Abstract
Gellan gum (GG) spongy-like hydrogels have been explored for different tissue engineering (TE) applications owing to their highly attractive hydrogel-like features, and improved mechanical resilience and cell performance. Although the whole process for the preparation of these materials is well-defined, we hypothesized that variations occurring during the freezing step lead to batch-to-batch discrepancies. Aiming to address this issue, two freezing devices were tested, to prepare GG spongy-like hydrogels in a more reproducible way. The cooling and freezing rates, the nucleation time and temperature, and the end freezing time were determined at different freezing temperatures (−20, −80, and −210 °C). The efficacy of the devices was assessed by analyzing the physicochemical, mechanical, and biological properties of different formulations. The cooling rate and freezing rate varied between 0.1 and 128 °C/min, depending on the temperature used and the device. The properties of spongy-like hydrogels prepared with the tested devices showed lower standard deviation in comparison to those prepared with the standard process, due to the slower freezing rate of the hydrogels. However, with this method, mean pore size was significantly lower than that with the standard method. Cell entrapment, adhesion, and viability were not affected as demonstrated with human dermal fibroblasts. This work confirmed that batch-to-batch variations are mostly due to the freezing step and that the tested devices allow fine tuning of the scaffolds’ structure and properties.
Highlights
Porous biomaterials have been extensively used in tissue engineering, holding a great promise for the regeneration and repair of damaged tissues as providers of a three-dimensional structure (3D)for the adhesion, growth, migration, and proliferation of cells [1]
The thermal profile is divided in two main phases, the cooling phase occurring until the nucleation temperature is reached, and the freezing phase which follows it, starting when the nucleation temperature is reached (Figure 2B)
The cooling rate was calculated from the linear region of the cooling phase, while the freezing rate was obtained from the linear region of the profile corresponding to the beginning of the freezing slope up to the end freezing temperature was reached (Figure 2B)
Summary
Porous biomaterials have been extensively used in tissue engineering, holding a great promise for the regeneration and repair of damaged tissues as providers of a three-dimensional structure (3D)for the adhesion, growth, migration, and proliferation of cells [1]. The architecture and isotropy of scaffolds influence overall cell behavior, as cells randomly distribute and project filopodia in scaffolds with random pores but elongate along the pores in radially or axially aligned scaffolds [5]. Host response and neotissue formation have been found to be influenced by scaffolds’ microstructure [6,7,8], the size [9] and direction [10] of the pores. Higher porosity is associated with a higher area to support cell adhesion in the first stage, and with faster biodegradability, providing additional space for tissue ingrowth [9]. A precise control of the pore size, architecture, and interconnectivity of scaffolds is needed
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