A quantitative metric for pattern fidelity of bioprinted cocultures.

This article describes a quantitative metric for coculture pattern fidelity and its use in the assessment of bioprinting systems. Increasingly, bioprinting is used to create in vitro cell and tissue models for the purpose of studying cell behavior and cell-cell interaction. To create meaningful models, a bioprinting system must be able to place cells in biologically relevant patterns with sufficient fidelity. A metric for assessing fidelity would be valuable for tuning experimental processes and parameters within a bioprinting system and for comparing performance between different systems. Toward this end, the "bioprinting fidelity index" (BFI), a metric which rates a bioprinted patterned coculture with a single number based on the proportions of correctly placed cells, is proposed. Additionally, a mathematical model of drop-on-demand printing is introduced, which predicts an upper bound on the BFI based on drop placement statistics. A proof-of-concept study was conducted in which patterned cocultures of D1 and 4T07 cells were produced in two different demonstration patterns. The BFI for the patterned cocultures was calculated and compared to the printing model fidelity prediction. The printing model successfully predicted the best BFI observed in the samples, and the BFI showed quantitatively that post-processing techniques negatively impacted the final fidelity of the samples. The BFI provides a principled method for comparing printing and post-processing methods.

Tannic acid cross-linked collagen scaffolds and their anti-cancer potential in a tissue engineered breast implant.

Tannic acid (TA) is a hydrolysable plant tannin, and it has been determined that TA functions as a collagen cross-linking agent through hydrogen-bonding mechanisms and hydrophobic effects. Since TA may have anti-tumor properties, it may be a viable cross-linking agent for collagen-based breast tissue scaffolds. The goal of this work was to determine if TA cross-linked scaffolds induce apoptotic processes in MCF-7 cancer cells, with minimal toxic effect on healthy D1 mesenchymal stem-like stromal cells. Cross-linked collagen scaffolds that were uniform, easily reproduced, easily characterized, and readily used in cell culture were manufactured. Thermal denaturation temperatures of the cross-linked scaffolds (68°C) were shown to be significantly higher when compared to those of uncross-linked scaffolds (55°C). Scanning electron microscopy images demonstrated the replacement of irregular collagen fibers with sheet-like structures upon cross-linking. The cross-linking solution concentration of TA that appears to be best for inducing apoptotic processes in MCF-7 cells, while minimizing toxic effect on D1 cells, is 1 mg/ml. At this concentration, the MCF-7 cell metabolic activity did not change over a 72-h period (i.e., proliferation was limited) while there was an increase in metabolic activity of D1 cells over the 72-h period. TA did appear to inhibit the production of lipid by D1 cells cultured in an adipogenic cocktail; in the future, the rate and duration of inhibition could be tailored to allow gradual bulking of the implant. The results suggest that the level of TA cross-linking can be modulated to provide optimal use in a tissue engineering composite.

Post-bioprinting processing methods to improve cell viability and pattern fidelity in heterogeneous tissue test systems.

Bioprinted tissue test systems show promise as a powerful tool for studying cell-cell interaction in heterogeneous, tissue-like co-culture. Several challenges were encountered while attempting to consistently fabricate samples with high viability and pattern fidelity. This paper evaluates four methods for processing samples after bioprinting but prior to adding media for incubation. These methods, composed of various combinations of three techniques meant to promote cell hydration, are evaluated with respect to sample viability and pattern preservation. In the best performing method, Hank's Balanced Salt Solution was applied immediately after fabrication and a collagen overlayer was applied one hour thereafter. The success of this method highlights the ability of the collagen substrate to absorb moisture, which promotes cell health without disturbing the cell's printed location. An addendum to the main study is an investigation of the limits of an HP26 print cartridge to deposit cells at a faster rate for the purpose of creating cell layers with densities that approach confluence.

The normal microenvironment directs mammary gland development.

Normal development of the mammary gland is a multidimensional process that is controlled in part by its mammary microenvironment. The mammary microenvironment is a defined location that encompasses mammary somatic stem cells, neighboring signaling cells, the basement membrane and extracellular matrix, mammary fibroblasts as well as the intercellular signals produced and received by these cells. These dynamic signals take numerous forms including growth factors, steroids, cell-cell or cell-basement membrane physical interactions. Cellular growth and differentiation of the mammary gland throughout the developmental stages are regulated by changes in these signals and interactions. The purpose of this review is to summarize current information and research regarding the role of the mammary microenvironment during normal glandular development.

Building off-the-shelf tissue-engineered composites.

Rapid advances in technology have created the realistic possibility of personalized medicine. In 2000, Time magazine listed tissue engineering as one of the 'hottest 10 career choices'. However, in the past decade, only a handful of tissue-engineered products were translated to the clinical market and none were financially viable. The reality of complex business planning and the high-investment, high-technology environment was not apparent, and the promise of tissue engineering was overstated. In the meantime, biologists were steadily applying three-dimensional benchtop tissue-culture systems for cellular research, but the systems were gelatinous and thus limited in their ability to facilitate the development of complex tissues. Now, the bioengineering literature has seen an emergence of literature describing biofabrication of tissues and organs. However, if one looks closely, again, the viable products appear distant. 'Rapid' prototyping to reproduce the intricate patterns of whole organs using large volumes of cellular components faces daunting challenges. Homogenous forms are being labelled 'tissues', but, in fact, do not represent the heterogeneous structure of the native biological system. In 2003, we disclosed the concept of combining rapid prototyping techniques with tissue engineering technologies to facilitate precision development of heterogeneous complex tissue-test systems, i.e. systems to be used for drug discovery and the study of cellular behaviour, biomedical devices and progression of disease. The focus of this paper is on the challenges we have faced since that time, moving this concept towards reality, using the case of breast tissue as an example.