Evaluation of Marine Agarose Biomaterials for Tissue Engineering Applications

Abstract

Five agarose types (D1LE, D2LE, LM, MS8 and D5) were evaluated in tissue engineering and compared for the first time using an array of analysis methods. Acellular and cellular constructs were generated from 0.3–3%, and their biomechanical properties, in vivo biocompatibility (as determined by LIVE/DEAD, WST-1 and DNA release, with n = 6 per sample) and in vivo biocompatibility (by hematological and biochemical analyses and histology, with n = 4 animals per agarose type) were analyzed. Results revealed that the biomechanical properties of each hydrogel were related to the agarose concentration (p < 0.001). Regarding the agarose type, the highest (p < 0.001) Young modulus, stress at fracture and break load were D1LE, D2LE and D5, whereas the strain at fracture was higher in D5 and MS8 at 3% (p < 0.05). All agaroses showed high biocompatibility on human skin cells, especially in indirect contact, with a correlation with agarose concentration (p = 0.0074 for LIVE/DEAD and p = 0.0014 for WST-1) and type, although cell function tended to decrease in direct contact with highly concentrated agaroses. All agaroses were safe in vivo, with no systemic effects as determined by hematological and biochemical analysis and histology of major organs. Locally, implants were partially encapsulated and a pro-regenerative response with abundant M2-type macrophages was found. In summary, we may state that all these agarose types can be safely used in tissue engineering and that the biomechanical properties and biocompatibility were strongly associated to the agarose concentration in the hydrogel and partially associated to the agarose type. These results open the door to the generation of specific agarose-based hydrogels for definite clinical applications such as the human skin, cornea or oral mucosa.

Fibrin and Marine-Derived Agaroses for the Generation of Human Bioartificial Tissues: An Ex Vivo and In Vivo Study

“…Although FA biomaterial showed promising results in tissue engineering of the cornea, skin, nerve, tendon, sclera and oral mucosa, the biocompatibility and biomechanical properties of this biomaterial still need to be improved [ 27 , 28 , 29 , 30 , 31 , 32 ]. In this regard, we recently demonstrated that different types of marine agaroses could have specific biological and biomechanical properties, suggesting that not all currently available marine agaroses polysaccharides show the same behavior both in vitro and in vivo [ 23 ]. In the present work, we evaluated several agaroses at increasing concentrations to determine their putative usefulness for the generation of human bioartificial tissues by tissue engineering.…”

“…Marine polysaccharides are other natural biomaterials that has been used in tissue engineering of different tissues and organs [ 20 ]. The inherent natural properties of these marine products such as biodegradability and biocompatibility, make agarose a perfect product to be used a scaffold in tissue engineering [ 21 , 22 , 23 ]. Agarose is a natural polysaccharide that exhibits controlled self-gelling properties, water solubility, and high biocompatibility, and it has been demonstrated that this material could enhance cell proliferation and activity [ 24 ].…”

“…Agarose is a natural polysaccharide that exhibits controlled self-gelling properties, water solubility, and high biocompatibility, and it has been demonstrated that this material could enhance cell proliferation and activity [ 24 ]. Several types of agaroses have been described so far, and we recently demonstrated that the concentration and the specific type of agarose used in tissue engineering can significantly influence the biomaterial biomechanical properties, biological functions, and in vivo biocompatibility [ 23 ]. Therefore, specific concentrations and types of agarose should be used for each specific application [ 23 ].…”

“…Several types of agaroses have been described so far, and we recently demonstrated that the concentration and the specific type of agarose used in tissue engineering can significantly influence the biomaterial biomechanical properties, biological functions, and in vivo biocompatibility [ 23 ]. Therefore, specific concentrations and types of agarose should be used for each specific application [ 23 ]. Although agarose has adequate biological and biomechanical properties, this product has several limitations, especially regarding cell differentiation [ 22 ].…”

Development of secretome-based strategies to improve cell culture protocols in tissue engineering

“…Cell proliferation was then analyzed with the WST-1 colorimetric assay (Cell proliferation reagent WST-1, Sigma/Aldrich Merck) as previously reported 28 , 29 . Briefly, water-soluble tetrazolium salt was added to the culture medium of each study group (HSEC cultured with the different types and concentrations of secretome at different time points), and cells were incubated in this mixture for 4 h at 37 °C.…”