s / Osteoarthritis and Cartilage 20 (2012) S54–S296 S250 medium also demonstrated time-dependency with viable cell number significantly increasing between 1 and 3 days (P<0.001). PDGF-AB and IGF-I concentrations in the cell supernatant decreased over the 6 day period (P<0.001 and P<0.05 respectively compared to 10% PRP alone). TGF-beta concentrations significantly increased after 6 days (P<0.001). VEGF concentrations increased significantly compared to 10% PRP after 3 and 6 days (P<0.05 and P<0.001 respectively). Growth factor concentrations remained low and insignificant in serum-starved controls at all time points. The addition of 10% PRP to cells stimulated phosphorylation of multiple pro-survival and pro-mitotic proteins within a 5 to 30 minute period. Conclusions: Growth factors released by activated platelets act upon human tendon cells to strongly increase viable cell number which would, in vivo, directly support the healing response. Multiple signalling pathways are activated by PRP, including classical proliferation and survival kinases. As expected, the concentration of some growth factors decreases over time. However, PRP strongly induced production of TGF-beta and VEGF by the tenocytes themselves. During the initial phases of healing these increases are likely to drive both matrix production/turnover and revascularisation. However excessive use of PRP may also cause fibrosis due to excessive TGF-driven cell proliferation and pain due to inappropriately high VEGF-driven neovascularisation. PRP contains a highly active cocktail of growth factors and produces both immediate and sustained effects on human tenocytes indicating considerable regenerative potential in vivo. 494 SERUM RELAXIN IS CORRELATED WITH RELAXIN RECEPTORS AND MMP-1 IN THE ANTERIOR OBLIQUE LIGAMENT J. Wolf , F. Scott , E. Etchell , A.E. Williams , S. Delaronde , K.B. King . Univ. of Connecticut, Farmington, CT, USA; Univ. of Colorado, Aurora, CO, USA; Bay Pines Veterans Admin. Hosp., Bay Pines, FL, USA Purpose: Introduction: Relaxin is a hormone produced in pregnancy, mediated at the cellular level by damage to the extracellular matrix. We hypothesized that relaxin was a target hormone involved in the development of trapeziometacarpal (TM) joint laxity, potentially as a cause of ligament loosening and attrition, and later osteoarthritis. The purpose of this study was to quantitatively evaluate relaxin receptors in the anterior oblique ligament of the thumb as well as the relationship of serum relaxin and matrix metalloproteases (MMPs) to these receptors. Methods: Consecutive patients undergoing carpometacarpal joint arthroplasty with trapeziectomy and ligament reconstruction by two hand surgeons were enrolled prospectively. Patients underwent examination of laxity using the Beighton-Horan measurement, blood draw for serum relaxin, and sampling of the anterior oblique ligament during surgery. Serum relaxin was determined by enzyme-linked immunosorbent assay (ELISA). The ligament underwent extraction of RNA and analysis with quantitative reverse-transcriptase polymerasechain reaction (qRT-PCR) for relaxin receptor (RXFP-1), MMP-1, and MMP-3. Results: A total of 49 patients were enrolled, including 30 women and 19 men with an average age of 62 (range, 43-78 years). Serum relaxin levels averaged 3.73 pg/ml (range, 0-9.45). The mean RXFP-1 receptor level was 5.23 x 106 attograms (ag), while MMP-1 and MMP-3 were expressed at much higher levels (0.022 ag and 0.318 ag, respectively). When compared to the serum relaxin levels in these patients, there was a significant relationship between serum relaxin and the log of RXFP-1 (p1⁄40.02) and MMP-1 (p1⁄40.05), using Pearson correlations and linear regression analysis. Conclusions: These findings confirm the presence of relaxin receptors in the anterior oblique ligament as well as MMPs 1 and 3, which are known to be upregulated by relaxin binding. The relationship between serum relaxin and RXFP-1 and MMP-1 suggests that as circulating relaxin levels rise, increased ligamentous attenuation could potentially occur in the anterior oblique ligament. This may be a factor in the development of trapeziometacarpal arthritis. Ă 495 ULTRASTRUCTURAL STUDY OF THE HEALING PROCESS IN TENOTOMIZED ACHILLES TENDONS TREATED WITH PLASMA RICH IN GROWTH FACTORS IN SHEEP J.M. Dominguez , J. Fernandez-Sarmiento , M.M. Granados , J. Morgaz , R. Navarrete , J.M. Carrillo , M. Rubio , M. Garcia-Balletbo , R. Cugat , J. Monterde , A. Blanco . Dept. of Animal Med. and Surgery, Univ. of Cordoba, Cordoba, Spain; Dept. of Animal Med. and Surgery, Univ. of Cardenal Herrera, Valencia, Spain; Deparment of Orthopaedic Surgery and Traumatology, Hosp. Quiron, Barcelona, Spain; Dept. of Comparative Pathology, Univ. of Cordoba, Cordoba, Spain Purpose: The aimwas to evaluate the effects of the ultrastructural healing process in tenotomized tendons treated with plasma rich in growth factors (PRGF) in sheep at 2, 4 and 8 weeks. Methods: Forty-two adult sheep were anaesthetized and the Achilles tendon was experimentally tenotomized and repaired. Animals were randomly assigned into six groups. Three groups received an infiltration of PRGF on the repaired area just after repairing the tenotomy and, one of them recived one more injection after one week, and two of them recived the infiltration every week during the following 3 weeks. The other three groups received a placebo injection with saline. At 2 weeks, one PRGF group (PRGF2) and one saline group (S2) were euthanized, and the other four groups were euthanized at 4 weeks (PRGF4; S4) and 8 weeks (PRGF8; S8). Samples of 2 mm in size were taken in a longitudinal direction of the tendon axis from the incision sites which exhibited active regeneration. Tendon samples were fixed with glutaraldehyde and examined with transmission electron microscopy. Collagen fibril diameter and priodicity were measured. The Mann-Whitney U test was perform to analyze the significance between groups. Results: At 2sd week, S2 group showed abundant vascularity, edema and inflammatory cells: B lymphocytes, plasma cells, eosinophils and macrophages. Collagen fibrils were thin, although there were some areas with hypertrophic fibrils. In PRFG2 group, low vascularization and inflammatory cells were seen but mature collagen fiber formation. At 4th week, S4 group still exhibited large areas with abundant vascular components and edema. Highlighting the presence of macrophages and fibroblasts. Collagen fibrils were scattered and anarchic surround the fibroblasts, they were thin, with a few unclear cross-striation masked by the edema. Collagen fibers thicken at specific points atypically, but keeping the striations, forming zones of accumulation of thickened fibers that give rise to microkeloid areas. PRGF4 group showed little sign of inflammation with low number of macrophages and vessels. Tenocytes become tenocyteblasts, considered highly active on the basis of their well-developed Golgi complex and rough