices as compared to softer materials. These results show that T cells can sense their 3D mechanical environment and alter both their potential for activation and their effector responses in different mechanical environments. A 3D scaffold of tunable stiffness and consistent microporosity offers a biomaterial advancement for both translational applications and reductionist studies on the impact of tissue microenvironmental factors on cellular behavior.

Recent innovations in immunoregulatory treatments have demonstrated both the impressive potential and vital role of T cells in fighting cancer. These treatments come at a cost, with systemic side effects including life-threatening autoimmunity and immune dysregulation the norm. Here, we developed an approach to locally synthesize immune therapies and in this way, avoid systemic toxicity. Rather than just encapsulating cytokines, we endowed our nanoparticles with transcriptional and translational machinery to make cytokines locally, in situ, and on demand (activated by light). We demonstrated the capabilities of these particles in vitro and in vivo, in a mouse model of melanoma, and showed that tumor-infiltrating T cells were more highly activated in the context of these "microfactory" particles that make the synthetic cytokine.

A tumour microenvironment abundant in regulatory T (T_reg) cells aids solid tumours to evade clearance by effector T cells. Systemic strategies to suppress T_reg cells or to augment immunity can elicit autoimmune side effects, cytokine storms and other toxicities. Here we report the design, fabrication and therapeutic performance of a biodegradable macroporous scaffold, implanted peritumourally, that releases a small-molecule inhibitor of transforming growth factor β to suppress T_reg cells, chemokines to attract effector T cells and antibodies to stimulate them. In two mouse models of aggressive tumours, the implant boosted the recruitment and activation of effector T cells into the tumour and depleted it of T_reg cells, which resulted in an 'immunological abscopal effect' on distant metastases and in the establishment of long-term memory that impeded tumour recurrence. We also show that the scaffold can be used to deliver tumour-antigen-specific T cells into the tumour. Peritumourally implanted immunomodulatory scaffolds may represent a general strategy to enhance T-cell immunity and avoid the toxicities of systemic therapies.