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3D Bioprinting of Amniotic Membrane-Based Nanocomposite for Tissue Engineering Applications: Evaluation of Rheological, Mechanical and Biological Properties

Kafili, Golara | 2023

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  1. Type of Document: Ph.D. Dissertation
  2. Language: Farsi
  3. Document No: 56520 (48)
  4. University: Sharif University of Technology
  5. Department: Institute for Nanoscience and Nanotechnology
  6. Advisor(s): Simchi, Abdolreza; Tamjid, Elnaz; Niknejad, Hassan
  7. Abstract:
  8. 3D bioprinting is an additive manufacturing method that facilitates the deposition of the desired cells and biomaterials at any pre-defined location. This technique also enables control over the internal structure and external dimensions of printed constructs. Among various biomaterials used as bioinks, the bioinks derived from decellularized extracellular matrixes (dECMs) have attracted significant attention due to their bioactivity and being a rich source of biochemical cues. Here in this study, the decellularized amnion membrane (dAM) has been selected as the main component of the bioink formulation because of its biocompatibility, low immunogenicity, antibacterial property, abundance, and ease of access. Despite of desired biological properties of the dAM-derived hydrogel, similar to other dECM-derived biomaterias, it lacks adequate mechanical properties and exhibits poor printability. Therefore, in the current study, the application of Laponite nanoparticles as a crosslinking agent and rheology modifier has been proposed for improving the printability of the dAM-derived bioink. In this research, we have prepared dAM-derived hydrogels with different concentrations and systematically investigated their rheological properties with various rheological tests, including temperature sweep test, time sweep test at physiological temperature, and frequency sweep test; biological properties using well-known methods, such as MTT or CCK-8 assays, live/dead fluorescence imaging, and in vitro wound healing experiment; as well as printability by assessing the printing quality of printed two-dimensional (2D) and three-dimensional (3D) constructs. Our results demonstrated that enhanced viscosity and mechanical properties (from 41.8 to 896.2 Pa) of dAM bioink by increasing concentration from 1 to 3 %w/v facilitates printing 2D patterns with superior shape fidelity. Nevertheless, the dAM bioink did not provide enough stability for printing 3D models. The dynamic mechanical modulus of the hydrogel was significantly enhanced by the incorporation of laponite nanoparticles into the dAM-sodium alginate hybrid hydrogel system (up to 16 folds, for example, storage modulus increased from ~ 0.5 kPa to 8.4 kPa by adding 1% Laponite), which facilitated 3D printing of free-standing constructs without compromising biological properties. Meanwhile, excess agglomeration of the nanoparticles in an ion-containing medium leading to nozzle clogging was observed at high Laponite concentrations (≥2%). Microstructural evaluations also revealed nanoparticle-induced changes in the pore structures of the hydrogel, i.e. a finer pore structure was obtained. Biological assays affirmed the biocompatibility of the nanoengineered hydrogels, while wound healing experiments revealed the positive effect of Laponite on fibroblast cell migration, as evidenced by ~ 30% enhancement in the in vitro wound healing rate after 36 h. Our results show that after introducing nanosilicates, loose interactions between the hydrophobic nanosilicate tactoids and hydrophilic polymer form laminated clay clusters surrounded by collagen fibers. At a high concentration of Laponite (1:1 w/w), liquid-solid phase separation may also occur. A decreased storage modulus (up to 80 %), swelling ratio (up to 50 %), and gelation rate (up to 16 %) are thus attained. Subsequently, electrosteric stabilization of the nanoparticles with amine-terminated polyethyleneglycol (AT-PEG) prevented aggregation of the nanoparticles in the hydrogel matrix and provided uniform distribution. In vitro cell studies also determine that the dAM-derived hydrogels containing AT-PEG-modified Laponite exhibit higher cell viability and cell adhesion. Generally, the results obtained in this study demonstrate that the developed bioinks based on dAM provide suitable structural integrity and biocompatibility, highlighting their potential for therapeutic applications, particularly skin tissue engineering and wound healing
  9. Keywords:
  10. Bioink ; Silica Nanoparticles ; Surface Functionality ; Wound Healing ; Decellularized Amnion-Derived Hydrogel ; Three Dimentional Bioprinting ; Rheological Properties ; Mechanical Properties ; Biological Properties ; Tissue Engineering

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