Gellan gum (GG) spongy-like hydrogels (SLH) have been widely explored for several tissue engineering
applications due to their attractive properties such as mechanical stability and cell-adhesion features. Those
are prepared from GG hydrogels after consecutive steps of freezing and freeze-drying to obtain dried polymeric networks (DPN) that after re-hydration give shape to spongy-like hydrogels1, 2. Different factors, such as, the freezing temperature, freezing rate and freezing time, have shown to affect the microstructural features of DPN and, consequently, the overall features of SLH1,2,3. Recurrent batch-to-batch variability suggested that other factors might affect spongy-like hydrogels preparation. Thus, in this work we aimed to further understand the influence of parameters, such as the diameter of the hydrogel (varying from 6 mm to 85.31 mm), the freezing temperature (-20ºC and -40ºC) and the freezing rate (0.1ºC/min and 1ºC/min), by using a freeze-dryer with a controlled and customized cycle of thermal treatment and drying steps. To attain this aim, GG DPN (0.75% and 1.5% (w/v)) were prepared and their microstructure was analyzed by micro-computed tomography and confirmed by scanning electron microscopy. As expected, there was a decrease of mean pore size with the increase of polymer amount, e.g. 138.20±29.52 μm (0.75% GG) vs 120.99±15.44 μm (1.5% GG) [0.1ºC/min, -20ºC, 6mm]. In respect to the influence of the diameter of the hydrogel, larger materials resulted in lower mean pore size, e.g. 138.20±29.52 μm (6 mm) vs 61.54±7.43 (22 mm) [0.1ºC/min, -20ºC, 0.75%]. Contrarily, the freezing temperature did not seem to affect the microstructure of the DPB, e.g. 120.99±15.44μm (-20ºC) vs 124.16±5.23μm (-40ºC) [0.1ºC/min, 1.5%, 6mm]. Disappointingly, conclusions regarding the effect of the freezing rate were not attained. Inconsistent data indicated that the equipment was not capable of reaching high freezing rates (1ºC/min). Nonetheless, this system could be of use to further tailor DPN microstructure according to specific applications for tissue engineering and to further standardize the processing of spongy-like hydrogels.
ACKNOWLEDGEMENTS: ERC for funding ECM-INK project (ERC-2016-CoG-726061) and FCT for LdS grant
(2020.01541.CEECIND/CP1600/CT0024).
[1] da Silva, L. P. et al. Acta Biomater.2014; 10, 4787–4797.
[2] da Silva L.P. et al. 2014. Horizon 2020 WO/2014/167513.
[3] Moreira, H. R. et al. Polymers. 2020; 12, 1–15
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