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CORRELATION BETWEEN THE SOFTNESS OF HYALURONIC ACID MICROPARTICULATE SCAFFOLDS AND THE PROLIFERATION OF HUMAN ADIPOSE MESENCHYMAL CELLS
Hyaluronic acid (HA) is a naturally occurring biomacromolecule found in most connective tissues and is particularly concentrated in synovial fluid. HA chemical structure is composed by disaccharide units of D-glucuronic acid and D-N-acetylglucosamine, linked together through alternating beta-1,4 and beta-1,3 glycosidic bonds. HA polymer structure is characterized by large and linear chains in which the number of repeat disaccharide units leads to a molecular mass of 400 Da. Nowadays, HA has been widely used as a viscosupplement in orthopedics, in plastic, intraocular and abdominal surgeries and wound healing. High molecular weight hyaluronic acid (HA) associated to platelet-rich plasma (PRP) has been used for better effects in wound healing and regenerative medicine.
However, the fluid or plain HA is very soluble in aqueous medium , and modifications are necessary to provide structural stability. HA molecular structure presents sites for covalent modification, such as carboxylic acid and hydroxyl, which have been explored for production of useful materials for biomedical applications, as well as to the production of scaffolds for cell cultivation.
Objetivos - Metodologia - Resultados - Discussão dos Resultados/Objectives - Methodology - Results - Discussion of Results/Objetivos - Metodología - Resultados - Discusión de los resultados
This study aimed to prepare and characterize various crosslinked HA based scaffolds, structured in microparticles, and to evaluate their performance on "in vitro" proliferation of human adipose mesenchymal cells (AdMSCs) stimulated by the growth factors from PRP. The rational for using crosslinked HA is to increase stability against degradation and increase the residence time "in vivo" compared to non-crosslinked fHA, as well as the structuration in microparticles provides a larger surface area for cell proliferation.
HA was crosslinked with 14-butanediol diglycidyl ether (BDDE), forming HA-BDDE gels, and autocrosslinked in its ester bonds by means of intermediate activation, giving HA- ACP gels. These gels also were treated with chitosan for attenuation of HA negative charge. The mean diameter of the microparticles was 200µm approximately. PRP was prepared by centrifugation of whole blood and activated by autologous thrombin and calcium chloride (aPRP). The gels were evaluated in a proportion of 200mg of crosslinked HA to 200 µL aPRP
The rheological characterization shown all gels were predominantly viscoelastic. The viscoelasticity increased from HA-BDDE:PRP > HA-ACP-CHI:PRP> HA-ACP:PRP> aPRP. Non-crosslinked HA did not present a gel behavior. The complex module, G*, which is related with rigidity of the material, decreased from HA-BDDE:PRP > HA-ACP-CHI:PRP> HA-ACP:PRP> aPRP The range of G* was from 1500 (HA-BDDE) to 47 (HA-ACP:PRP) and 21 (aPRP). The "in vitro" culture of AdMSCs shown higher proliferation in HA-ACP-CHI:PRP scaffolds and aPRP. The lower cell proliferation was obtained in HA-BDDE:PRP . Therefore, despite the much larger surface area provided by the fibrin fibers in aPRP, and also the instability of the HA-ACP:PRP microparticles, there was a clear correlation between the softness (lower rigidity) and AdMSCs proliferation. The softness surface of the HA-ACP-CHI :PRP microparticles added to its viscoelasticity provided a favorable environment to adhesion and proliferation of AdMSCs.
Considerações Finais/Final considerations/Consideraciones finales
These results are relevant to the development of HA based scaffolds for applications in healing and regenerative trerapies.
Palavras-chave/Key words/Palabras clave
Hyaluronic acid, Microparticles, Scaffolds, Human Adipose Mesenchymal cells
Scaffold (surface topology and softness)
MARIA HELENA ANDRADE SANTANA, Angela Cristina Luzo, Andréa Martins Shimojo