Encapsulation of adipose stromal vascular fraction cells in alginate hydrogel spheroids using a direct-write three-dimensional printing system.

The study of tissue function in vitro has been aided by the development of three-dimensional culture systems that more accurately duplicate the complex cell components of tissues and organs. Bioprinting of cells provides a rapid tissue fabrication technique that can be used to evaluate normal and pathologic conditions in vitro as well as to construct complex three-dimensional tissue structures for implantation in regenerative medicine therapies. Studies were performed using a direct write three-dimensional bioprinting system to fabricate adipose-derived stromal vascular fraction cell spheroids. Human fat-derived stromal vascular fraction cells were mixed in 1.5% (w/v) alginate solutions, and fabrication conditions were varied to produce an array of spheroids. The spheroids were placed in spinner culture, and spheroid integrity and encapsulated cell viability were assessed for 16 days. Results establish the ability to tightly control adipose SVF spheroids in the range of 800-1500 µm. Fabrication conditions were used to control spheroid size, and the results illustrate the ability to construct spheroids of precise size and shape. The adipose SVF cell population remains viable and the spheroid integrity was maintained for 16 days in suspension culture. The direct-write printing of adipose stromal vascular fraction cell containing spheroids provides a rapid fabrication technology to support in vitro microphysiologic system studies.

Determinants of microvascular network topologies in implanted neovasculatures.

OBJECTIVE: During neovascularization, the end result is a new functional microcirculation composed of a network of mature microvessels with specific topologies. Although much is known concerning the mechanisms underlying the initiation of angiogenesis, it remains unclear how the final architecture of microcirculatory beds is regulated. To begin to address this, we determined the impact of angiogenic neovessel prepatterning on the final microvascular network topology using a model of implant neovascularization. METHODS AND RESULTS: We used 3D direct-write bioprinting or physical constraints in a manner permitting postangiogenesis vascular remodeling and adaptation to pattern angiogenic microvascular precursors (neovessels formed from isolated microvessel segments) in 3D collagen gels before implantation and subsequent network formation. Neovasculatures prepatterned into parallel arrays formed functional networks after 4 weeks postimplantation but lost the prepatterned architecture. However, maintenance of uniaxial physical constraints during postangiogenesis remodeling of the implanted neovasculatures produced networks with aligned microvessels, as well as an altered proportional distribution of arterioles, capillaries, and venules. CONCLUSIONS: Here we show that network topology resulting from implanted microvessel precursors is independent from prepatterning of precursors but can be influenced by a patterning stimulus involving tissue deformation during postangiogenesis remodeling and maturation.

Engineering a titanium and polycaprolactone construct for a biocompatible interface between the body and artificial limb.

Intraosseous transcutaneous amputation prostheses may be able to overcome the problems that stem from the nonuniform distribution of pressure seen in the conventional stump-socket prosthetic replacement devices. Transcutaneous devices have had limited success in amputees. By optimizing the attachment of the skin to the prosthetic, intraosseous transcutaneous amputation prostheses may become clinically viable options. This report details studies evaluating the development of a modified titanium construct with a specially machined surface to increase the adherence of tissue as well as scaffold. A computer-aided biology tool was used to fabricate polycaprolactone (PCL) scaffolds with a specific three-dimensional architecture. To extrude the PCL, it was dissolved in acetic acid to produce a 70% PCL liquid. A scaffold with a porosity of >50% was fabricated to have a tensile strength similar to skin. The presence of a specially machined surface greatly increased the adhesion of the PCL scaffold to the titanium constructs. When the 70% PCL was properly neutralized by heating at 55 degrees C and washing in 90% ethanol (EtOH), there was only a decrease (10%) in the viability of cells seeded onto the PCL constructs when compared with the cells in culture. The antibacterial properties of titanium dioxide anatase, silver nanoparticles, and chlorhexidine diacetate mixed in either type I collagen or hyaluronic acid (HA) were assessed. The addition of 1% (w/w) chlorhexidine diacetate in HA resulted in a 71% decrease in bacteria seen in nontreated HA. These results show promise in developing a novel engineered titanium and PCL construct that promotes effective adhesion between the titanium-skin interface.

Tissue engineering by self-assembly of cells printed into topologically defined structures.

Understanding the principles of biological self-assembly is indispensable for developing efficient strategies to build living tissues and organs. We exploit the self-organizing capacity of cells and tissues to construct functional living structures of prescribed shape. In our technology, multicellular spheroids (bio-ink particles) are placed into biocompatible environment (bio-paper) by the use of a three-dimensional delivery device (bio-printer). Our approach mimics early morphogenesis and is based on the realization that the genetic control of developmental patterning through self-assembly involves physical mechanisms. Three-dimensional tissue structures are formed through the postprinting fusion of the bio-ink particles, in analogy with early structure-forming processes in the embryo that utilize the apparent liquid-like behavior of tissues composed of motile and adhesive cells. We modeled the process of self-assembly by fusion of bio-ink particles, and employed this novel technology to print extended cellular structures of various shapes. Functionality was tested on cardiac constructs built from embryonic cardiac and endothelial cells. The postprinting self-assembly of bio-ink particles resulted in synchronously beating solid tissue blocks, showing signs of early vascularization, with the endothelial cells organized into vessel-like conduits.