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Silva, et al.
which cells are exposed during extrusion. Moreover, an outermost tunica adventitia layer made of
the viability of constructs extruded through nozzle surrounding fibroblast and collagen; an ideal
2 is only significantly different from those extruded tissue-engineered blood vessel should consist of
through nozzle 1 when the inlet pressure is high (40 those three layers, this coaxial nozzle can reduce
kPa), which indicates that layer thickness affects the wall thickness of this kind of tissues obtained
viability but to a lesser extent than the extrusion in other investigations close mimicking the
[31]
pressure. However, setting an 80 % cell viability wall thickness of small arteries and veins in a real
threshold, these results suggest that all three coaxial human body.
nozzles could be used for the bioprinting of hollow
tubular structures by applying an inlet extrusion 4 Conclusions
pressure below 34 kPa. In silico simulations were performed for studying
The main goal of this research is to optimize the the pressure distribution exerted on cells during the
development of triple-layered coaxial nozzles for bioprinting process, as well as the outlet velocity at
facilitating the fabrication of biomimetic tissues the tip of three different flow channels. Our results
and organ-like constructs for tissue engineering confirmed those of previously reported studies and
and regenerative medicine applications. demonstrate the usefulness of in silico experiments in
Furthermore, the enhanced development of triple- helping to optimize in vitro experiments. The results
layered coaxial nozzles can help to solve the issues can be useful in guiding the future development of
regarding vascularization, which remain as one of improved multi-layered coaxial nozzles.
the key bottlenecks of the field . The next step Three triple-layered coaxial nozzles with
[30]
in our research is to apply these same concepts different Gauges were first studied in silico
in the development of a four-layered coaxial regarding varying pressures and then successfully
nozzle, whose advantages compared to the coaxial designed for the fabrication of single-layered
nozzle presented here will be remarked. With hollow tubular structures of different dimensions.
one more layer, it will be easier to close mimic All nozzles displayed adequate bioprinting
the complexity of the vascular network, taking
into account that the current state of the art tissue conditions to guarantee cell viability above 80 %
conventional culture technique is limited to only in alginate-based hydrogels when extrusion
triple co-culture (3 types of cells). Advancements pressure was kept below 34 kPa, meaning they
are all suitable for bioprinting with bioinks with
in tissue culture techniques are necessary to similar composition or rheological properties to
address the bottleneck of maturing bioprinted the one studied in this project. Moreover, herein
multi-cellular 3D tissue constructs into functional reported coaxial nozzles to allow the formation
tissues with a wide range of cells and biomaterials of perfusable cannular structures with dimensions
with differentiated layer co-culture within one that fall within the range of human arteries, which
single bioprinted construct. means they could be further exploited for the
The novelty of this work is adding value in the
research field of bioprinting with a triple-layered fabrication of multicellular vascular networks
coaxial nozzle development that has the potential and vessel-like constructs with applications on
to closely mimic the complexity of vascular tissue engineering. Our work paves the way for
networks found in the native human body in the rational development of coaxial nozzles useful
terms of histological and morphological of this for bioprinting multi-layered vascular channels
vascular constructs, as well this development can or vessel-like constructs that truly resemble those
replicate the wall thickness of a native blood vessel found in native organs and organisms.
that generally comprised three layers with the Acknowledgments
innermost tunica intima layer made by continuous
endothelium cells followed by the middle tunica We would like to acknowledge the financial support
layer made of elastic, smooth muscle cells and received from the Colombian Administrative

International Journal of Bioprinting (2020)–Volume 6, Issue 4 103