The systemic influence of platelet-derived growth factors on bone marrow mesenchymal stem cells in fracture patients.

BACKGROUND: Fracture healing is a complex process regulated by a variety of cells and signalling molecules which act both locally and systemically. The aim of this study was to investigate potential changes in patients' mesenchymal stem cells (MSCs) in the iliac crest (IC) bone marrow (BM) and in peripheral blood (PB) in relation to the severity of trauma and to correlate them with systemic changes reflective of inflammatory and platelet responses following fracture. METHODS: ICBM samples were aspirated from two trauma groups: isolated trauma and polytrauma (n = 8 and 18, respectively) at two time-points post-fracture and from non-trauma controls (n = 7). Matched PB was collected every other day for a minimum of 14 days. BM MSCs were enumerated using colony forming-fibroblast (CFU-F) assay and flow cytometry for the CD45-CD271+ phenotype. RESULTS: Regardless of the severity of trauma, no significant increase or decrease in BM MSCs was observed following fracture and MSCs were not mobilised into PB. However, direct positive correlations were observed between changes in the numbers of aspirated BM MSCs and time-matched changes in their serum PDGF-AA and -BB. In vitro, patients' serum induced MSC proliferation in a manner reflecting changes in PDGFs. PDGF receptors CD140a and CD140b were expressed on native CD45-CD271+ BM MSCs (average 12% and 64%, respectively) and changed over time in direct relationship with platelets/PDGFs. CONCLUSIONS: Platelet lysates and other platelet-derived products are used to expand MSCs ex vivo. This study demonstrates that endogenous PDGFs can influence MSC responses in vivo. This indicates a highly dynamic, rather than static, MSC nature in humans.

Single-platform quality control assay to quantify multipotential stromal cells in bone marrow aspirates prior to bulk manufacture or direct therapeutic use.

BACKGROUND AIMS: The manufacture of multipotential stromal cell (MSC)-based products is costly; therefore, a rapid evaluation of bone marrow (BM) 'quality' with respect to MSC content is desirable. The aim of this study was to develop a rapid single-platform assay to quantify MSC in BM aspirates. METHODS: Aspirated MSC were enumerated using the CD45(-/low) CD271(bright) phenotype and AccuCheck counting beads and compared with a classic colony-forming unit-fibroblast (CFU-F) assay. The phenotype of CD45(-/low) CD271(bright) cells was defined using a range of MSC (CD73, CD105, CD90) and non-MSC (CD31, CD33, CD34, CD19) markers. The effect of aspirated BM volume on MSC yield was also determined. RESULTS: CD45(-/low) CD271(bright) cells had a classic MSC phenotype (CD73(+) CD105(+) CD90(+)). Their numbers correlated positively with CFU-F counted manually (R = 0.81, P < 0.001) or using automatic measurements of surface area occupied by colonies (R = 0.66, P < 0.001). Simultaneous enumeration of CD34(+) cells revealed donor variability ranges compatible with standard International Society of Hematotherapy and Graft Engineering (ISHGE) protocols. Aspirating larger marrow volumes gave a significant several-fold reduction in the frequency of CFU-F and CD45(-/low) CD271(bright) cells per milliliter. Therefore aspirated MSC yields can be maximized through a standardized, low-volume harvesting technique. CONCLUSIONS: Absolute quantification of CD45(-/low) CD271(bright) cells was found to be a reliable method of predicting CFU-F yields in BM aspirates. This rapid (< 40 min) procedure could be suitable for intra-operative quality control of BM aspirates prior to volume reduction/direct injection in orthopedics. In the production of culture-expanded MSC, this assay could be used to exclude samples containing low numbers of MSC, resulting in improved consistency and quality of manufactured MSC batches.

Proximal tibial fractures: early experience using polyaxial locking-plate technology.

Between 2004 and 2009, 60 patients with proximal tibial fractures were included in this prospective study. All fractures were treated with the polyaxial locked-plate fixation system (DePuy, Warsaw, IN, USA). Clinical and radiographic data, including fracture pattern, changes in alignment, local and systemic complications, hardware failure and fracture union were analysed. The mean follow-up was 14 (12-36) months. According to the Orthopaedic Trauma Association (OTA) classification, there were five 41-A, 28 41-B and 27 41-C fractures. Fractures were treated percutaneously in 30% of cases. Double-plating was used in 11 cases. All but three fractures progressed to union at a mean of 3.2 (2.5-5) months. There was no evidence of varus collapse as a result of polyaxial screw failure. No plate fractured, and no screw cut out was noted. There was one case of lateral joint collapse (>10°) in a patient with open bicondylar plateau fracture. The mean Knee Society Score at the time of final follow-up was 91 points, and the mean functional score was 89 points. The polyaxial locking-plate system provided stable fixation of extra-articular and intra-articular proximal tibial fractures and good functional outcomes with a low complication rate.

Novel anticoagulants: new evidence for emerging drugs and their potential application in major lower limb surgery.

For decades, parenteral drugs, such as the low molecular weight heparins and unfractionated heparins or vitamin K antagonists, have been used as anticoagulants for prevention of venous thromboembolism following major lower limb surgery. However, these regiments have limitations that rendered the quest for new anticoagulants mandatory. Recently, research has been focused on the development of orally active small molecules that directly target thrombin or activated factor X (FXa). These regiments exhibit a number of characteristics that an "ideal" anticoagulant should possess. Currently, two agents, dabigatran etexilate and rivaroxaban, which inhibit thrombin and FXa, respectively have been approved in the European Union and Canada for venous thromboprophylaxis in patients undergoing elective hip- or knee-replacement surgery. Other agents are at an early or late stage of clinical evaluation. In this study, we summarize the current evidence for these new developed or under development drugs regarding their applications in the filed of lower limb orthopaedic surgery.

The timing of drug administration for thromboprophylaxis following orthopaedic surgery: evidence and controversies related to treatment initiation and duration.

Patients undergoing major orthopaedic surgery of the lower extremities or spine are at increased risk of venous thromboembolism (VTE). Although consensus exists as to the need for routine thromboprophylaxis in high risk patients, some aspects of this approach, such as the timing of the first dose and overall duration of the anticoagulation regimen, are subject to debate. Reviewing the available literature, there appears to be little evidence to support initiation of thromboprophylaxis more than 12 hours before surgery. Perioperative thromboprophylaxis (2 hours pre to 6 hours post -op) has been associated with an increased risk of bleeding complications whilst initiating prophylaxis more than 12 hours after surgery appears to increase the incidence of subsequent thromboembolic complications. Overall evidence would appear to support initiation of thromboprophylaxis 6 to 9 hours postoperatively, though further confirmatory studies investigating this variable in isolation would be useful to guide clinical decision making. Although evidence exists supporting extended duration thromboprophylaxis after major orthopaedic procedures, further evidence is required, using clinically important end points, prior to adoption of such an approach in all patients. Stratification of prophylaxis duration, based on risk factors for thromboembolic or bleeding complications, would seem a more rational approach than strict adherence to guidelines.