ORGAN PRINTING - 3D Bioprinting and Nanotechnology in Tissue Engineering and Regenerative Medicine - page 329

hydrostatic pressure increase chondrogenesis ( Sumanasinghe et al., 2006 ; Sumanasinghe et al., 2006 ;
Sen et al., 2008 ; Arnsdorf et al., 2009 ; Li et al., 2004 ; Riddle et al., 2006 ; Haudenschild et al., 2008 ;
Wagner et al., 2008 ; Castillo and Jacobs, 2010 ). A fundamental topic requiring study is the effect
of microstructural cues from engineered scaffolds, for instance, the dimensional scale and geometry of
the 3D polymer substrates. A number of micro- and nanofabrication-based cell platforms, particularly
with spatially controlled features, have been advanced for investigating fundamental questions in cell
behavior and tissue development ( Whitesides et al., 2001 ; Théry, 2010 ; Chen et al., 1997 ; Quake and
Scherer, 2000 ; Huang et al., 2008 ). While these micro- and nanoscale manufacturing techniques have
yielded insight into structural cues on stem cell differentiation, scalable bioadditive manufacturing pro-
cesses with embedded microstructural cues for creating functional tissue to meet critical organ deficit
size demands have remained unexplored. It has been shown that, given the same material composition,
electrospun nanofiber ( < 1 m m) substrates seeded with MSCs differentiate into bone-forming cells,
while solid freeform fabricated ( > 100 m m) 3D substrates similarly seeded with MSCs yield undif-
ferentiated cells ( Farooque et al., 2014 ; Kumar et al., 2011 ; Kumar et al., 2012 ; Huang et al., 2013 ).
Therefore, the author's working hypothesis is that there exists a threshold dimensional scale on the
order of the single cell of presenting microstructural cues that will induce MSC differentiation into
bone-forming cells. There is a need, therefore, to model and develop enabling bioadditive manufactur-
ing processes to fill in the critical dimensional gap between polymer electrospinning and conventional
RP technologies. As a global measure for the process outcome of presenting stem cells with a 3D
microstructural niche for differentiation, one possibility is advancing subcellular cell shape dynamics
as a quantitative metric. Although there are published reports on single-cell imaging with structural
classification of 3D cell shape dimensionality, current limitations of cell shape metrology that need
to be addressed include single time point imaging and accounting for the number and distribution of
subcellular structural elements ( Farooque et al., 2014 ; Kumar et al., 2011 ).
15.1.3 MICRO-ORGAN PRINTING AS PHYSIOLOGICAL AND DISEASE PLATFORMS
15.1.3.1 Microextusion-based Printed Liver Micro-organ on a Chip
A near-future application of organ printing techniques is the creation of cell-based physiological
and disease platforms, which we will refer to in this chapter as micro-organ on a chip. Specifically,
liver-on-a-chip technology has generated significant research interest in in vitro drug metabolism and
toxicity studies. However, to date, conventional 2D monolayer cultures of primary hepatocytes cause
hepatocytes to lose their morphology and liver-specific functions, including the activity of a group of
metabolizing enzymes located in the endoplasmic reticulum called cytochrome P-450 oxidases ( Bur-
khardt et al., 2013 ). The authors have addressed this obstacle with 3D culture models with layered
microextrusion-based printing approaches. Others have also embedded matrices made from natural
or synthetic hydrogels and scaffolds constructed from classical biocompatible soft polymers, which
enable hepatocytes to express their phenotype in a 3D microenvironment toward increasing their vi-
ability and maintaining their functionality over longer culture time periods ( Chang et al., 2008a ; Chang
et al., 2008b ; Miranda et al., 2010 ). It has also been shown that hepatocyte cell physiological behavior
is favored under constant perfusion conditions ( Goral and Yuen, 2012 ). In this regard, microfluidics
has been adapted through layer-by-layer biofabrication approaches, thus enabling the relatively new
“organ-on-a-chip” technology ( Lee et al., 2013 ). This development has led to the creation of micro-
analytical micro-organ devices, whose 3D, dynamic nature distinguishes them from the conventional