Different composition of leucocytes in cortical and cancellous bone healing in a mouse model.

OBJECTIVES: Cortical and cancellous bone healing processes appear to be histologically different. They also respond differently to anti-inflammatory agents. We investigated whether the leucocyte composition on days 3 and 5 after cortical and cancellous injuries to bone was different, and compared changes over time using day 3 as the baseline. METHODS: Ten-week-old male C56/Bl6J mice were randomized to either cancellous injury in the proximal tibia or cortical injury in the femoral diaphysis. Regenerating tissues were analyzed with flow cytometry at days 3 and 5, using panels with 15 antibodies for common macrophage and lymphocyte markers. The cellular response from day 3 to 5 was compared in order to identify differences in how cancellous and cortical bone healing develop. RESULTS: Between day 3 and 5, the granulocytes increased in the cancellous model, whereas the lymphocytes (T cells, B cells, NK cells) and monocytes (CD11b+, F4/80+, CD206+, CD14+) increased in the cortical model. CONCLUSION: These results suggest an acute type of inflammation in cancellous bone healing, and a more chronic inflammation in cortical healing. This might explain, in part, why cancellous healing is faster and more resistant to anti-inflammatory drugs than are diaphyseal fractures.Cite this article: L. Tätting, O. Sandberg, M. Bernhardsson, J. Ernerudh, P. Aspenberg. Different composition of leucocytes in cortical and cancellous bone healing in a mouse model. Bone Joint Res 2018;7:620-628. DOI: 10.1302/2046-3758.712.BJR-2017-0366.R2.

Undisturbed local bone formation capacity in patients with atypical femoral fractures: a case series.

We excised the fracture site in 8 patients with incomplete atypical femoral fractures by drilling an 11-mm-diameter hole. New bone formation could be seen in the hole within a normal time frame. Delayed healing of these fractures might be unrelated to an impaired capacity to form bone. INTRODUCTION: Incomplete atypical femoral fractures (undisplaced cracks) heal slowly or not at all, and often progress to a complete fracture with minimal trauma. The impaired healing has been attributed to an impaired biologic healing capacity related to bisphosphonate use, or, alternatively, to the mechanical environment within the fracture crack. This study aimed to investigate the capacity for bone formation after resection of the fracture site. METHODS: Between 2008 and 2014, we recruited eight patients with incomplete atypical femoral fractures. All used oral bisphosphonates before the fracture for on average 8 years (range 4 to 15) and complained of thigh pain. The fractures were stabilized with reamed cephalomedullary nails. During surgery, the fracture site in the lateral cortex was resected with a cylindrical drill (diameter 11.5 mm). The cylindrical cortical defect allowed radiographic evaluation of new bone formation, and the patients were followed clinically and radiologically for 24 months (range 15 to 92). RESULTS: After 3 months, newly formed bone could be seen in the cortical defects in all patients. After 13-26 months, the previous defects showed continuous cortical bone. At final follow-up, all patients reported full recovery of pre-surgical complaints. No complications occurred and no reoperations were performed. CONCLUSIONS: New bone formation occurred within a time frame that appears normal for healing of cortical bone defects. This suggests that the capacity to form new bone is intact.

Local bisphosphonate reduces migration and formation of radiolucent lines adjacent to cemented acetabular components.

AIMS: Post-operative migration of cemented acetabular components as measured by radiostereometric analysis (RSA) has a strong predictive power for late, aseptic loosening. Also, radiolucent lines predict late loosening. Migration has been reduced by systemic bisphosphonate treatment in randomised trials of hip and knee arthroplasty. Used as a local treatment, a higher local dose of bisphosphonate can be achieved without systemic exposure. We wished to see if this principle could be applied usefully in total hip arthroplasty (THA). PATIENTS AND METHODS: In this randomised placebo-controlled, double-blinded trial with 60 participants, we compressed gauze soaked in bisphosphonate solution (ibandronate) or saline against the acetabular bone bed immediately before cementing the acetabular component. RSA, classification of radiolucent lines, the Harris Hip Score (HHS) and the Western Ontario McMasters Universities Osteoarthritis Index (WOMAC) were carried out at three-, six-, 12-, and 24-month follow-up. RESULTS: Migration of the cemented acetabular component relative to the pelvis was reduced by movement almost half in the ibandronate group, when measured as maximum total point or as movement of the femoral head (p = 0.001 and 0.004, respectively). Radiolucent lines after one year were classified as absent, partial or complete, and correlated with treatment (rho 0.37; p = 0.004). Only three of 30 patients in the ibandronate group had complete lines, compared with 13 of 28 in the placebo group (p = 0.002). There were no significant effects on HHS or WOMAC score. CONCLUSION: Considering the power of RSA to predict loosening of cemented acetabular components, and the likelihood that radiolucent lines indicate risk of loosening, these data suggest that local treatment with a bisphosphonate can reduce the risk of late aseptic loosening. Cite this article: Bone Joint J 2017;99-B:317-24.

Randomised trial of bisphosphonate-coated dental implants: Radiographic follow-up after five years of loading.

The results of a randomised trial with bisphosphonate-coated dental implants have been reported previously. Each patient received one coated and one uncoated implant in a double-blind split-mouth design study. After 6 months of osseointegration, resonance frequency analysis indicated better fixation of the coated implants. Reduced marginal bone resorption was also shown. However, it was not known whether the advantage of the bisphosphonate coating would persist over time. The radiographic results at 5 years after implant installation are reported herein. A blinded investigator measured marginal resorption on fresh radiographs obtained for 14 of the 16 patients (two had died) and compared these with the post-implantation images. Non-parametric statistics were used. All implants functioned well. The median marginal bone loss for control implants was found to be 0.70mm, which is less than usually reported in the literature. The bisphosphonate-coated implants showed even less resorption (median 0.20mm). The median difference within each pair of implants after 5 years of use was 0.34mm (95% confidence interval 0.00-0.75mm; P=0.04). The present data suggest that bisphosphonate-coated implants enable prolonged preservation of the marginal bone.

Glucocorticoids inhibit shaft fracture healing but not metaphyseal bone regeneration under stable mechanical conditions.

OBJECTIVES: Healing in cancellous metaphyseal bone might be different from midshaft fracture healing due to different access to mesenchymal stem cells, and because metaphyseal bone often heals without a cartilaginous phase. Inflammation plays an important role in the healing of a shaft fracture, but if metaphyseal injury is different, it is important to clarify if the role of inflammation is also different. The biology of fracture healing is also influenced by the degree of mechanical stability. It is unclear if inflammation interacts with stability-related factors. METHODS: We investigated the role of inflammation in three different models: a metaphyseal screw pull-out, a shaft fracture with unstable nailing (IM-nail) and a stable external fixation (ExFix) model. For each, half of the animals received dexamethasone to reduce inflammation, and half received control injections. Mechanical and morphometric evaluation was used. RESULTS: As expected, dexamethasone had a strong inhibitory effect on the healing of unstable, but also stable, shaft fractures. In contrast, dexamethasone tended to increase the mechanical strength of metaphyseal bone regenerated under stable conditions. CONCLUSIONS: It seems that dexamethasone has different effects on metaphyseal and diaphyseal bone healing. This could be explained by the different role of inflammation at different sites of injury. Cite this article: Bone Joint Res 2015;4:170-175.

Osteoporosis: the emperor has no clothes.

Current prevention strategies for low-trauma fractures amongst older persons depend on the notions that fractures are mainly caused by osteoporosis (pathophysiology), that patients at high risk can be identified (screening) and that the risk is amenable to bone-targeted pharmacotherapy (treatment). However, all these three notions can be disputed. PATHOPHYSIOLOGY: Most fracture patients have fallen, but actually do not have osteoporosis. A high likelihood of falling, in turn, is attributable to an ageing-related decline in physical functioning and general frailty. SCREENING: Currently available fracture risk prediction strategies including bone densitometry and multifactorial prediction tools are unable to identify a large proportion of patients who will sustain a fracture, whereas many of those with a high fracture risk score will not sustain a fracture. TREATMENT: The evidence for the viability of bone-targeted pharmacotherapy in preventing hip fracture and other clinical fragility fractures is mainly limited to women aged 65-80 years with osteoporosis, whereas the proof of hip fracture-preventing efficacy in women over 80 years of age and in men at all ages is meagre or absent. Further, the antihip fracture efficacy shown in clinical trials is absent in real-life studies. Many drugs for the treatment of osteoporosis have also been associated with increased risks of serious adverse events. There are also considerable uncertainties related to the efficacy of drug therapy in preventing clinical vertebral fractures, whereas the efficacy for preventing other fractures (relative risk reductions of 20-25%) remains moderate, particularly in terms of the low absolute risk reduction in fractures with this treatment.

Trigger finger, tendinosis, and intratendinous gene expression.

The pathogenesis of trigger finger has generally been ascribed to primary changes in the first annular ligament. In contrast, we recently found histological changes in the tendons, similar to the findings in Achilles tendinosis or tendinopathy. We therefore hypothesized that trigger finger tendons would show differences in gene expression in comparison to normal tendons in a pattern similar to what is published for Achilles tendinosis. We performed quantitative real-time polymerase chain reaction on biopsies from finger flexor tendons, 13 trigger fingers and 13 apparently healthy control tendons, to assess the expression of 10 genes which have been described to be differently expressed in tendinosis (collagen type 1a1, collagen 3a1, MMP-2, MMP-3, ADAMTS-5, TIMP-3, aggrecan, biglycan, decorin, and versican). In trigger finger tendons, collagen types 1a1 and 3a1, aggrecan and biglycan were all up-regulated, and MMP-3and TIMP-3 were down-regulated. These changes were statistically significant and have been previously described for Achilles tendinosis. The remaining four genes were not significantly altered. The changes in gene expression support the hypothesis that trigger finger is a form of tendinosis. Because trigger finger is a common condition, often treated surgically, it could provide opportunities for clinical research on tendinosis.

Prevention of osteonecrosis of the jaw by mucoperiosteal coverage in a rat model.

There is evidence for a link between the use of systemic bisphosphonates and osteonecrosis of the jaw (ONJ). This condition has the appearance of chronic osteomyelitis, and antibiotics prevent the development of ONJ in animal models. Clinically, ONJ can sometimes be treated successfully by mucoperiosteal coverage. If ONJ is indeed primarily caused by bacterial infection, immediate coverage of the extraction alveolus might reduce the risk of ONJ developing in risk patients. Therefore, we studied whether immediate mucoperiosteal coverage after tooth extraction could prevent the development of ONJ in a rat model. Thirty rats were randomly allocated to three groups (10 in each): (1) group I (controls): extraction, no drug treatment; (2) group II (non-coverage): extraction, dexamethasone plus alendronate; (3) group III (coverage): extraction, dexamethasone plus alendronate, plus coverage with a mucoperiosteal flap. Rats were examined for macroscopic ONJ-like wounds after 2 weeks. All animals in the non-coverage group developed large ONJ-like changes. The coverage and control groups showed an intact overlying mucosa in all rats. Findings were confirmed with histology. Bisphosphonates and dexamethasone caused ONJ-like lesions after tooth extraction in a rat model. This was prevented by immediate mucoperiosteal coverage. The risk of ONJ in patients using bisphosphonates might be reduced by mucoperiosteal coverage after tooth extraction.

Effect of local vs. systemic bisphosphonate delivery on dental implant fixation in a model of osteonecrosis of the jaw.

Locally applied bisphosphonates may improve the fixation of metal implants in bone. However, systemic bisphosphonate treatment is associated with a risk of osteonecrosis of the jaw (ONJ). We hypothesized that local delivery of bisphosphonate from the implant surface improves the fixation of dental implants without complications in a setting where systemic treatment induces ONJ. Forty rats were randomly allocated to 4 groups of 10. All groups received a titanium implant inserted in an extraction socket. Group I received the implants only. Group II received dexamethasone (0.5 mg/kg). Group III received dexamethasone as above plus alendronate (200 µg/kg). Group IV received zoledronate-coated implants and dexamethasone as above. The animals were sacrificed 2 weeks after tooth extraction. All 10 animals with systemic alendronate treatment developed large ONJ-like changes, while all with local treatment were completely healed. Implant removal torque was higher for the bisphosphonate-coated implants compared with the other groups (p < 0.03 for each comparison). Micro-computed tomography of the maxilla showed more bone loss in the systemic alendronate group compared with groups receiving local treatment (p = 0.001). Local bisphosphonate treatment appears to improve implant fixation in a setting where systemic treatment caused ONJ.

Atypical fractures do not have a thicker cortex.

UNLABELLED: An association between atypical fractures and general cortical thickness of the femoral shaft is often suggested in the literature. Our radiographic measurements of 59 atypical and 218 ordinary fractures now exclude a difference larger than 10 % in mean femoral cortical thickness (sum of lateral and medial) with 95 % confidence. INTRODUCTION: An increased general cortical thickness in patients with fatigue fracture of the femoral shaft (atypical fractures) is commonly suggested. However, there are scarce data to support this. METHODS: In a published nationwide Swedish study, we identified by radiographic review 59 women with an atypical fracture during 2008. The femoral cortical thickness index (thickness/femoral diameter) of these women was now compared with the 218 ordinary fractures that occurred in the same region of the femur in a case-control design. The cortical thickness index 5 cm below the lesser trochanter was the primary variable. RESULTS: Patients with atypical fractures were younger. Without correction for age, they had a thicker cortex (i.e., higher index). However, the difference in cortical thickness disappeared after age correction. The 95 % CI excludes a group mean difference exceeding 10 % of total mean thickness. Similarly, there was no significant difference in cortical thickness between patients with or without bisphosphonate treatment or between the ipsi- and contralateral femurs in patients with an atypical fracture. CONCLUSION: The concept of a generally increased cortical thickness in patients with atypical fractures should be reconsidered.

Growth hormone does not stimulate early healing in rat tendons.

Growth Hormone stimulates bone growth and fracture repair. It acts mainly by increasing the systemic levels of IGF-1. Local treatment with IGF-1 appears to stimulate tendon healing. We therefore hypothesized that systemic treatment with Growth Hormone would also stimulate tendon healing. Rat Achilles tendons were transected and left to heal. 4 groups were studied. Intramuscular injections of botulinum toxin A (Botox) were used to reduce loading in 2 groups. The animals were randomized to twice daily injections of Growth Hormone (n=2×10) or saline (n=2×10), and killed after 10 days. Healing was assessed by mechanical testing. Muscle paralysis induced by Botox reduced the strength of the healing tendon by two thirds. Growth Hormone increased femoral and tibial length in the unloaded, and femoral and tibial weight in the loaded group. Body weight and muscle weight were increased in both. In contrast, there was no increase in the strength of the healing tendons, regardless of mechanical loading status. An increase in peak force of the loaded healing tendons by more than 5% could be excluded with 95% confidence. In spite of its stimulatory effects on other tissues, Growth Hormone did not appear to stimulate tendon or tendon repair.

Trigger finger and tendinosis.

The pathogenesis of trigger finger has generally been ascribed to primary changes in the pulley. Histological examination of the affected tendons has rarely been done. We studied biopsies from tendons of trigger fingers from 29 patients and compared these to biopsies from six intact tendons. We used a modified Movin score, which describes the tendinosis of the Achilles tendon. Trigger finger tendons had a high score (14.2; SD, 2.2) consistent with tendinosis, while the controls were almost normal (2.5; SD, 1.9). This suggests that the tendon is also affected, and that trigger finger is a form of tendinosis.

Early E-modulus of healing Achilles tendons correlates with late function: similar results with or without surgery.

Non-operative treatment of Achilles tendon ruptures is associated with an increased risk of rerupture. We hypothesized that this is due to inferior mechanical properties during an early phase of healing, and performed a randomized trial, using a new method to measure the mechanical properties. Tantalum markers were inserted in the tendon stumps, and tendon strain at different loadings was measured by stereo-radiography (Roentgen stereophotogrammetric analysis) at 3, 7 and 19 weeks and 18 months after injury. Thirty patients were randomized to operative or non-operative treatment. The primary out-come variable was an estimate for the modulus of elasticity at 7 weeks. Strain per force, cross-sectional area and tendon elongation were also measured. The functional outcome variable was the heel-raise index after 18 months. There was no difference in the mean modulus of elasticity or other mechanical or functional variables between operative and non-operative treatments at any time-point, but strain per force at 7 and 19 weeks had a significantly larger variation in the non-operative group. This group, therefore, might contain more outliers with poor healing. The modulus of elasticity at 7 weeks correlated with the heel-raise index after 18 months in both treatment groups (r(2) =0.75; P=0.0001). This correlation is an intriguing finding.

Local bisphosphonate release versus hydroxyapatite coating for stainless steel screw fixation in rat tibiae.

Implant fixation in bone can be improved by a coating that delivers bisphosphonates locally, or by a hydroxyapatite (HA) coating. In this study, we compared these different types of coatings. For mechanical testing, 30 rats were assigned into three groups, and similar screws were implanted bilaterally in the proximal tibiae. The rats received screws that were either uncoated, coated with nano-crystalline hydroxyapatite or coated with a bisphosphonate releasing protein matrix. After 4 weeks, one screw was subjected to pull-out testing, and the contra-lateral one to torsion testing. For morphology, 30 rats were assigned to similar treatment groups, but received only one screw each. Bisphosphonates enhanced the pull-out force by 41% (P = 0.02) compared to controls, HA increased the pull-out force although not significantly. Conversely, HA increased the maximal torque by 64% (P = 0.02). Morphometry showed higher bone volume around bisphosphonate screws in comparison to HA-coated screws (P < 0.001) and controls (P < 0.001). The results suggest that bisphosphonates improve fixation by increasing the amount of surrounding bone, whereas HA mainly improves bone to implant attachment.

Bisphosphonate coating might improve fixation of dental implants in the maxilla: a pilot study.

This pilot study evaluates the clinical stability of bisphosphonate-coated dental implants placed using a two-stage surgical procedure in five patients. Each patient received seven regular Brånemark implants, one of which was coated with bisphosphonate in a fibrinogen matrix. The coated implant was inserted where the bone was expected to have the least favourable quality. The level of the marginal bone around each implant was measured by intraoral periapical radiographs and implant stability was recorded using resonance frequency measurements. Frequency values (ISQ) were obtained peroperatively before flap closure and after 6 months at abutment connection. At abutment connection the bisphosphonate-coated implants were removed en bloc in two patients for histological examination. An animal experiment had previously confirmed that gamma-sterilization did not reduce bioactivity of the bisphosphonate coating. In each patient, the bisphosphonate-coated implant showed the largest improvement in ISQ level of all implants. Their values at the start tended to be lower, and the absolute value at 6 months did not differ. No complications occurred with the coated implants. Histology showed no abnormalities. Improvement in ISQ values was an expected effect of the bisphosphonate coating, but could be due to the choice of insertion site. This finding warrants a randomized, blinded study.

Evidence for anti-osteoporosis therapy in acute fracture situations--recommendations of a multidisciplinary workshop of the International Society for Fracture Repair.

The International Society for Fracture Repair convened a multidisciplinary workshop to assess the current evidence around the interaction between anti-osteoporosis drugs and the healing of incident fractures, with a view to making recommendations for clinical practice. The consensus was that there is no evidence-based reason to withhold anti-resorptive therapy while a fracture heals, whether or not the patient was taking such therapy when the fracture occurred. The workshop also considered existing models of service provision for secondary prevention and concluded that the essential ingredient for reliable delivery is the inclusion of a dedicated coordinator role. Several unresolved issues were defined as subjects for further research, including the question of whether continuous long-term administration of anti-resorptives may impair bone quality. The rapidly changing area requires re-assessment of drugs and their interaction with fracture healing in the near future.

Elevated intraperitoneal matrix metalloproteinases-8 and -9 in patients who develop anastomotic leakage after rectal cancer surgery: a pilot study.

OBJECTIVE: Experimental studies suggest that matrix metalloproteinase (MMP) enzymes mediate the early tissue breakdown that leads to a decrease in intestinal anastomotic strength. Patients with upregulation of MMPs in intestinal biopsies have an increased rate of anastomotic leakage. We measured MMPs and their inhibitors [tissue inhibitors of metalloproteinases (TIMPs)] in postoperative intraperitoneal fluid after rectal cancer surgery, and hypothesized that they would be elevated in patients who later would develop anastomotic leakage. METHOD: Twenty-nine patients with rectal carcinoma underwent low anterior resection of the rectum for cancer. Intraperitoneal fluid was collected via a pelvic drain at a median of 4 h postoperatively. MMP-1, -2, -3, -7, -8, -9 and -13 were determined using particle-based multiplex flow-cytometry. TIMP-1 and -2 were measured by enzyme-linked immunosorbent assays. MMP-9 was considered the main outcome variable. RESULTS; Ten patients developed anastomotic leakage. These patients had increased intraperitoneal MMP-9 [median difference (m.d.) 29%; P = 0.03] and MMP-8 (m.d. 58%; P = 0.02), compared with patients who did not develop leakage. There were no differences between the groups for other MMPs and TIMPs. CONCLUSION: Matrix metalloproteinase-8 and -9 appear to have an important role in the development of anastomotic leakage and may be promising pharmacological targets to protect anastomotic integrity. We suggest further investigation of MMPs as markers for anastomotic leakage.

Elevation of systemic matrix metalloproteinases 2 and 7 and tissue inhibitor of metalloproteinase 2 in patients with a history of Achilles tendon rupture: pilot study.

OBJECTIVES: In this study, serum levels of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) between patients with a history of Achilles tendon rupture and blood donor controls were compared, and their relation to mechanical properties of the tendons during healing were studied. METHODS: More than 3 years after injury, serum levels of MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9 and MMP-13, TIMP-1 and TIMP-2 in eight patients who had Achilles tendon rupture were measured. Twelve blood donors served as controls. During the early phase of healing, the tendon modulus of elasticity was calculated from radiostereometric data and tendon cross-sectional area. RESULTS: Patients with a history of Achilles tendon rupture had increased levels of MMP-2 (median difference 10%, p=0.01), MMP-7 (median difference 15%, p=0.02) and TIMP-2 (median difference 36%, p=0.02), compared with controls. Levels of MMP-7, measured 3 years after injury, correlated inversely to tendon modulus of elasticity (r(s)=20.83, p=0.02) and positively to tendon elongation (r(s)=0.74, p=0.05) during the early phase of healing. There was a trend towards positive correlation between MMP-7 and cross-sectional area during the early phase of healing (r(s)=0.67, p=0.08). CONCLUSIONS: Patients with a history of Achilles tendon rupture appear to have elevated levels of MMP-2, MMP-7 and TIMP-2 in serum. In these pilot data, the view that the MMP-TIMP system is involved in tendinopathy is supported and that disturbances in proteolytic control might be generalised are suggested.

Experimental results of combining bisphosphonates with allograft in a rat model.

Soaking bone grafts in a bisphosphonate solution before implantation can prevent their resorption and increase the local bone density in rats and humans. However, recent studies suggest that pre-treatment of allografts with bisphosphonate can prevent bone ingrowth into impaction grafts. We tested the hypothesis that excessive amounts of bisphosphonate would also cause a negative response in less dense grafts. We used a model where non-impacted metaphyseal bone grafts were randomised into three groups with either no bisphosphonate, alendronate followed by rinsing, and alendronate without subsequent rinsing, and inserted into bone chambers in rats. The specimens were evaluated histologically at one week, and by histomorphometry and radiology at four weeks. At four weeks, both bisphosphonate groups showed an increase in the total bone content, increased newly formed bone, and higher radiodensity than the controls. In spite of being implanted in a chamber with a limited opportunity to diffuse, even an excessive amount of bisphosphonate improved the outcome. We suggest that the negative results seen by others could be due to the combination of densely compacted bone and a bisphosphonate. We suggest that bisphosphonates are likely to have a negative influence where resorption is a prerequisite to create space for new bone ingrowth.