3D printing structures with a cell bearing gelatin bioink and transglutaminase crosslinking.
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1
Nanyang Technological University, School of Materials Science and Engineering, Singapore
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2
Nanyang Technological University, School of Biological Sciences, Singapore
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3
The Technion – Israel Institute of Technology, Faculty of Biotechnology and Food Engineering, Israel
Introduction: Free form fabrication (FFF), also referred to as additive manufacturing and 3D printing, offers architectural control for the production of biomaterial structures. Deposition control allowed by FFF is considerably better than alternative techniques such as electrospinning. Most FFF methods are not cytocompatible due to the heat or solvent required for polymer delivery, hence cell seeding can only occur post fabrication. Here we employ FFF to bioprint a cell bearing bioink to create cellurized structures with both human umbilical vein endothelial cells (HUVEC) and HEK293 cells. A gelatin hydrogel based bioink was used due to the proteins cytocompatible nature. Gelatin hydrogels however are soluble hence not stable without crosslinking. In this study gelatin cross linking was performed by the activity of microbial transglutaminase (Tgase).
Methods: Bioprinting was performed using back pressure assisted syringe mechanism on a computer controlled robotic dispensing system with XYZ table. The bioprinting was optimized for gelatin concentration, viscosity, temperature, printing speed and back pressure to allow effective bioprinting and a sufficient level of cell viability.
HEK293 cells and HUVEC cell were assessed for their ability to endure bioprinting, crosslinking reaction and culture within a bioprinted hydrogel structure
Results: It was found that the lower the gelatin concentration, the better the cell spreading and proliferation. It was found 3% gelatin with 2%PEO additive produced the most precise hydrogel trace and best cell spreading in 2D patterning. However, bioinks with less than 5% gelatin concentration could not be built up in 3D. 5% gelatin proved to allow building of 3D hydrogel structures with best cell viability. Increased gelatin concentration in the bioink reduced cell viability due to detrimental shear stress. Tgase had to be included in the bioink during dispensing for effective gelatin crosslinking, thus giving a limiting time window for the bioprinting to occur before the crosslinked gelatin clogs the printing nozzle. However the crosslinking time and printing window could be extended by reducing the TGase concentration.
Conclusion: 3D structures were printed in 5% gelatin bioink crosslinked with Tgase. Living HUVEC and HEK293 cells were included in the bioink before dispensing and fabrication of a cell bearing structure. The crosslinked gelatin structure remained stable for several days and viable bioprinted cells were observed to spread and proliferate within the hydrogel.
In summary, gelatin crosslinked by Tgase forms an effective bioink that support the deposition of cells directly into the emerging scaffold.
Singapore National Research Foundation
References:
[1] Irvine SA, Agrawal A, Lee BH, Chua HY, Low KY, Lau BC, Machluf M, Venkatraman S. Printing cell-laden gelatin constructs by free-form fabrication and enzymatic protein crosslinking. Biomed Microdevices. 2015 Feb;17(1):16. doi: 10.1007/s10544-014-9915-8.
Keywords:
Hydrogel,
Bioprinting,
Rapid prototyping,
3D scaffold
Conference:
10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016.
Presentation Type:
General Session Oral
Topic:
Biomaterials in printing
Citation:
Irvine
S,
Agrawal
A,
Lee
B,
Chua
H,
Low
K,
Lau
B,
Machluff
M and
Venkatraman
S
(2016). 3D printing structures with a cell bearing gelatin bioink and transglutaminase crosslinking..
Front. Bioeng. Biotechnol.
Conference Abstract:
10th World Biomaterials Congress.
doi: 10.3389/conf.FBIOE.2016.01.01924
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Received:
27 Mar 2016;
Published Online:
30 Mar 2016.
*
Correspondence:
Dr. Scott Irvine, Nanyang Technological University, School of Materials Science and Engineering, Singapore, Singapore, Email1
Dr. Bae Hoon Lee, Nanyang Technological University, School of Materials Science and Engineering, Singapore, Singapore, LEEBH@ntu.edu.sg
Dr. Hui Yee Chua, Nanyang Technological University, School of Materials Science and Engineering, Singapore, Singapore, CHUA0660@ntu.edu.sg
Dr. Boon Chong Lau, Nanyang Technological University, School of Materials Science and Engineering, Singapore, Singapore, BCLAU1@e.ntu.edu.sg
Dr. Subbu Venkatraman, Nanyang Technological University, School of Materials Science and Engineering, Singapore, Singapore, subbu@Nus.edu.sg