Clinical experiences in the use of a gentamicin-coated titanium nail in tibia fractures.

Despite the improvement of surgical techniques surgical site infections (SSIs) still remain clinically challenging in high risk patients undergoing osteosynthesis for tibia fractures. The use of an antibiotic coated implant might reduce the adhesion of bacteria on the implant surface and could therefore reduce the rate of implant-related infection or osteomyelitis. A gentamicin-coated tibia nail was evaluated in a prospective study. Four centers enrolled 100 patients (99 treated) with fresh open or closed tibia fractures, or for non-union revision surgery and followed them for 18 months. Data collected included infection events, radiographs, SF-12, EQ-5D, Iowa Ankle score, and the WOMAC questionnaire. Sixty-eight of the 99 treated patients suffered from a fresh fracture, while in 31 patients, the intramedullary nail was implanted for revision purposes, including non-unions due to infection. Fifteen (22%) of the fresh fractures were GA Type III. The follow-up rate was 87% and 82% at 12 months and 18 months, respectively. Deep surgical site infections occurred in 3 fresh fractures and two in revision surgeries. We did not observe any local or systemic toxic effects related to gentamicin during this study. The use of the antibiotic coated nail is an option in patients with a high infection risk, like open factures or infected non unions, in the prevention of the onset of an implant-related infection or osteomyelitis.

[Tibial defects and infected non-unions : Treatment results after Masquelet technique]. / Tibiadefekt- und Infektpseudarthrosen : Behandlungsergebnisse nach Masquelet-Technik.

BACKGROUND: The treatment of non-unions with large bone defects or osteitis is a major challenge in orthopedic and trauma surgery. A new concept of therapy is a two-step procedure: Masquelet technique according to the diamond concept. METHODS: Between February 2010 and June 2014, 55 patients with tibia non-unions or infections were treated in a two-step Masquelet technique in our center. The patients' average age was 48 (median 50; minimum 15-maximum 72) with an average BMI (body mass index) of 28 (27; 18-52). There were 10 (18 %) female and 45 (82 %) male patients in the group. All study patients went through a follow up. Bone healing and clinical functional data were collected, as well as data according to subjective patient statements about pain and everyday limitations. RESULTS: In 42 cases (76.4 %) the outcome was a sufficient bony consolidation. On average, the time to heal was 10.3 (8, 5; 3-40) months, defect gaps were 4 cm (3 cm; 0,6-26 cm), and on average the patients had had 6 (median 4; range 1-31) previous operations . In all cases patients received osteosynthesis as well as a defect filling with RIA (reamer-irrigator-aspirator), and growth factor BMP-7 (bone morphogenetic protein-7). In 13 cases (23.6 %) there was no therapeutic success. In the evaluation of the SF12 questionnaire the mental health score increased from 47.4 (49.1; 27.6-65.7) to 49.8 (53.0; 28.7-69.4) and the well-being score from 32.7 (32.7;16.9-55.7) to 36.6 (36.5; 24.6-55.9). CONCLUSION: The two-step bone grafting method in the Masquelet technique used for tibia non-unions according to the diamond concept is a promising treatment option. Its application for tibia shaft non-unions with large bone defects or infections means a high degree of safety for the patient.

Immunological characterization of the early human fracture hematoma.

The initial inflammatory phase of fracture healing is of great importance for the clinical outcome. We aimed to develop a detailed time-dependent analysis of the initial fracture hematoma. We analyzed the composition of immune cell subpopulations by flow cytometry and the concentration of cytokines and chemokines by bioplex in 42 samples from human fractures of long bones <72 h post-trauma. The early human fracture hematoma is characterized by maturation of granulocytes and migration of monocytes/macrophages and hematopoietic stem cells. Both T helper cells and cytotoxic T cells proliferate within the fracture hematoma and/or migrate to the fracture site. Humoral immunity characteristics comprise high concentration of pro-inflammatory cytokines such as IL-6, IL-8, IFNγ and TNFα, but also elevated concentration of anti-inflammatory cytokines, e.g., IL-1 receptor antagonist and IL-10. Furthermore, we found that cells of the fracture hematoma represent a source for key chemokines. Even under the bioenergetically restricted conditions that exist in the initial fracture hematoma, immune cells are not only present, but also survive, mature, function and migrate. They secrete a cytokine/chemokine cocktail that contributes to the onset of regeneration. We hypothesize that this specific microenvironment of the initial fracture hematoma is among the crucial factors that determine fracture healing.

Treatment of nonunions in fractures of the humeral shaft according to the Diamond Concept.

METHODS: Between 2005 and 2012, 50 patients (23 female, 27 male) with nonunion of the humeral shaft were included in this retrospective study. The mean age was 51.3 years (14 to 88). The patients had a mean of 1.5 prior operations (sd 1.2;1 to 8). All patients were assessed according to a specific risk score in order to devise an optimal and individual therapy plan consistent with the Diamond Concept. In 32 cases (64%), a change in the osteosynthesis to an angular stable locking compression plate was performed. According to the individual risk an additional bone graft and/or bone morphogenetic protein-7 (BMP-7) were applied. RESULTS: A successful consolidation of the nonunion was observed in 37 cases (80.4%) with a median healing time of six months (IQR 6). Younger patients showed significantly better consolidation. Four patients were lost to follow-up. Revision was necessary in a total of eight (16%) cases. In the initial treatment, intramedullary nailing was most common. DISCUSSION: The use of locking compression plates in combination with autologous cancellous bone graft has been shown to be a safe and effective treatment. In more complex cases, the use of the Masquelet technique and BMP-7 may be indicated at the first revision operation. TAKE HOME MESSAGE: Our results suggest the Diamond Concept is a successful treatment strategy for nonunions of the humeral shaft.

[Treatment of Mason type II radial head fractures by percutaneous reduction]. / Die Behandlung von Radiusköpfchenfrakturen Typ Mason II durch perkutane Drahtreposition.

BACKGROUND: The therapeutic algorithm for the treatment of Mason type II radial head fractures is still controversially discussed. This study describes the technique of percutaneous fracture reduction without additional internal fixation of the radial head as an alternative to open reduction and presents the results of the method. MATERIAL AND METHODS: The data from 26 out of 30 patients with a Mason type II radial head fracture who had been consecutively treated with percutaneous fracture reduction were evaluated retrospectively. The analysis comprised the disabilities of shoulder and hand (DASH) score, the Mayo elbow performance score (MEPS) and data from the radiological examinations. RESULTS: The average follow-up time was 21 months (range 6-47 months). In 22 cases (85 %) an anatomical reduction could be achieved, 2 cases (8 %) showed a complete redislocation of the fragment and 2 cases (8 %) a partial redislocation. The average DASH score was 5.6 points (range 0-56) and the average MEPS was 93.8 (range 60-100). Only 4 patients (15 %) reported persisting functional impairment with a DASH score >10. CONCLUSIONS: The method of percutaneous reduction of radial head fractures without additional internal fixation in Mason type II fractures has been demonstrated to be a good alternative to open reduction.

Local gentamicin application does not interfere with bone healing in a rat model.

For the prophylaxis and treatment of bony infections antibiotics are locally used. Since several decades antibiotics mixed with bone cement (methylmethacrylate) are successfully used in prosthetic surgery and a gentamicin coated tibial nail is approved in Europe for fracture stabilization. The goal of the present study was to investigate if gentamicin, locally applied from a polymeric coating of intramedullary nails, might interfere with the bone healing process. Female Sprague Dawley rats (n = 72) were used and the tibiae were intramedullary stabilized with Kirschner-wires (k-wires) after osteotomy. This model was established earlier and shows a delayed healing with a prolonged inflammatory reaction. The open approach is clinically more relevant compared to a closed one because it mimics the clinically critical case of an open fracture, which has a higher risk of infection. The k-wire was either coated with the polymer poly(d,l-lactide) (control group) or with 10% gentamicin incorporated into the polymer (gentamicin group). In vivo µCT analyses were performed at days 10, 28, 42, and 84 after osteotomy. Mechanical torsional testing and histological evaluation were done at the days of sacrifice: 28, 42, and 84. The µCT analyses revealed an increase in tissue mineral density (TMD) over the healing period in both groups. In the control group, the torsional stiffness and maximum load did not reach the values of the intact contralateral side at any time point. At day 84 the gentamicin treated tibiae, however, showed significantly better maximum load compared to the control group. The histology showed no bony bridging in the control, whereas in 2 of 5 calluses of the gentamicin group mineralized bridging occurred. Significantly more mineralized tissue was measured in the gentamicin group. This study shows that the local gentamicin application does not negatively interfere with the long term healing process. Local infection prophylaxis is effective without negative effects on bone healing.

Increase in soluble CD95L during subacute phases after human spinal cord injury: a potential therapeutic target.

STUDY DESIGN: A pilot study measuring the levels of serum-soluble CD95 ligand (CD95L) in eight spinal cord-injured patients. OBJECTIVES: To determine the soluble concentration of CD95L in spinal cord injury (SCI) patients after trauma. METHODS: We collected blood samples from eight patients with acute traumatic SCI. Soluble CD95L serum levels were determined using an enzyme-linked immunosorbent assay. American Spinal Injury Association (ASIA) was determined according to ASIA classification. The patients were monitored, and venous blood was drawn after arrival at the hospital on the 1st and 3rd day and during the 1st, 2nd, 4th, 8th and 12th weeks after trauma. RESULTS: The average patient age was 48.1 years (18-86 years). Three patients were paraplegic (two incomplete, one complete), five were quadriplegic (one complete, four incomplete). The serum concentration of soluble CD95L (sCD95L) decreased during the 1st week (41 ng(- l)) and increased after the 2nd week in all eight patients. It peaked during the 4th week (68.5 ng (- l)) and reached a plateau during the 12th week (76.2 ng (- l)). There are many possible explanations for not being able to detect a statistical significance, one of course being the small sample size. CONCLUSION: Promising results for anti-CD95L therapy have already been documented in lab studies with rodents. Anti-CD95L blocks the pro-apoptotic and proinflammatory activity of membrane-bound CD95L during the acute phase of SCI. We observed that sCD95L levels are elevated during the subacute and intermediate phases of SCI. It would be of great interest to study a larger group of patients to determine whether higher sCD95 levels are correlated with improved or impaired neurological outcome or with increasing levels of autoimmune components in peripheral blood.

Effect of ß-tricalcium phosphate coated with zoledronic acid on human osteoblasts and human osteoclasts in vitro.

The combination of a bone graft material with bisphosphonates (BPs) might be advantageous for an optimal balance of material resorption and stimulation of bone formation. This study investigated the effect of ß-tricalcium phosphate (ß-TCP) bone grafts coated with zoledronic acid (ZOL) on osteoblast-like cells and osteoclast-like cells (OLC). As a drug carrier, the polymer poly(D,L-lactide) was used and three different concentrations of ZOL were tested. ß-TCP coated with ZOL stimulated the production of osteocalcin (OC), osteoprotegerin, and sRANKL in osteoblast-like cells. The polymer coating alone caused a significant increase in collagen type 1 and OC production. OLC viability was inhibited and the tartrate-resistant acidic phosphatase isoform-5b was significantly decreased after cultivation on polymer-coated ß-TCP for 12 days. The three different concentrations of ZOL decreased cell viability and no TRAPiso-5b was detectable, indicating a strong reduction of the TRAPiso-5b after 12 days in culture. After 21 days in culture, only the higher ZOL concentrations significantly reduced cell viability and TRAPiso-5b. The results of this study show that coating of ß-TCP with ZOL has stimulating effects on osteoblast-like cells. Additionally, an inhibition of osteoclasts was seen. The combination of this bone grafting material with BPs might, therefore, be effective in the treatment of large bone defects.

Core decompression combined with implantation of a demineralised bone matrix for non-traumatic osteonecrosis of the femoral head.

INTRODUCTION: Core decompression is the standard surgical procedure in the treatment of early stage non-traumatic osteonecrosis of the femoral head (ONFH). However, there is still a debate whether decompression in combination with supplementary augmentation by bone grafts, growth factors, or cell implementation is superior to conventional decompression alone. This study evaluated patients after core decompression combined with an augmentation by a demineralised bone matrix, and particularly aimed to report long-term conversion rates to total hip replacement (THR). MATERIALS AND METHODS: 14 patients with 18 hips suffering from ONFH (Ficat stage I-IIB) underwent this surgical procedure. All patients underwent radiographic and MRI investigations at baseline and at follow-up periods of 12 and 24 months. The clinical follow-up was done using the Merle d'Aubigné-score for an average period of 9 years after surgery. RESULTS: 14 of the 18 subjects (77 %) achieved at least a good clinical result after 2 years. The Merle d'Aubigné-score improved significantly after 12 (p = 0.0001) and 24 months (p = 0.0002). However, the MRI volumetric analysis showed an increased necrotic bone volume from 3.16 ± 0.54 to 3.88 ± 0.62 cm(3) (p = 0.04). Within 9 years, 13 out of 18 cases (72 %) required further surgery by THR. Only 7 out of 18 subjects (39 %) reported an ongoing postoperative clinical benefit, and would retrospectively redo the same surgical approach again. The five patients that did not require THR were still satisfied after 9 years. CONCLUSIONS: In patients with early- stage femoral head osteonecrosis core decompression combined with the implantation of a demineralised bone matrix leads to a limited, temporary pain relief as seen in core decompression alone. However, long-term results were not encouraging with a high rate of conversion to arthroplasty. Therefore, core decompression with implantation of a demineralised bone matrix may be not appropriate to avoid THR in the long term.

Local inhibition of angiogenesis results in an atrophic non-union in a rat osteotomy model.

Long bone and in particular tibia fractures frequently fail to heal. A disturbed revascularisation is supposed to be a major cause for impaired bone healing or the development of non-unions. We aim to establish an animal model, which reliably mimics the clinical situation. Human microvascular endothelial cells (HMEC-1) and primary human osteoblast like cells (POBs) were cultured with different angiogenesis-inhibitors (Fumagillin, SU5416, Artesunate and 3,5,4'-Trimethoxystilbene) released out of poly(D,L-Lactide) (PDLLA) coated k-wires and cell activity was determined. Discs containing PDLLA or PDLLA + Fumagillin/Artesunate were placed at the chorionallantoic membrane of hen eggs and the effect on vessel formation and egg vitality was observed. Tibia osteotomy was performed in rats and stabilised with K-wires coated with PDLLA + Fumagillin or with PDLLA only (control group). The healing was compared at different time points to the PDLLA control. Fumagillin and Artesunate inhibited the activity of HMEC-1 with minor effect on POBs. Artesunate caused embryonic death, whereas Fumagillin had no effects on egg vitality, but reduced the blood vessels. In the animal study all rats showed an impaired healing with reduced biomechanical stability. The Fumagillin treated tibiae had a significantly decreased callus size at day 42 and 84, less blood vessels in the early callus, a reduced histological callus size at day 10, 28 and 84, as well as an altered callus composition. This study presents a less vascularised, atrophic, tibia non-union and can be used in further investigations to analyse the pathology of atrophic non-union and to test new interventions.

Immunologically restricted patients exhibit a pronounced inflammation and inadequate response to hypoxia in fracture hematomas.

For patients who are known to have an impaired immune system, bone healing is often impaired. Therefore, it has been suggested that an effectively functioning immune system will have an influence on the quality of bone healing. Here, we demonstrate that cells within the fracture hematoma of immunologically restricted patients (1) exhibit a disturbed osteogenic differentiation (normal SPP1 but diminished RUNX2 expression), (2) show a strong inflammatory reaction (high IL8 and CXCR4), and (3) react on local hypoxia (high expression of HIF1A) but with inadequate target gene responses (diminished LDHA and PGK1 expression). Thus, it is already within the early inflammatory phase of fracture healing that the local gene expression in fracture hematomas of immunologically restricted patients points toward a critical regeneration.

Characterization of tendon cell cultures of the human rotator cuff.

Rotator cuff tears are common soft tissue injuries of the musculoskeletal system that heal by formation of repair tissue and may lead to high retear rates and joint dysfunction. In particular, tissue from chronic, large tendon tears is of such degenerative nature that it may be prone to retear after surgical repair. Besides several biomechanical approaches, biologically based strategies such as application of growth factors may be promising for increasing cell activity and production of extracellular tendon matrix at the tendon-to-bone unit. As a precondition for subsequent experimental growth factor application, the aim of the present study was to establish and characterize a human rotator cuff tendon cell culture. Long head biceps (LHB)- and supraspinatus muscle (SSP)- tendon samples from donor patients undergoing shoulder surgery were cultivated and examined at the RNA level for expression of collagen type-I, -II and -III, biglycan, decorin, tenascin-C, aggrecan, osteocalcin, tenomodulin and scleraxis (by Real-time PCR). Finally, results were compared to chondrocytes and osteoblasts as control cells. An expression pattern was found which may reflect a human rotator cuff tenocyte-like cell culture. Both SSP and LHB tenocyte-like cells differed from chondrocyte cell cultures in terms of reduced expression of collagen type-II (p

In vitro testing of the osteoinductive potential of different bony allograft preparations.

INTRODUCTION: Bony allografts are used frequently in the clinic for bone defect filling, however, less comparative data concerning their osteoinductive potential are available. AIM: The purpose of the present study was the comparative analysis of different allograft preparations. From five donors, we investigated fresh-frozen cancellous bone (native), peracetic acid­ethanol sterilized (PES) cancellous bone, cortical bone and demineralised bone matrix (DBM). In addition, two commercially available DBM products from five different donors were analyzed: Allomatrix® (Wright Medical Technology Inc.) and DBX putty® (Synthes GmbH). For positive control and as a clinically used growth factor, BMP-2 was chosen. METHOD: To investigate the osteoinductivity C2C12 cells were cultured with the different materials and the effect on cell proliferation and alkaline phosphatase activity were measured. RESULT: Proliferation was significantly enhanced by the native cancellous bone, Allomatrix, and BMP-2 and decreased by the PES-processed cancellous bone. The osteogenic differentiation was significantly enhanced by BMP-2 and the two commercial DBM products and decreased by PES-sterilized cancellous bone. All tested materials revealed a high donor-dependent variability. This is the first comparative study on the osteoinductivity of bony allografts frequently used in clinic.

Stabilisation of vertical unstable distal clavicular fractures (Neer 2b) using locking T-plates and suture anchors.

Distal clavicular fractures are associated with an increased risk of delayed union and non-union, and therefore operative treatment is favoured. Fragment dislocation and instability result from detachment of the coracoclavicular ligaments. Various methods for operative treatment can be found in the literature, but no gold standard has been established. In this retrospective study, we present a new surgical technique using a locking T-plate for osseous stabilisation in combination with vertical stabilisation by suture anchors. Between October 2006 and December 2007, eight people underwent surgery for unstable distal clavicular fracture (Neer type 2b). Subsequently one patient could only be contacted by phone and was excluded from further analysis. Mean follow-up for the remaining seven individuals was 8.3 months. The Constant and DASH scores were evaluated and stress radiographs were performed to check for vertical instability. In all cases bony union was achieved within 6 weeks postoperatively. No intraoperative or early postoperative complications were observed. All but one patient regained excellent shoulder function, the mean Constant and DASH scores were 93.3 and 15.3, respectively. Coracoclavicular distance was successfully restored with a mean 1mm (range 0-2mm) side-to-side difference. Early clinical and radiographic results of this new method are promising, with good to excellent outcome in all cases.

Cocultures of osteoblasts and osteoclasts are influenced by local application of zoledronic acid incorporated in a poly(D,L-lactide) implant coating.

The antiresorptive activity of bisphosphonates such as zoledronic acid (ZOL) has been shown in vitro to be because of their effect on osteoclasts and osteoblasts. However, whether the effect of ZOL on monocultures might be reproducible on cocultures and whether cell interactions might influence this effect has not been described. The aim of the study was to investigate the effect of ZOL on cocultures of osteoblasts and osteoclasts in vitro. ZOL was incorporated in an implant coating based on poly(D,L-lactide) in different concentrations (10-50 microM). Cell number was measured, and procollagen I synthesis, osteoprotegerin (OPG) secretion and soluble receptor activator of nuclear factor-kappaB ligand (sRANKL) were analyzed. Moreover, TRAP-positive cells and resorption lacunas on dentin chips were counted. Results showed that cell viability was not affected when treated with doses equivalent up to 50-microM ZOL-coated implants (ZOL-CI). Procollagen I and OPG synthesis was highest when treated with 10 microM ZOL-CI, whereas sRANKL showed no significant decrease when treated with the investigated concentrations of ZOL-CI. TRAP-positive cells were decreased when treated with ZOL-CI in a dose-dependent manner. Resorption activity of osteoclasts was not significantly decreased when treated with investigated concentrations of ZOL-CI. Exposure to specific concentrations of ZOL-CI showed a beneficial effect on osteoblast differentiation and protein synthesis. Formation of osteoclast was decreased, whereas a significant decrease in sRANKL secretion and resorption activity of osteoclasts could not be shown. The investigated effect on cocultures might be clinically useful to support fracture healing and to reduce orthopedic implant loosening.

Effect of mechanical stimulation on osteoblast- and osteoclast-like cells in vitro.

Bone-forming osteoblasts and bone-resorbing osteoclasts play an important role during maintenance, adaptation and healing of bone, and both cell types are influenced by physical activity. The aim of the present study was to investigate the effect of a narrow mechanical stimulation window on osteoblast- and osteoclast-like cells. Primary human cells were cultured on a bone-like structure (dentine) and three-point bending with approximately 1,100 microstrain was applied to the dentine at varying frequencies (0.1 and 0.3 Hz) and duration (1, 3 and 5 min daily over 5 days) resulting in different patterns of mechanical stimulation of osteoblast- and osteoclast-like cells. The longest stimulation (5 min at 0.1 Hz) induced a significant increase in osteoblast alkaline phosphatase activity and a significant decrease in osteoprotegerin (OPG) production, and resulted in a significant increase in the soluble receptor activator of NF-kappaB ligand (sRANKL)/OPG ratio towards sRANKL in comparison to the unstimulated osteoblast-like cells. All stimulations caused a significant decrease in collagen type 1 synthesis. Stimulation for 1 min at 0.3 Hz decreased the fusion and resorption activity of the osteoclast-like cells. These results demonstrate a direct effect of mechanical stimuli on osteoblast-like cells as well as on osteoclast formation and activity in vitro. The change in the sRANKL/OPG ratio towards the stimulation of osteoclastogenesis stresses the necessity to investigate the effect of the same stimulation parameter on the co-culture of both cell types.

Aprotinin application has no negative effect on osseous implant integration: a biomechanical and histomorphometric investigation in a rat model.

Intraoperative blood loss requiring allogenic blood transfusion (ABT) is a common problem in major orthopedic surgery. Since transfusion related side effects up to fatal consequences due to blood type incompatibility cannot be excluded completely, it is desirable to reduce the amount of blood loss and transfusions to a minimum. Encouraging results in the application of aprotinin, a natural protease-inhibitor with antifibrinolytic, bleeding-reducing properties, in thoracic-, heart- and abdominal surgery led to the use of aprotinin also in orthopedic surgery. One important safety issue in the use of aprotinin in orthopedic surgery is a possible negative effect on the osseous integration of an implant due to the multiple interactions of aprotinin with several enzymatic systems. In this study, we therefore investigated the influence of aprotinin on the osseous ingrowth of a titanium-implant in a rat model. Forty female Sprague-Dawley rats underwent unilateral retrograde nailing of the femur. Animals were divided in two groups, one receiving i.v. aprotinin intraoperatively, the other group receiving the same amount as saline solution. After 56 days animals were killed and from each group half of the femora were prepared for biomechanical testing, the other half for histological examination. The push-out experiment revealed no significant difference between the aprotinin-group and the control-group, both showing comparable shear stresses. In addition, the histomorphometrical analysis showed comparable implant integration between both groups. The results demonstrate that perioperative aprotinin application has no negative effect on osseous implant integration in a rat model.

The effect of zoledronic acid incorporated in a poly(D,L-lactide) implant coating on osteoblasts in vitro.

Bisphosphonates such as zoledronic acid (ZOL) are used in diseases associated with osteoclast-mediated bone loss. However, their antiresorptive activity is partly due to their effect on osteoblasts. Local application might increase the therapeutical fence and their local efficiency and reduce systemic side effects. Aim of the study was to investigate the effect of ZOL on human osteoblasts like cells in vitro with special focus on the synthesis of factors mediating osteoclast differentiation (RANKL, OPG). ZOL was incorporated in an implant coating based on poly(D,L-lactide) (PDLLA) in different concentrations (10-150 microM). Control groups were treated with uncoated implants, PDLLA-coated implants, and ZOL pure substance in corresponding concentrations. After an experimental period of 144 h, primary human osteoblasts were stained with alamar blue and cell viability was measured. Procollagen I synthesis, osteoprotegerin (OPG) secretion, and soluble receptor activator of nuclear factor-kappaB ligand (sRANKL) were analyzed. Results showed that cell viability was not affected when treated with doses equivalent up to 100 microM ZOL-coated implants (ZOL-CI). Procollagen I synthesis was highest when treated with 50 microM ZOL-CI. OPG increased significantly in the 10 microM ZOL-CI group, whereas sRANKL decreased significantly with different concentrations of ZOL-CI. Higher concentrations or exposure to the pure substance showed a decrease in cell viability, collagen I, OPG, and sRANKL synthesis. In conclusion, exposure to specific concentrations of ZOL-CI showed a beneficial effect on osteoblast differentiation and protein synthesis without influencing their proliferation. Changes in sRANKL and OPG production may contribute to the inhibition of osteoclastic bone resorption. This local antiresorptive effect might be clinically useful in osseous implant integration and fracture healing.

Quantification of various growth factors in different demineralized bone matrix preparations.

Besides autografts, allografts, and synthetic materials, demineralized bone matrix (DBM) is used for bone defect filling and treatment of non-unions. Different DBM formulations are introduced in clinic since years. However, little is known about the presents and quantities of growth factors in DBM. Aim of the present study was the quantification of eight growth factors important for bone healing in three different "off the shelf" DBM formulations, which are already in human use: DBX putty, Grafton DBM putty, and AlloMatrix putty. All three DBM formulations are produced from human donor tissue but they differ in the substitutes added. From each of the three products 10 different lots were analyzed. Protein was extracted from the samples with Guanidine HCL/EDTA method and human ELISA kits were used for growth factor quantification. Differences between the three different products were seen in total protein contend and the absolute growth factor values but also a large variability between the different lots was found. The order of the growth factors, however, is almost comparable between the materials. In the three investigated materials FGF basic and BMP-4 were not detectable in any analyzed sample. BMP-2 revealed the highest concentration extractable from the samples with approximately 3.6 microg/g tissue without a significant difference between the three DBM formulations. In DBX putty significantly more TGF-beta1 and FGFa were measurable compared to the two other DBMs. IGF-I revealed the significantly highest value in the AlloMatrix and PDGF in Grafton. No differences were accessed for VEGF. Due to the differences in the growth factor concentration between the individual samples, independently from the product formulation, further analyzes are required to optimize the clinical outcome of the used demineralized bone matrix.

Use of bone morphogenetic proteins for treatment of non-unions and future perspectives.

Still a major problem in orthopedic and trauma surgery is the delayed healing or the non-union of long bone fractures. Demographic data reveal that due to the steadily rising age of the population, complications with the musculoskeletal system will increase during the next years. Bone morphogenetic proteins (BMPs) have successfully been applied in clinic for the treatment of delayed healing and non-unions. The broad difference concerning the indication, timing of treatment, dosage and application technique of BMPs calls for the need to perform further prospective studies in order to standardize the treatment and furthermore optimize the procedures or even develop new therapeutic strategies. For example, the application technique may be improved and in some cases injectable BMP preparations could be of use. Also the coating of implants with growth factors might be valuable in order to stimulate bone healing and to prevent delayed healing or non-union. This article tries to discuss some of the open questions, however can and will not reflect the absolute standard of care. To make the BMP treatment a standard of care, more clinical data and long time experiences are necessary. The intramedullary application of BMP in combination with autologous or allogenic bone grafts or bone substitutes after debridement and stabilization with implants seems to be an adequate procedure for treatment of atrophic non-unions. However, the total number of patients is too small to draw final conclusions. Further clinical studies need to be performed in the future.