Collagen-Chitosan inks for 3D-printing

ANA CAROLINA HEIDENREICH; PEREZ RECALDE MERCEDES; ANA GONZALEZ WUSENER; ELIDA HERMIDA

Mar del Plata

Simposio; XVI Simposio Latinoamericano de Polímeros; 2018

INTEMA, Universidad Nacional de Mar del Plata

INTRODUCTIONCollagen and chitosan are two biopolymers often usedin hydrogels for tissue engineering applications1,2. Bothhave complementary benefits: the non-rapidbiodegradability and good mechanical properties ofchitosan and the cellular adhesion of collagen. Blendsof them using different techniques were developed forcorneal applications, skeletal muscle tissue, islettransplantation, endothelial and skin tissue3?7,9 amongothers. But, as far as we know, these two biomaterialshave not been used for 3D-bioprinting yet. With theaim of developing chitosan-collagen inks, rheologicalstudies were performed. Dynamic viscosity of differentCollagen:Chitosan inks were measured and correlatedwith their capacity to be printed (printability: Pr).EXPERIMENTAL METHODSInks preparation: Type I rat tail collagen was extractedaccording to a protocol given in the literature8, leadingto a solution 0.72%w/v in acetic acid 1:1000. Lowmolecular weight chitosan (Sigma Aldrich®) wasdissolved in acetic acid to get concentrations between1 and 3%w/v. The inks were obtained mixing equalparts of collagen and chitosan solutions.Rheology: The flow tests were set at 25 ºC and undershear rate between 0,015-100 Hz, by duplicate, using arheometer TA Discovery-Hybrid R3.Printing and Measure of Printability: Extrusion of theinks were assessed using the 3D bioprinter 3Donor,made by Life SI, Arg. Printing fidelity, usually called,printability, was measured using square grids; its areasand perimeters were accurately measured by opticalmicroscopy and normalized by the square circularity.The closer to 1 the better the capacity of the gel toproduce a square, according to the model.RESULTS AND DISCUSSIONFig.1 shows viscosity vs. shear rate curves for three inkswith different chitosan concentrations and the twocomponents alone. Vertical lines indicate three shearrates applied to the inks while being printed at threedifferent printing conditions (1, 2 and 3).Table 1. Printability for Col 0.36% Chi 1% at the indicated flow ratesFlow rate [ul/s] Shear Rate [1/s] Pr0.19±0.09 27.7±12.67 0.97±0.020.42±0.04 60.61±6.6 0.95±0.050.35±0.07 50.16±10.16 0.97±0.03Collagen exhibits a thixotropic behavior within theshear rate range tested while the chitosan solutionbehaves as a Newtonian fluid. Also the viscosity of thethree blends decreases as the shear rate increases;furthermore, the solution becomes more viscous as thechitosan concentration increases.Figure 1. Apparent viscosity in a shear rate sweep for collagen 0.36% w/v and different chitosan concentrations. Shear rates associatedto the 3 printing conditions are also indicated.The ink Col 0.36% Chi 1% has a suitable viscosity forprinting purposes and is also compatible with cellularviability4,5,7; therefore, its printability was measured atthe 3 different printing conditions, as shown in Table 1.Pr values are higher than 0.85 in all cases so, the flowrate can be the highest to reduce printing time.CONCLUSIONHomogenous blends of chitosan and collagenhydrogels could be printed with acceptable shapefidelity and adequate printing process. The passagefrom hydrogel precursors, present in inks, to hydrogelafter printing is an important challenge since bothpolymers have to be crosslinked to get the desiredsubstrate morphology. Our preliminary results showthat pH neutralisation and crosslinking activatorsallows to achieve a desired substrate.