In vitro performance of injectable chitosan-tripolyphosphate scaffolds combined with platelet-rich plasma.

This study aimed to evaluate the in vitro biological effectiveness of chitosan microparticles crosslinked with sodium tripolyphosphate (TPP) in combination with activated pure platelet-rich plasma (aP-PRP) as an injectable composite scaffold for growth factors release, cell proliferation and osteogenic differentiation. Two main novelties were addressed in the field of scaffolds in regenerative medicine: the first is the approach including simultaneously the three vertices of the proliferation triangle formed by the capabilities genic progenitor cells, conductive scaffolds and inductive growth factors, which are provided by platelet rich plasma (PRP); secondly, the approach of an injectable composite scaffolds formed by the fibrin network from aP-PRP and the chitosan microparticles crosslinked with TPP. The microparticles were prepared by vortexing the chitosan and TPP solutions. The ionic crosslinking of chitosan with TPP was made at mass ratios of 2:1, 5:1, and 10:1 at pH 4.0. P-PRP was obtained via the controlled centrifugation of whole blood. The composite scaffolds were prepared by adding the microparticles to immediately activated P-PRP. The results showed that the microparticles had adequate physicochemical and mechanical properties for injection. Furthermore, the microparticles controlled the release of growth factors from P-PRP. The proliferation of human adipose-derived mesenchymal stem cells was lower than in aP-PRP alone but significant at a 2:1 chitosan-TPP mass ratio. Osteogenic differentiation was stimulated at all studied mass ratios, as indicated by the alkaline phosphatase activity. These results offer perspectives for optimizing the composite scaffold, and to prove its potential as an injectable scaffold in regenerative medicine.

Sterilization of auto-crosslinked hyaluronic acid scaffolds structured in microparticles and sponges.

This work evaluated the effects of UV irradiation, plasma radiation, steam and 70% ethanol treatments on the sterilization and integrity of auto-crosslinked hyaluronic acid (HA-ACP) scaffolds structured in microparticles and sponges aiming in vivo applications for regenerative medicine of bone tissue. The integrity of the microparticles was characterized by rheological behavior, while for the sponges, it was characterized by scanning electron microscopy, Fourier transform infrared spectroscopy and differential scanning calorimetry. The effectiveness of the sterilization treatment was verified by the number of microorganism colonies in the samples after the treatments. In conclusion, plasma radiation was the best treatment for the sponges, while steam sterilization in the autoclave at 126°C (1.5 kgf/cm2) for 5 min was the best treatment for the microparticles.

Performance of PRP associated with porous chitosan as a composite scaffold for regenerative medicine.

This study aimed to evaluate the in vitro performance of activated platelet-rich plasma associated with porous sponges of chitosan as a composite scaffold for proliferation and osteogenic differentiation of human adipose tissue-derived mesenchymal stem cells. The sponges were prepared by controlled freezing (-20, -80, or -196°C) and lyophilization of chitosan solutions (1, 2, or 3% w/v). The platelet-rich plasma was obtained from controlled centrifugation of whole blood and activated with calcium and autologous serum. The composite scaffolds were prepared by embedding the sponges with the activated platelet-rich plasma. The results showed the performance of the scaffolds was superior to that of activated platelet-rich plasma alone, in terms of delaying the release of growth factors and increased proliferation of the stem cells. The best preparation conditions of chitosan composite scaffolds that coordinated the physicochemical and mechanical properties and cell proliferation were 3% (w/v) chitosan and a -20°C freezing temperature, while -196°C favored osteogenic differentiation. Although the composite scaffolds are promising for regenerative medicine, the structures require stabilization to prevent the collapse observed after five days.

The crosslinking degree controls the mechanical, rheological, and swelling properties of hyaluronic acid microparticles.

Viscosupplements, used for treating joint and cartilage diseases, restore the rheological properties of synovial fluid, regulate joint homeostasis and act as scaffolds for cell growth and tissue regeneration. Most viscosupplements are hydrogels composed of hyaluronic acid (HA) microparticles suspended in fluid HA. These microparticles are crosslinked with chemicals to assure their stability against enzyme degradation and to prolong the action of the viscosupplement. However, the crosslinking also modifies the mechanical, swelling and rheological properties of the HA microparticle hydrogels, with consequences on the effectiveness of the application. The aim of this study is to correlate the crosslinking degree (CD) with these properties to achieve modulation of HA/DVS microparticles through CD control. Because divinyl sulfone (DVS) is the usual crosslinker of HA in viscosupplements, we examined the effects of CD by preparing HA microparticles at 1:1, 2:1, 3:1, and 5:1 HA/DVS mass ratios. The CD was calculated from inductively coupled plasma spectrometry data. HA microparticles were previously sized to a mean diameter of 87.5 µm. Higher CD increased the viscoelasticity and the extrusion force and reduced the swelling of the HA microparticle hydrogels, which also showed Newtonian pseudoplastic behavior and were classified as covalent weak. The hydrogels were not cytotoxic to fibroblasts according to an MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay.