Platelet Rich Plasma and Hyaluronic Acid Blend for the Treatment of Osteoarthritis: Rheological and Biological Evaluation

Abstract

Introduction: Osteoarthritis (OA) is the most common musculoskeletal disease. Current treatments for OA are mainly symptomatic and inadequate since none results in restoration of fully functional cartilage. Hyaluronic Acid (HA) intra-articular injections are widely accepted for the treatment of pain associated to OA. The goal of HA viscosupplementation is to reduce pain and improve viscoelasticity of synovial fluid. Platelet-rich plasma (PRP) has been also employed to treat OA to possibly induce cartilage regeneration. The combination of HA and PRP could supply many advantages for tissue repair. Indeed, it conjugates HA viscosupplementation with PRP regenerative properties. The aim of this study was to evaluate the rheological and biological properties of different HA compositions in combination with PRP in order to identify (i) the viscoelastic features of the HA-PRP blends, (ii) their biological effect on osteoarthritic chondrocytes and (iii) HA formulations suitable for use in combination with PRP.

Materials and methods: HA/PRP blends have been obtained mixing human PRP and three different HA at different concentrations: 1) Sinovial, 0.8% (SN); 2) Sinovial Forte 1.6% (SF); 3) Sinovial HL 3.2% (HL); 4) Hyalubrix 1.5% (HX). Combinations of phosphate buffered saline (PBS) and the four HA types were used as control. Rheological measurements were performed on an Anton PaarMCR-302 rheometer. Amplitude sweep, frequency sweep and rotational measurements were performed and viscoelastic properties were evaluated. The rheological data were validated performing the tests in presence of Bovine Serum Albumin (BSA) up to ultra-physiological concentration (7%). Primary osteoarthritic chondrocytes were cultured in vitro with the HA and PRP blends in the culture medium for one week. Cell viability, proliferation and glycosaminoglycan (GAG) content were assessed.

Results: PRP addition to HA leads to a decrease of viscoelastic shear moduli and increase of the crossover point, due to a pure dilution effect. For viscosupplements with HA concentration below 1% the viscoelasticity is mostly lost. Results were validated also in presence of proteins, which in synovial fluid are more abundant than HA. Chondrocytes proliferated overtime in all different culture conditions. The proliferation rate was higher in chondrocytes cultured in the media containing PRP compared to the cultures with different HA alone. GAG content was significantly higher in chondrocytes cultured in PRP and HL blend.

Discussion: We investigated the rheological and biological properties of four different HA concentrations when combined with PRP giving insights on viscoelastic and biological properties of a promising approach for future OA therapy. Our data demonstrate that PRP addition is not detrimental to the viscosupplementation effect of HA. Viscosupplements containing low HA concentration are not indicated for combination with PRP, as the viscoelastic properties are lost. Although having the same rheological behavior of SF and HX, HL was superior in stimulating extracellular matrix production in vitro.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1

(A) Viscosity curves of the pure viscosupplements. The graph represents the viscosity as a function of the shear rate. For all specimens the typical shear-thinning behavior of HA is observed. SN displays viscosity one order of magnitude lower than the other specimens. (B) Mechanical spectra of the pure viscosupplements. The graph represents the shear storage modulus G' (triangles) and the shear loss modulus G" (circle) as a function of the angular frequency.

Fig 2

(A): Comparison of the mechanical spectrum of pure HX and its combinations: pure HX (black); HX+PBS 1:1 (red); HX+PRP 1:1 (blue); HX+PRP pre-added to 7% BSA (dark yellow). All HX combinations show a similar rheological profile, which is dictated by the dilution effect (B): Comparison of the mechanical spectrum of pure HL and its combinations: pure HL (black); HL+PBS 1:1 (red); HL+PRP 1:1 (blue); HL+PRP pre-added to 7% BSA (dark yellow). All HL combinations show a similar rheological profile, which is dictated by the dilution effect.

Fig 3

(A): Mechanical spectra of HL and SF in combination with PRP. (B): Viscosity curve of HL and SF in combination with PRP.

Fig 4

(A): Cell viability and proliferation overtime of chondrocytes cultured under different conditions normalized to the rates of chondrocytes cultured in growing media at day 0, 3 and 7. (B): GAG content normalized to DNA of chondrocytes cultured for 7 days under different conditions compared to the GAG content of chondrocytes cultured in growing media (*: p<0.05).