Assembling and decorating hyaluronan hydrogels with twin protein superglues to mimic cell-cell interactions

Simple polymeric scaffolds have yielded dramatic effects on cell behavior. For more sophisticated phenotypes, precise and efficient chemistries are desired to incorporate proteins into these scaffolds. Here we derivatize hyaluronan with an elastin-like polypeptide containing telechelic SpyTags (HA-SpyTag). Our second network component, the TriCatcher protein, had two SpyCatchers and a terminal SnoopCatcher. Mixing HA-SpyTag with TriCatcher led to rapid hydrogel formation, via spontaneous amidation. SnoopCatcher allowed modular network decoration with SnoopTagJr-linked adhesion molecules, through orthogonal transamidation. This programmed scaffold enables the testing of how individual matrix-anchored protein interactions affect cell behavior. Epithelial cell adhesion molecule (EpCAM) regulates cell behavior and migration, with important effects in cancer. EpCAM-anchoring to the hydrogel induced disassembly of non-malignant mammary spheres in 3D culture. Integrating signaling proteins into biomaterials via efficient biocompatible chemistry should reveal key cues to control cell behavior

Assembling and decorating hyaluronan hydrogels with twin protein superglues to mimic cell-cell interactions

Simple polymeric scaffolds have yielded dramatic effects on cell behavior. For more sophisticated phenotypes, precise and efficient chemistries are desired to incorporate proteins into these scaffolds. Here we derivatize hyaluronan with an elastin-like polypeptide containing telechelic SpyTags (HA-SpyTag). Our second network component, the TriCatcher protein, had two SpyCatchers and a terminal SnoopCatcher. Mixing HA-SpyTag with TriCatcher led to rapid hydrogel formation, via spontaneous amidation. SnoopCatcher allowed modular network decoration with SnoopTagJr-linked adhesion molecules, through orthogonal transamidation. This programmed scaffold enables the testing of how individual matrix-anchored protein interactions affect cell behavior. Epithelial cell adhesion molecule (EpCAM) regulates cell behavior and migration, with important effects in cancer. EpCAM-anchoring to the hydrogel induced disassembly of non-malignant mammary spheres in 3D culture. Integrating signaling proteins into biomaterials via efficient biocompatible chemistry should reveal key cues to control cell behavior

Layer-by-Layer Assembly of Heparin and Peptide-Polyethylene Glycol Conjugates to Form Hybrid Nanothin Films of Biomatrices.

We investigated the utility of a heparin/peptide-polyethylene glycol conjugate system to build layer-by-layer (LbL) structures, to assemble tailored multilayer-biomatrices for cell culture. The LbL assembly balances the advantages of polyelectrolyte systems and protein-based systems. Human umbilical vein endothelial cells showed distinct responses to the film thickness and structure; the presence, density, and spatial arrangement of a cell adhesion ligand within the nanothin film; and the pretreatment of the film with morphogens. The LbL technique presents a versatile tool for modifying cell culture substrates with defined and diverse biochemical and structural features, for investigating cell-material interactions

Noncovalent hydrogel beads as microcarriers for cell culture.

Hydrogel beads as microcarriers could have many applications in biotechnology. However, bead formation by noncovalent cross-linking to achieve high cell compatibility by avoiding chemical reactions remains challenging because of rapid gelation rates and/or low stability. Here we report the preparation of homogeneous, tunable, and robust hydrogel beads from peptide-polyethylene glycol conjugates and oligosaccharides under mild, cell-compatible conditions using a noncovalent crosslinking mechanism. Large proteins can be released from beads easily. Further noncovalent modification allows for bead labeling and functionalization with various compounds. High survival rates of embedded cells were achieved under standard cell culture conditions and after freezing the beads, demonstrating its suitability for encapsulating and conserving cells. Hydrogel beads as functional system have been realized by generating protein-producing microcarriers with embedded eGFP-secreting insect cells

Cytocompatible, injectable, and electroconductive soft adhesives with hybrid covalent/noncovalent dynamic network

Synthetic conductive biopolymers have gained increasing interest in tissue engineering, as they can provide a chemically defined electroconductive and biomimetic microenvironment for cells. In addition to low cytotoxicity and high biocompatibility, injectability and adhesiveness are important for many biomedical applications but have proven to be very challenging. Recent results show that fascinating material properties can be realized with a bioinspired hybrid network, especially through the synergy between irreversible covalent crosslinking and reversible noncovalent self‐assembly. Herein, a polysaccharide‐based conductive hydrogel crosslinked through noncovalent and reversible covalent reactions is reported. The hybrid material exhibits rheological properties associated with dynamic networks such as self‐healing and stress relaxation. Moreover, through fine‐tuning the network dynamics by varying covalent/noncovalent crosslinking content and incorporating electroconductive polymers, the resulting materials exhibit electroconductivity and reliable adhesive strength, at a similar range to that of clinically used fibrin glue. The conductive soft adhesives exhibit high cytocompatibility in 2D/3D cell cultures and can promote myogenic differentiation of myoblast cells. The heparin‐containing electroconductive adhesive shows high biocompatibility in immunocompetent mice, both for topical application and as injectable materials. The materials could have utilities in many biomedical applications, especially in the area of cardiovascular diseases and wound dressing

A Self-Assembled Matrix System for Cell-Bioengineering Applications in Different Dimensions, Scales, and Geometries

Stem cell bioengineering and therapy require different model systems and materials in different stages of development. If a chemically defined biomatrix system can fulfill most tasks, it can minimize the discrepancy among various setups. By screening biomaterials synthesized through a coacervation-mediated self-assembling mechanism, a biomatrix system optimal for 2D human mesenchymal stromal cell (hMSC) culture and osteogenesis is identified. Its utility for hMSC bioengineering is further demonstrated in coating porous bioactive glass scaffolds and nanoparticle synthesis for esiRNA delivery to knock down the SOX-9 gene with high delivery efficiency. The self-assembled injectable system is further utilized for 3D cell culture, segregated co-culture of hMSC with human umbilical vein endothelial cells (HUVEC) as an angiogenesis model, and 3D bioprinting. Most interestingly, the coating of bioactive glass with the self-assembled biomatrix not only supports the proliferation and osteogenesis of hMSC in the 3D scaffold but also induces the amorphous bioactive glass (BG) scaffold surface to form new apatite crystals resembling bone-shaped plate structures. Thus, the self-assembled biomatrix system can be utilized in various dimensions, scales, and geometries for many different bioengineering applications

Endogenous glucocorticoids control host resistance to viral infection through the tissue-specific regulation of PD-1 expression on NK cells.

Controlling the balance between immunity and immunopathology is crucial for host resistance to pathogens. After infection, activation of the hypothalamic-pituitary-adrenal (HPA) axis leads to the production of glucocorticoids. However, the pleiotropic effects of these steroid hormones make it difficult to delineate their precise role(s) in vivo. Here we found that the regulation of natural killer (NK) cell function by the glucocorticoid receptor (GR) was required for host survival after infection with mouse cytomegalovirus (MCMV). Mechanistically, endogenous glucocorticoids produced shortly after infection induced selective and tissue-specific expression of the checkpoint receptor PD-1 on NK cells. This glucocorticoid-PD-1 pathway limited production of the cytokine IFN-gamma by spleen NK cells, which prevented immunopathology. Notably, this regulation did not compromise viral clearance. Thus, the fine tuning of NK cell functions by the HPA axis preserved tissue integrity without impairing pathogen elimination, which reveals a novel aspect of neuroimmune regulation