Synthetic 3D Extracellular Matrices (ECM) find application in cell studies, regenerative medicine and drug discovery. While cells cultured in a monolayer may exhibit unnatural behavior and develop very different phenotypes and genotypes than in vivo, great efforts in materials chemistry have been devoted to reproduce in vitro the in vivo cell mi-croenvironments, which requires a fine tuning of the biochemical and structural actors varying in a spatiotemporal fashion to regulate the cell fate. This review will present the fundamentals of ECM, cover the chemical and structural features of the scaffolds used to generate ECM mimics, discuss the nature of the signaling biomolecules required and exploited to generate bioresponsive cell microenvironments able to induce a specific cell fate, and highlights the syn-thetic strategies involved to create functional 3D ECM mimics.

The development of 3D printable hydrogels based on the crosslinking between chitosan and gelatin is proposed. Chitosan and gelatin were both functionalized with methyl furan groups. Chemical modification was performed by reductive amination with methyl furfural involving the lysine residues of gelatin and the amino groups of chitosan to generate hydrogels with tailored properties. The methyl furan residues present in both polymers were exploited for efficient crosslinking via Diels-Alder ligation with PEG-Star-maleimide under cell-compatible conditions. The obtained chitosan-gelatin hybrid was employed to formulate hydrogels and 3D printable biopolymers and its processability and biocompatibility were preliminarily investigated.

A protocol based on the combination of different analytical methodologies is proposed to standardize the experimental conditions for reproducible formulations of hybrid hydrogels. The final hybrid material, based on the combination of gelatin and chitosan functionalized with methylfuran and cross-linked with 4-arm-PEG-maleimide, is able to mimic role, dynamism, and structural complexity of the extracellular matrix. Physical–chemical properties of starting polymers and finals constructs were characterized exploiting the combination of HP-SEC-TDA, UV, FT-IR, NMR, and TGA.