Abstract
Platelet-rich plasma (PRP) associated with high molecular weight hyaluronic acid (HA) has been clinically used for tissue regeneration in orthopedics. Despite the recognized beneficial clinical outcomes (e.g., early pain control, improvement of patients’ functional limitation and longer-term effectiveness compared to PRP and HA alone in mild and moderate osteoarthritis treatments), its use is still challenging and controversial due to lack of standardization of association practical protocols. Moreover, most studies neglect the matrix structure, that generates the ultimate properties of the association among platelets, fibrin network and the microparticles. In the present work, we aimed to analyze the influence of the PRP/HA association with a controlled matrix structure on the stability, rheological behavior, release of growth factors and in vitro proliferation of human adipose-derived mesenchymal cells (h-AdMSCs). The attenuation of the negative charge of HA was also evaluated. Pure PRP (P-PRP) (i.e., plasma enriched with platelets and poor in leukocytes) was prepared by centrifugation and activated with serum and calcium chloride (AP-PRP). Autocrosslinked hyaluronic acid (AHA) was prepared by organocatalyzed auto-esterification and structured in microparticles (MPAHA) by shearing. The attenuation of the negative charge of MPAHA was performed with chitosan (CHT) by polyelectrolyte complexation yielding MPAHA-CHT. The results showed that microparticles (MPs) have viscoelastic properties, extrusion force and swelling ratio appropriate for injectable applications. The association of AP-PRP with the controlled structure of MPAHA and MPAHA-CHT formed a matrix composed of platelets and of a fibrin network with fibers around 160 nm located preferably on the surface of the MPs with an average diameter of 250 μm. Moreover, AP-PRP/MPAHA and AP-PRP/MPAHA-CHT associations were non-toxic and supported controlled growth factor (PDGF-AB and TGF-β1) release and in vitro proliferation of h-AdMSC with a similar pattern to that of AP-PRP alone. The best h-AdMSC proliferation was obtained with the AP-PRP/MPAHA-CHT75:25 indicating that the charge attenuation improved the cell proliferation. Thus, the association of AP-PRP with the controlled structure of HA can be a valuable approach for orthopedic applications.
Highlights
Platelet-rich plasma (PRP) is a concentrate of platelets and other components of plasma, which has been widely used in regenerative medicine [1,2,3,4]
The spectrum of Autocrosslinked hyaluronic acid (AHA) showed the characteristic bands of polysaccharides (Supplementary Figure S4) between 3600 and 2800 cm−1 attributed to O–H stretching vibration and between 3000 and 2800 cm−1 attributed to the C–H stretching vibrations and bands of N–H stretching vibrations overlapped with the O–H bands, as expected
We studied a matrix consisted of platelets, fibrin network and hyaluronic acid (HA) crosslinked microparticles
Summary
Platelet-rich plasma (PRP) is a concentrate of platelets and other components of plasma, which has been widely used in regenerative medicine [1,2,3,4]. The regenerative potential of PRP is based on the release of growth factors that occurs with platelet degranulation. The activation of PRP with autologous serum and CaCl2 promotes the release of growth factors, which stimulate cell proliferation and differentiation. PRP has been associated with high molecular weight hyaluronic acid (HA), mainly for orthopedic therapies [2,5]. HA is a glycosaminoglycan formed by a variable number of repeating units of D-glucuronic acid and N-acetylglucosamine. Besides its role in lubrication and joints protection against shocks, HA exhibits anti-inflammatory effects, relieves pain, provides signalization of specific receptors (such as CD44), activates intrinsic repair processes of the cartilage, and normalizes the endogenous production of HA [2,5,6,7]
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