Autologous protein solution prepared from the blood of osteoarthritic patients contains an enhanced profile of anti-inflammatory cytokines and anabolic growth factors.

The objective of this clinical study was to test if blood from osteoarthritis (OA) patients (n = 105) could be processed by a device system to form an autologous protein solution (APS) with preferentially increased concentrations of anti-inflammatory cytokines compared to inflammatory cytokines. To address this objective, APS was prepared from patients exhibiting radiographic evidence of knee OA. Patient metrics were collected including: demographic information, medical history, medication records, and Knee Injury and Osteoarthritis Outcome Score (KOOS) surveys. Cytokine and growth factor concentrations in whole blood and APS were measured using enzyme-linked immunosorbent assays. Statistical analyses were used to identify relationships between OA patient metrics and cytokines. The results of this study indicated that anti-inflammatory cytokines were preferentially increased compared to inflammatory cytokines in APS from 98% of OA patients. APS contained high concentrations of anti-inflammatory proteins including 39,000 ± 20,000 pg/ml IL-1ra, 21,000 ± 5,000 pg/ml sIL-1RII, 2,100 ± 570 pg/ml sTNF-RI, and 4,200 ± 1,500 pg/ml sTNF-RII. Analysis of the 82 patient metrics indicated that no single patient metric was strongly correlated (R(2) > 0.7) with the key cytokine concentrations in APS. Therefore, APS can be prepared from a broad range of OA patients.

The effect of thrombin activation of platelet-rich plasma on demineralized bone matrix osteoinductivity.

BACKGROUND: Demineralized bone matrix is an osteoinductive and osteoconductive material that is often used in orthopaedic surgery to induce bone formation. Autologous platelet-rich plasma, which contains proliferative and chemoattractant growth factors, has been used as a demineralized bone matrix adjuvant with mixed results. One variable during clinical use appears to be whether the platelet-rich plasma is activated with thrombin or is implanted in a liquid form with intact platelets. The objective of the present study was to determine if platelet-rich plasma can increase the osteoinductivity of demineralized bone matrix when used without thrombin activation. METHODS: The bioactivity of the demineralized bone matrix was evaluated in vitro by determining alkaline phosphatase production by C2C12 myoblast cells. The effect of thrombin activation on platelet-rich plasma was studied in vitro by evaluating osteosarcoma and bone marrow stromal cells for cell number and transforming growth factor-beta1 activation. Demineralized bone matrices supplemented with platelet-rich plasma, with or without thrombin activation, were implanted intramuscularly in athymic rats and were examined at fourteen, twenty-eight, and fifty-six days. Histological samples were analyzed for osteogenesis and chondrogenesis. Osteogenesis was further characterized on the basis of alkaline phosphatase activity. RESULTS: In vitro, thrombin triggered an immediate release of growth factors from the platelet-rich plasma, and the platelet-rich plasma increased the number of both osteosarcoma and stromal cells in a dose-dependent manner. Thrombin activation of the platelet-rich plasma eliminated such stimulatory effects. In vivo, the platelet-rich plasma stimulated chondrogenesis on Day 14 and osteogenesis on Days 28 and 56, whereas thrombin-activated platelet-rich plasma acted as an inhibitor of such events. In addition, inflammatory cells were detected in demineralized bone matrix samples that were mixed with thrombin-activated platelet-rich plasma. These cells were not present in matrix mixed with platelet-rich plasma alone. CONCLUSIONS: Platelet-rich plasma significantly increased in vivo demineralized bone matrix osteoinductivity only when used without thrombin activation.

Demineralized bone matrix graft: a scientific and clinical case study assessment.

Osteoinductive demineralized bone matrix results from bone demineralization and is attributed to matrix-associated bone morphogenetic proteins. The osteoinductive potential can vary with donor. Many bioassay methods are available to screen donors, each with its own interpretation, so performance of more than one may be of value. Furthermore, little is known about the relationship between bioassay results and clinical outcomes. A study designed to meaningfully explore these issues would require assay of a large number of donors as well as clinical utilization in a large patient population. A preliminary study was undertaken to gain initial perspective. Using demineralized bone matrix derived from one 33-year-old female donor, 2 methods of bioassay and a clinical case study were performed. The levels of bone morphogenetic proteins 2, 4, and 7 in lyophilized demineralized bone matrix powder were measured (19.65 +/- 0.30 ng/g, 2.49 +/- 0.19 ng/g, and 82.03 +/- 6.89 ng/g, respectively). Also, putty (Osteostim DBM Putty), prepared from powder, was intramuscularly implanted in athymic rats and de novo bone formation quantified (6.7% +/- 3.5% new bone formation with 49% +/- 17% of the implant area associated with new bone formation). The putty, in conjunction with internal fixation, was used in the revision of a medial malleolar nonunion of an obese, 76-year-old woman. Radiographic union with excellent graft incorporation was achieved by 12 weeks postoperatively, with maintenance of an acceptable clinical result during the 14-month follow-up period. These results are interpreted in the broader context of demineralized bone grafting, in general, and an outline for further study is presented.