Alexandra L. Rutz

Three-Dimensional Printing of High-Content Graphene Scaffolds for Electronic and Biomedical Applications

The exceptional properties of graphene enable applications in electronics, optoelectronics, energy storage, and structural composites. Here we demonstrate a 3D printable graphene (3DG) composite consisting of majority graphene and minority polylactide-co-glycolide, a biocompatible elastomer, 3D-printed from a liquid ink. This ink can be utilized under ambient conditions via extrusion-based 3D printing to create graphene structures with features as small as 100 μm composed of as few as two layers (<300 μm thick object) or many hundreds of layers (>10 cm thick object). The resulting 3DG material is mechanically robust and flexible while retaining electrical conductivities greater than 800 S/m, an order of magnitude increase over previously reported 3D-printed carbon materials. In vitro experiments in simple growth medium, in the absence of neurogenic stimuli, reveal that 3DG supports human mesenchymal stem cell (hMSC) adhesion, viability, proliferation, and neurogenic differentiation with significant upregulation of glial and neuronal genes. This coincides with hMSCs adopting highly elongated morphologies with features similar to axons and presynaptic terminals. In vivo experiments indicate that 3DG has promising biocompatibility over the course of at least 30 days. Surgical tests using a human cadaver nerve model also illustrate that 3DG has exceptional handling characteristics and can be intraoperatively manipulated and applied to fine surgical procedures. With this unique set of properties, combined with ease of fabrication, 3DG could be applied toward the design and fabrication of a wide range of functional electronic, biological, and bioelectronic medical and nonmedical devices.

A Multimaterial Bioink Method for 3D Printing Tunable, Cell‐Compatible Hydrogels

A multimaterial bio-ink method using polyethylene glycol crosslinking is presented for expanding the biomaterial palette required for 3D bioprinting of more mimetic and customizable tissue and organ constructs. Lightly crosslinked, soft hydrogels are produced from precursor solutions of various materials and 3D printed. Rheological and biological characterizations are presented, and the promise of this new bio-ink synthesis strategy is discussed.

A bioprosthetic ovary created using 3D printed microporous scaffolds restores ovarian function in sterilized mice

Emerging additive manufacturing techniques enable investigation of the effects of pore geometry on cell behavior and function. Here, we 3D print microporous hydrogel scaffolds to test how varying pore geometry, accomplished by manipulating the advancing angle between printed layers, affects the survival of ovarian follicles. 30° and 60° scaffolds provide corners that surround follicles on multiple sides while 90° scaffolds have an open porosity that limits follicle–scaffold interaction. As the amount of scaffold interaction increases, follicle spreading is limited and survival increases. Follicle-seeded scaffolds become highly vascularized and ovarian function is fully restored when implanted in surgically sterilized mice. Moreover, pups are born through natural mating and thrive through maternal lactation. These findings present an in vivo functional ovarian implant designed with 3D printing, and indicate that scaffold pore architecture is a critical variable in additively manufactured scaffold design for functional tissue engineering.

Hyperelastic “bone”: A highly versatile, growth factor–free, osteoregenerative, scalable, and surgically friendly biomaterial

A new, mechanically elastic biomaterial can be custom 3D-printed, is surgically friendly, and promotes robust bone regeneration. Building better bones What if we could create custom bone implants that would trigger their own replacement with real bone? Jakus and colleagues have done just this with a promising biomaterial that can be 3D-printed into many shapes and easily deployed in the operating room. Made mainly of hydroxyapatite and either polycaprolactone or poly(lactic-co-glycolic acid), this “hyperelastic bone” can be 3D-printed at up to 275 cm3/hour, the authors report. It also promoted bone growth in vitro, in mice and rats, and in a case study of skull repair in a rhesus macaque. Its effectiveness, fast, easy synthesis, and ease of use in surgery set it apart from many of the materials now available for bone repair. Despite substantial attention given to the development of osteoregenerative biomaterials, severe deficiencies remain in current products. These limitations include an inability to adequately, rapidly, and reproducibly regenerate new bone; high costs and limited manufacturing capacity; and lack of surgical ease of handling. To address these shortcomings, we generated a new, synthetic osteoregenerative biomaterial, hyperelastic “bone” (HB). HB, which is composed of 90 weight % (wt %) hydroxyapatite and 10 wt % polycaprolactone or poly(lactic-co-glycolic acid), could be rapidly three-dimensionally (3D) printed (up to 275 cm3/hour) from room temperature extruded liquid inks. The resulting 3D-printed HB exhibited elastic mechanical properties (~32 to 67% strain to failure, ~4 to 11 MPa elastic modulus), was highly absorbent (50% material porosity), supported cell viability and proliferation, and induced osteogenic differentiation of bone marrow–derived human mesenchymal stem cells cultured in vitro over 4 weeks without any osteo-inducing factors in the medium. We evaluated HB in vivo in a mouse subcutaneous implant model for material biocompatibility (7 and 35 days), in a rat posterolateral spinal fusion model for new bone formation (8 weeks), and in a large, non-human primate calvarial defect case study (4 weeks). HB did not elicit a negative immune response, became vascularized, quickly integrated with surrounding tissues, and rapidly ossified and supported new bone growth without the need for added biological factors.

Advancing the field of 3D biomaterial printing.

3D biomaterial printing has emerged as a potentially revolutionary technology, promising to transform both research and medical therapeutics. Although there has been recent progress in the field, on-demand fabrication of functional and transplantable tissues and organs is still a distant reality. To advance to this point, there are two major technical challenges that must be overcome. The first is expanding upon the limited variety of available 3D printable biomaterials (biomaterial inks), which currently do not adequately represent the physical, chemical, and biological complexity and diversity of tissues and organs within the human body. Newly developed biomaterial inks and the resulting 3D printed constructs must meet numerous interdependent requirements, including those that lead to optimal printing, structural, and biological outcomes. The second challenge is developing and implementing comprehensive biomaterial ink and printed structure characterization combined with in vitro and in vivo tissue- and organ-specific evaluation. This perspective outlines considerations for addressing these technical hurdles that, once overcome, will facilitate rapid advancement of 3D biomaterial printing as an indispensable tool for both investigating complex tissue and organ morphogenesis and for developing functional devices for a variety of diagnostic and regenerative medicine applications.

Protein-Based Hydrogels

Protein-based hydrogels are composed of isolated or enriched proteins from natural extracellular matrix. Inherent and controllable bioactivity makes these hydrogels promising candidates as smart biomaterials for drug delivery, tissue engineering and regenerative medicine, and other applications. Desirable characteristics for these applications include natural cell binding, cell degradable, and growth factor-binding sequences. This chapter covers the unique properties of a variety of proteins (collagen, gelatin, fibrin, silk, elastin, keratin, and decellularized, tissue-specific extracellular matrix) as well as hydrogel synthesis, fabrication, modification, and established applications. Conditions of solubility and the mechanism of the sol–gel transition are discussed. Since each protein presented undergoes self-assembly to form a gel network, gelation parameters that affect this assembly and subsequently the gel ultrastructure are specifically presented. Emerging applications and technologies for protein-based hydrogels are also briefly mentioned.