Cochlear organoids reveal transcriptional programs of postnatal hair cell differentiation from supporting cells.

We explore the changes in chromatin accessibility and transcriptional programs for cochlear hair cell differentiation from postmitotic supporting cells using organoids from postnatal cochlea. The organoids contain cells with transcriptional signatures of differentiating vestibular and cochlear hair cells. Construction of trajectories identifies Lgr5+ cells as progenitors for hair cells, and the genomic data reveal gene regulatory networks leading to hair cells. We validate these networks, demonstrating dynamic changes both in expression and predicted binding sites of transcription factors (TFs) during organoid differentiation. We identify known regulators of hair cell development, Atoh1, Pou4f3, and Gfi1, and the analysis predicts the regulatory factors Tcf4, an E-protein and heterodimerization partner of Atoh1, and Ddit3, a CCAAT/enhancer-binding protein (C/EBP) that represses Hes1 and activates transcription of Wnt-signaling-related genes. Deciphering the signals for hair cell regeneration from mammalian cochlear supporting cells reveals candidates for hair cell (HC) regeneration, which is limited in the adult.

Highly tunable elastomeric silk biomaterials.

Elastomeric, fully degradable and biocompatible biomaterials are rare, with current options presenting significant limitations in terms of ease of functionalization and tunable mechanical and degradation properties. We report a new method for covalently crosslinking tyrosine residues in silk proteins, via horseradish peroxidase and hydrogen peroxide, to generate highly elastic hydrogels with tunable properties. The tunable mechanical properties, gelation kinetics and swelling properties of these new protein polymers, in addition to their ability to withstand shear strains on the order of 100%, compressive strains greater than 70% and display stiffness between 200 - 10,000 Pa, covering a significant portion of the properties of native soft tissues. Molecular weight and solvent composition allowed control of material mechanical properties over several orders of magnitude while maintaining high resilience and resistance to fatigue. Encapsulation of human bone marrow derived mesenchymal stem cells (hMSC) showed long term survival and exhibited cell-matrix interactions reflective of both silk concentration and gelation conditions. Further biocompatibility of these materials were demonstrated with in vivo evaluation. These new protein-based elastomeric and degradable hydrogels represent an exciting new biomaterials option, with a unique combination of properties, for tissue engineering and regenerative medicine.

Enzymatically cross-linked alginic-hyaluronic acid composite hydrogels as cell delivery vehicles.

An injectable composite gel was developed from alginic and hyaluronic acid. The enzymatically cross-linked injectable gels were prepared via the oxidative coupling of tyramine modified sodium algiante and sodium hyaluronate in the presence of horse radish peroxidase (HRP) and hydrogen peroxide (H2O2). The composite gels were prepared by mixing equal parts of the two tyraminated polymer solutions in 10U HRP and treating with 1.0% H2O2. The properties of the alginate gels were significantly affected by the addition of hyaluronic acid. The percentage water absorption and storage modulus of the composite gels were found to be lower than the alginate gels. The alginate and composite gels showed lower protein release compared to hyaluronate gels in the absence of hyaluronidase. Even hyaluronate gels showed only approximately 10% protein release after 14 days incubation in phosphate buffer solution. ATDC-5 cells encapsulated in the injectable gels showed high cell viability. The composite gels showed the presence of enlarged spherical cells with significantly higher metabolic activity compared to cells in hyaluronic and alginic acid gels. The results suggest the potential of the composite approach to develop covalently cross-linked hydrogels with tuneable physical, mechanical, and biological properties.