Callus Volume Research Articles

Interleukin-6 (IL-6) deficiency enhances intramembranous osteogenesis following stress fracture in mice.

Interleukin-6 (IL-6) is highly upregulated in response to skeletal injury, suggesting it plays a role in the inflammatory phase of fracture repair. However, the impact of IL-6 on successful repair remains incompletely defined. Therefore, we investigated the role of IL-6 in two models of fracture repair (full fracture and stress fracture) using 12-week old IL-6 global knockout mice (IL-6 KO) and wild type (WT) littermate controls. Callus morphology and mineral density 14days after full femur fracture did not differ between IL-6 knockout mice and controls. In contrast, IL-6 KO mice had an enhanced bone response 7days after ulnar stress fracture compared to WT, with increased total callus volume (p=0.020) and callus bone volume (p=0.045). IL-6 KO did not alter the recruitment of immune cells (Gr-1 or F4/80 positive) to the stress fracture callus. IL-6 KO also did not alter the number of osteoclasts in the stress fracture callus. Using RNA-seq, we identified differentially expressed genes in stress fracture vs. contralateral control ulnae, and observed that IL-6 KO resulted in only modest alterations to the gene expression response to stress fracture (SFx). Wnt1 was more highly upregulated in IL-6 KO SFx callus at both day 1 (fold change 12.5 in KO vs. 5.7 in WT) and day 3 (fold change 4.7 in KO vs. 1.9 in WT). Finally, using tibial compression to induce bone formation without bone injury, we found that IL-6 KO directly impacted osteoblast function, increasing the propensity for woven bone formation. In summary, we report that IL-6 knockout enhanced formation of callus and bone following stress fracture injury, likely through direct action on the osteoblast's ability to produce woven bone. This suggests a novel role of IL-6 as a suppressor of intramembranous bone formation.

Effect of Selective Androgen Receptor Modulator Enobosarm on Bone Healing in a Rat Model for Aged Male Osteoporosis

Enobosarm (ostarine, MK-2866, or GTx-024) is a non-steroidal selective androgen receptor modulator. This study evaluated the effect of various regimens of enobosarm (EN) on bone healing in an orchiectomized rat model for aged male osteoporosis and compared it to testosterone (T) treatment. Ninety eight-month-old male Sprague Dawley rats were either orchiectomized (Orx) or left intact (Non-Orx) and divided into groups (n = 15/group): (1) Non-Orx; (2) Orx; (3) Orx+T-th; (4) Orx+EN-th; (5) Orx+T-pr; and (6) Orx+EN-pr. Prophylaxis (Pr) treatments were applied immediately after Orx for up to 18 weeks. Therapy (Th) treatments were applied 12 weeks after Orx for up to 6 weeks. Bilateral tibia osteotomy with plate osteosynthesis was performed 12 weeks after Orx in all groups. EN and T were mixed with the diet; the daily dosage was 0.35 ± 0.06 and 41 ± 8 mg/kg BW, respectively. Both T treatments improved bone healing by increasing callus volume and area, bone volume and density, and cortical width; they had no effect on prostate or levator ani weight. EN-pr increased the callus area and callus density and decreased cortical density, but increased prostate weight. The effect of T-pr and T-th on bone was stronger than EN-pr. EN-th affected bone healing negatively by reducing callus density and area and delaying osteotomy bridging. Levator ani weight was increased in both EN groups. EN treatment after fracture is not advisable in aged males. EN-pr treatment as a therapy for bone healing in men could be further investigated; endocrinological side effects must be closely monitored.

Assessment of bone healing using (Ti,Mg)N thin film coated plates and screws: Rabbit femur model.

Magnesium (Mg) based implants such as plates and screws are often preferred to treat bone defects because of the positive effects of magnesium in bone growth and healing. Their low corrosion resistance, however, leads to fast degradation and consequently failure before healing was completed. Previously, we developed Mg doped titanium nitrate (TiN) thin film coatings to address these limitations and demonstrated that <10 at% Mg doping led to enhanced mineralization in vitro. In the present study, in vivo performance of (Ti,Mg)N coated Ti6Al4V based plates and screws were studied in the rabbit model. Bone fractures were formed on femurs of 16 rabbits and then fixed with either (Ti,Mg)N coated (n = 8) or standard TiN coated (n = 8) plates and screws. X-ray imaging and μCT analyses showed enhanced bone regeneration on fracture sites fixed with (Ti,Mg)N coated plates in comparison with the Mg free ones. Bone mineral density, bone volume, and callus volume were also found to be 11.4, 23.4, and 42.8% higher, respectively, in accordance with μCT results. Furthermore, while TiN coatings promoted only primary bone regeneration, (Ti,Mg)N led to secondary bone regeneration in 6 weeks. These results indicated that Mg presence in the coatings accelerated bone regeneration in the fracture site. (Ti,Mg)N coating can be used as a practical method to increase the efficiency of existing bone fixation devices of varying geometry.

Effect of kirenol on the interaction between the WNT/β-Catenin and RUNX2/TCF/LEF1 pathways in fracture healing in vivo.

This study aimed to determine the effects of a natural diterpenoid, kirenol, on fracture healing in vivo in an experimental rat model of femur fracture and investigate its potential mechanism of action via the Wnt/β-catenin pathway. In this study, 64 male Wistar albino rats aged 5-7 weeks and weighing 261-348 g were randomly divided into 8 groups from A to L, with eight rats in each group. Standardized fractures were created in the right femurs of the rats and then fixed with an intramedullary Kirschner wire. Four experimental groups were administered 2 mg/kg/day kirenol (Groups C and G) and 4 mg/kg/day (Groups D and H) kirenol by oral gavage.Thereafter, the animals were sacrificed at two time points as follows: on the 10th day (Groups B, C and D) and on the 21st day (Groups F, G and H) after the surgery; fracture healing in each group was assessed radiologically and histopathologically. The Radiographic Union scale of tibia fracture scoring system was used in the radiological examination; callus volume and density were measured using computed tomography. In the histopathologic examination, the scoring system described by Huo et al. was used. Additionally, the mechanism of action was evaluated based on the analyses of protein expression of Wnt3a, LRP5, TCF-LEF1, β-catenin, and Runx-2 proteins using western blot analysis. Among the animals sacrificed on the 10th day after the surgery, the highest histopathological and radiological scores were observed in Group D (p<0.05). Furthermore, the callus density (p<0.05) was highest in Group D. Among the animals sacrificed on the 21st day, the highest histopathological and radiological scores were found in Group H, although the differences among the groups were not significant (p>0.05). The callus volume and density were the highest in Groups G and H, respectively, although the differences among groups were not significant. Kirenol may improve fracture healing in a dose-dependent manner with the early activation of the Wnt/β-catenin pathway and the activation of the Runx-2 pathway.

Light Intensity-Induced Morphogenetic Response and Enhanced β-Sitosterol Accumulation in Date Palm (Phoenix dactylifera L. cv. Hayani) Callus Culture

The present study, for the first time, assessed the influence of light intensity on morphogenesis of callus cultures and accumulation of β-sitosterol, a key phytosterol compound in date palm. High frequencies of calli with maximum biomass were induced from shoot tip explants in Murashige and Skoog basal medium supplemented with 10 mg/l 2,4 dichlorophenoxyacetic acid + 3 mg/l 2-isopentyladenine. Such proliferating callus cultures of date palm cv. Hayani were incubated in specified medium under different light intensities [0, 14, and 42 µmol/m2/s photosynthetic photon flux density (PPFD)]. The collected data exhibited an increase in callus volume, globularization, newly differentiated somatic embryos, degree of browning, and β-sitosterol contents of callus cultures. Various effects of different light intensities on the above-mentioned traits were recorded. Morphogenesis of callus under dark condition (0 µmol/m2/s PPFD) resulted in a higher degree of globularization and produced maximum number of newly differentiated somatic embryos. Whereas, under gradually increasing light intensities (14 and 42 µmol/m2/s PPFD), a surge in the accumulation of β-sitosterol content in the proliferating callus was observed. Thus, from the present study, it can be deduced that a proper application of suitable light intensity on proliferating callus has beneficial influence on morphogenesis and a higher production of β-sitosterol in date palm.

Discussion on bone healing rate and expression of cytokines and growth factors in rat models with simple fracture and brain injury

This research aims to analyze the differences in bone healing rate, cytokines content, and growth factor content in rats with fracture and traumatic brain injury. In this study, rat models of tibial fracture (fracture group) and tibial fracture combined with brain injury (combined group) were first constructed. Callus and brain tissue sections of the two groups of rat models were made, and the pathological changes were analyzed by hematoxylin – eosin staining. Tissue sections were made to compare the differences in callus volume, trabecular width, and trabecular bone ratio between the two groups of rats. Taking normal rats as controls, serum samples were collected from rats in the control group, fracture group, and combined group. The differences of levels in nerve growth factor (NGF), insulin-like growth factor 1 (IGF1), bone morphogenetic protein 7 (BMP7), and transforming growth factor β1 (TGFβ1) in serum of different groups of rats were detected by enzyme-linked immunosorbent assay, and the expression of NGF, IGF 1, BMPR7, and TGFβR1 in rat callus was detected by Western-Blot. It was found that the rate of bone healing in the combined group was faster than that in the fracture group. The bone callus volume, trabecular width, and trabecular proportion of the rats in the combined group were significantly higher than those in the fracture group (P < 0.05). The content of NGF, IGF1, BMP7, and TGFβ1 in the serum of the fracture group and the combined group and the expression of their receptor proteins were significantly higher than that of the control group (P < 0.05), but the content of NGF, IGF1, BMP7, and TGF 1 in the serum and the expression of their receptor proteins of the combined group were significantly higher than that of the fracture group (P < 0.05). Accordingly, tibial fractures combined with brain injury can accelerate bone healing, which may be caused by regulating the expression of NGF, IGF1, BMP7, and TGFβ1 and accelerating the binding of its receptors.

Prognostic Significance of Alkaline Phosphatase Activity and Callus Volume in Operatively Treated Bone Fractures

Total and bone alkaline phosphatase are indicators of bone formation, a process essential in bone healing. The aim of this study was to assess the relationship of both total and bone alkaline phosphatase with the course and features of healing in surgically treated long bone fractures as compared to the callus volume. In this study, total and bone alkaline phosphatase levels and the callus volume were measured in two patients with long bone fractures. Fracture healing was rapid in one patient and slow in the other. Depending on the healing outcome, on day 7 an increase in the case of slow healing and a decrease in the case of rapid healing was noted for both total and bone alkaline phosphatase. In the case of slow healing, the callus volume was significant whereas in the case of rapid healing the callus was almost invisible on day 7. This result indicates a possible prognostic significance of aforementioned clinical biochemical and radiographic parameters in the monitoring of long bone fracture healing.

The Promotive Effects of PTHrP on the Union of Femoral Shaft Fracture in a Mouse Model

In order to explore the effects of PTH-related peptide on fracture healing in mice, the intervention effects and mechanism of action of PTH-related peptide on femoral shaft fracture models in mice of different genotypes were analysed to obtain a new approach for promoting the healing fractures clinically. A total of 32 8-week old PTH-related peptide wild-type and PTH-related peptide gene knockout heterozygous (PTH-related peptide +/-) mice were selected as the test system and standard femoral shaft fracture mouse models were constructed by intramedullary wire fixation. After model construction, all mice were randomly divided into the wild type control, the PTH-related peptide +/- control, the wild type treated and the PTH-related peptide +/- treated groups, with 8 mice in each group. Mice in both treated groups were administered with 80 μg/kg/day of PTH-related peptide (1-34) subcutaneously and the control groups received subcutaneous injections of saline. After 2 weeks, all mice were sacrificed and the fractured right femur of each mouse in each group was taken for X-ray examination and micro-CT scan to observe the union of the fracture. Next, the formed callus tissues were paraffin-embedded and stained to analyse and compare the bone formation changes of osteoblasts in the fracture sites of mice. Compared to the wild-type group, the saline-treated mice in the PTH-related peptide +/- group had clear fracture lines with callus volume and bone mineral density significantly reduced; after PTH-related peptide treatment, the fractures in the PTH-related peptide +/- group were tightly connected and a large number of osseous calluses were formed; in addition, the density of the callus was higher, which was significantly different from those in the control group (p<0.05). Compared to the wild-type group, the area of cartilage callus of the saline-treated PTH-related peptide +/- group increased by 97 % (p<0.05); after PTH-related peptide administration, the total callus area and osseous callus area increased significantly, while the area of cartilage osseous significantly decreased compared to the control group and after PTH-related peptide administration, the number of osteoblasts and the percent positive area of collagen I in mice were significantly increased compared to the control group. In conclusion, endogenous PTH-related peptide deficiency could lead to delayed healing of fracture, which could be compensated by PTH-related peptide administration, thereby accelerating the formation of calluses and promoting fracture healing in mice.

Angiotensin receptor blockade mimics the effect of exercise on recovery after orthopaedic trauma by decreasing pain and improving muscle regeneration.

Our tibial fracture orthopaedic injury model in mice recapitulates the major manifestations of complex trauma, including nociceptive sensitization, bone fracture, muscle fibrosis and muscle fibre loss. Delayed exercise after complex orthopaedic trauma results in decreased muscle fibrosis and improved pain Losartan, an angiotensin-receptor blocker with anti-fibrotic abilities, recapitulates the effect of exercise on post-injury recovery and may provide an enhanced recovery option for those who are unable to exercise after injury ABSTRACT: Chronic pain and disability after limb injury are major public health problems. Early mobilization after injury improves functional outcomes for patients, although when and how to implement rehabilitation strategies remains a clinical challenge. Additionally, whether the beneficial effects of exercise can be reproduced using pharmacological tools remains unknown and may benefit patients who are unable to exercise as a result of immobilization. We developed a murine model of orthopaedic trauma combining tibia fracture and pin fixation with muscle damage. Behavioural measures included mechanical nociceptive thresholds and distances run on exercise wheels. Bone healing was quantified using microcomputed tomagraphic scanning, and muscle fibre size distribution and fibrosis were followed using immunohistochemistry. We found that the model provided robust mechanical allodynia, fibrosis and a shift to smaller average muscle fibre size lasting up to 5weeks from injury. We also observed that allowing 'late' (weeks 1-2) rather than 'early' (weeks 0-1) exercise after injury resulted in greater overall running activity and greater reversal of allodynia. In parallel, the late running paradigm was associated with reduced muscle fibrosis, earlier increase in muscle fibre diameter and a short-term benefit in reducing callus volume. Providing the anti-fibrotic angiotensin receptor blocker losartan to mice in drinking water reduced both allodynia and muscle fibrosis. Combining losartan and late exercise provided no additional benefit. We conclude that early healing after orthopaedic trauma must be allowed prior to the initiation of exercise to achieve optimal pain, functional and physiological outcomes and that losartan is a viable candidate for translational studies.

Knockout of TLR4 promotes fracture healing by activating Wnt/β-catenin signaling pathway

ObjectivesThe aim of this study was to investigate the effect of Toll like receptor 4 (TLR4) on fracture healing. MethodsThe open tibial fracture models in TLR4 knockout (TLR4−/−) and wild type (WT) C57BL-6 J mice were established. The radiological examination, tartrate-resistant acid phosphatase (TRAP) staining, Micro-CT scan and biological torsion test were performed on 7, 14 and 21 days after operation. Enzyme Linked Immunosorbent Assay (ELISA) kit was used to detect the expression levels of tumor necrosis factor-α (TNF-α), interleukin-1 beta (IL-1β) and interleukin 6 (IL-6). Western blotting was used to detect the expression of β-catenin, Wingless-type MMTV integration site family, member 4 and 5B (Wnt4 and Wnt5B), proliferating cell nuclear antigen (PCNA) and bone morphogenetic protein-2 (BMP-2) of the callus tissue obtained from mice. ResultsTLR4 knockout promoted fracture healing, reduced the number of osteoclasts, increased bone callus volume (BV) and callus mineralized volume fraction (BV/TV%) (P < 0.05), increased the maximum torque and torsional stiffness of callus (P < 0.05), reduced TNF-α, IL-1β and IL-6 expression (P < 0.01), and increased the expression levels of β-catenin, Wnt4, Wnt5B, PCNA and BMP-2 (P < 0.01). ConclusionTLR4 knockout reduced inflammatory and promoted fracture healing by activating Wnt/β-catenin signaling pathway.

Nanofilm generated non-pharmacological anabolic bone stimulus.

Stimulus-responsive nanomaterials have mainly been employed to ablate or destroy tissues or to facilitate controlled release of drugs or biologics. Herein, we demonstrate the potential of stimulus-responsive nanomaterials to promote tissue regeneration via a non-pharmacological and noninvasive strategy. Thin nanofilms of an optically-absorbing organic dye or nanoparticle (single-walled graphene nanoribbons [SWOGNR]) were placed over (without touching the skin) a rodent femoral fracture site. A nanosecond pulsed near-infrared laser diode was employed to generate photoacoustic (PA) signals from the nanofilms. X-ray micro-computed tomography (microCT), histology, and mechanical testing results showed that daily PA stimulations of upto 45 min for 6 weeks (complete fracture healing) do not adversely affect bone regeneration and quality. Further, microCT and histological analysis showed 10 min daily stimulation for 2 weeks significantly increases bone quantity at the fracture sites of rats exposed to the nanoparticle-generated PA signals. In these rats, up to threefold increase in bone volume to callus volume ratio and twofold increase in bone mineral density within the callus were noted, compared to rats that were not exposed to the photoacoustic signals. The results taken together indicate that nanofilm-generated photoacoustic signals serve as an anabolic stimulus for bone regeneration. The results, in conjugation with the ability of these nanofilms to serve as PA contrast agents, present opportunities toward the development of integrated noninvasive imaging and noninvasive or invasive treatment strategies for bone loss due to disease or trauma.

Bone marrow concentrate efficacy in treatment of chronic aseptic nonunion

&lt;p class="abstract"&gt;&lt;strong&gt;Background:&lt;/strong&gt; Bone marrow is a rich source of osteoprogenitor cells that proliferate and differentiate into osteoblasts. Traditionally, autologous iliac bone grafts have been used in treatment of nonunion. This technique has its associated morbidity, which include donor site pain, infection, scarring, and nerve injury. The use of percutaneous bone marrow stem cells (BMSC) avoids these morbidities with comparable clinical results.&lt;/p&gt;&lt;p class="abstract"&gt;&lt;strong&gt;Methods:&lt;/strong&gt; We studied twenty patients of aseptic nonunion where bone marrow was aspirated from the anterior iliac crest, concentrated on a cell separator, and then injected into the nonunion site under c-arm guidance. Each nonunion received a relatively same amount of concentrated bone marrow. The volume of callus was calculated by dimensions obtained from RISPACS software in our institution.&lt;/p&gt;&lt;p class="abstract"&gt;&lt;strong&gt;Results:&lt;/strong&gt; There was an average of 29,418 mm3 of callus formation with a minimum of 4,455 mm&lt;sup&gt;3&lt;/sup&gt; and maximum of 68,460. The average time of union was 12 weeks with minimum of 6 weeks and maximum of 24 weeks.&lt;/p&gt;&lt;p class="abstract"&gt;&lt;strong&gt;Conclusions:&lt;/strong&gt; The percutaneous bone marrow concentrate injection provides an effective and a safe alternative method for the treatment of a chronic long standing non-union.&lt;/p&gt;

NLRP3 inflammasome inhibitor glyburide expedites diabetic-induced impaired fracture healing.

Localized inflammation is accompanied by the diabetic-induced fracture. The present study aims to investigate the therapeutic effects of glyburide, an NLRP3 inflammasome inhibitor, in a diabetic-induced fracture model. An animal model of diabetic-induced fracture was established and the mice were administrated with metformin or glyburide for 3 weeks. Quantitative polymerase chain reaction (qPCR) and Western blotting were used to evaluate the relative expressions of IFN-γ, TNF-α, and IL-6. Micro-computed tomography (μCT) scanning was applied to evaluate bone callus formation. Histopathology examinations of fractured femur sections were performed using Tartrate-resistant acid phosphatase (TRAP) staining and Alcian blue and orange G staining. Bone strength was evaluated using Torsional testing. Our results showed that treatment of glyburide significantly decreased the expressions of IFN-γ, TNF-α, and IL-6 in the fracture calluses in diabetic-induced fracture model, while bone callus volume and bone volume fraction were increased. Additionally, our results also demonstrated that treatment of glyburide rescued the increase of osteoclasts in the bone-cartilage interface. Apart from decreasing a percentage of cartilage area and increasing the percentage of bone and fibrotic tissue area, treatment of glyburide increased the maximum torque and yield torque of fractures. These results implied that glyburide might be used as a potential drug candidate for diabetic-induced fracture.

Development of a novel murine delayed secondary fracture healing in vivo model using periosteal cauterization

IntroductionDelayed union and nonunion development remain a major clinical problematic complication during fracture healing, with partially unclear pathophysiology. Incidences range from 5 to 40% in high-risk patients, such as patients with periosteal damage. The periosteum is essential in adequate fracture healing, especially during soft callus formation. In this study, we hypothesize that inducing periosteal damage in a murine bone healing model will result in a novel delayed union model.Materials and methodsA mid-shaft femoral non-critically sized osteotomy was created in skeletally mature C57BL/6 mice and stabilized with a bridging plate. In half of the mice, a thin band of periosteum adjacent to the osteotomy was cauterized. Over 42 days of healing, radiographic, biomechanical, micro-computed tomography and histological analysis was performed to assess the degree of fracture healing.ResultsAnalysis showed complete secondary fracture healing in the control group without periosteal injury. Whereas the periosteal injury group demonstrated less than half as much maximum callus volume (p < 0.05) and bridging, recovery of stiffness and temporal expression of callus growth and remodelling was delayed by 7–15 days.ConclusionThis paper introduces a novel mouse model of delayed union without a critically sized defect and with standardized biomechanical conditions, which enables further investigation into the molecular biological, biomechanical, and biochemical processes involved in (delayed) fracture healing and nonunion development. This model provides a continuum between normal fracture healing and the development of nonunions.

The influence of gap size on the development of fracture union with a micro external fixator

Increasingly, the rat femoral fracture model is being used for preclinical investigations of fracture healing, however, the effect of gap size and its influence on mechanobiology is not well understood. We aimed to evaluate the influence of osteotomy gap on osteotomy healing between the previously published extremes of guaranteed union (0.5 mm) and non-union (3 mm) using this model.A femoral osteotomy in 12–14 week old female Wistar rats was stabilised with a micro fixator (titanium blocks, carbon fiber bars) with an osteotomy gap of 1.0 mm (n = 5), 1.5 mm (n = 7), 2.0 mm (n = 6). After five weeks, the left femur was retrieved. The osteotomy gap was scanned using X-ray microtomography and then histologically evaluated. The radiographic union rate (complete mineralised bone bridging across the osteotomy) was three times higher for the 1.0 mm than the 2.0 mm gap. The 1.0 mm gap had the largest callus (0.069μm3) and bone volume (0.035μm3). Callus and bone volume were approximately 50% smaller within the 2.0 mm gap.Using cadaveric rat femurs stabilised with the external fixator, day 0 mechanical assessment of construct stiffness was calculated on materials testing machine displacement vs load output. The construct stiffness for the 1.0, 1.5 and 2.0 mm gaps was 32.6 ± 5.4, 32.5 ± 2.4, and 32.4 ± 8.3 N/mm (p = 0.779). Interfragmentary strain (IFS) was calculated using the change in osteotomy gap displacement as measured using microstrain miniature differential reluctance transducer spanning the osteotomy gap. Increasing the gap size significantly reduced the IFS (p = 0.013). The mean ‘day 0’ IFS for the 1.0, 1.5 and 2.0 mm gaps were 11.2 ± 1.3, 8.4 ± 1.5 and 6.1 ± 1.2% respectively.A 1.5 mm gap resulted in a delayed fracture healing by 5 weeks and may represent a useful test environment for fracture healing therapy. Increasing gap size did not affect construct stiffness, but did reduce the ‘day 0’ IFS, with a doubling of non-union and halving of bone volume measured between 1.0 and 2.0 mm gaps.

Effect of initial micro-movement of a fracture gap fastened by composite prosthesis on bone healing

Micro-movements are considered to have strong influence on the secondary healing process of fractured long bones, however, the translational shear movement is assumed to have a negative effect on the healing of such fractures. Numerical simulations were carried out with initially applied individual and combined, axial and shear displacements (ranging 0.1 mm to 1.0 mm for a 3 mm fracture at day 1) for a tibial fracture fastened by flexible composite prostheses. The best individual case of healing was seen with an initial axial displacement of 0.1 mm (86% reduced movement compared to the worst, 1.0 mm translational movement) while in combined cases, 0.1–0.1 mm (axial-shear combination) gave the best healing results (0.000157 mm) at day 56. Moreover, using composite intramedullary (IM) nail and bone plate as a superior implant resulted in 157.81% and 201% improved callus volumes with a promising bending stiffness of 20.62 and 19.03 Nm/θ, respectively, at day 154. But the excessive shear movement for the initial 4–6 weeks using an IM nail delayed fracture healing.

Grape seed extract supplement increases bone callus formation and mechanical strength: an animal study

BackgroundThe positive effects of grape seed proanthocyanidin extract (GSPE) on bone health, which is a potent antioxidant, are known but its effects on fracture healing are not sufficiently covered in the literature. This study aims to investigate the effects of GSPE on fracture healing and biomechanics of healing bone.Materials and methodsSixty-four adult Wistar-Albino male rats were divided into 8 groups of 8 animals in each group. Osteotomy was performed to the right femurs of all groups except the negative control (G1) and positive control (G2) groups, and intramedullary Kirchner wire was used for fixation. GSPE was given to half of the rats (G2-G4-G6-G8) 100 mg/kg/day by oral gavage. The rats were sacrificed on the tenth (G3–G4), twentieth (G5–G6), and thirtieth (G1–G2–G7–G8) days, respectively, and histopathological, radiological, and biomechanical examinations were performed.ResultsHistopathological examination of the specimens from the callus tissues revealed that bone healing was more prominent in the groups supplemented with GSPE (G4, G6, G8). There was a statistically significant improvement in radiological recovery scores and callus volumes in groups with GSPE. When biomechanical strengths were evaluated, it was found that GSPE increased bone strength not only in fracture groups but also in the positive control group (G2).ConclusionsAs a result, this study showed that GSPE, a potent anti-oxidant, had a positive effect on bone healing and improved mechanical strength of the healing bone.

Impact of Explants, Plant Growth Regulators and their Interaction on Micropropagation of Impatiens balsamina, L.

The aim of this study is to improve the propagation of Impatiens balsamina, L by an efficient protocol for in vitro propagation was successfully established through tissue culture technique. Cotyledonary node with sections of cotyledon, section of cotyledons and hypocotyls were derived from 21 days in vitro germinating seeds after surface sterilization. They were cultured for shoot multiplication and callus induction on MS medium fortified with various concentrations of BAP, TDZ or combination between BAP and NAA. It was determined that cotyledonary node with sections of cotyledon on MS medium supplemented with 4.5 mg/L BAP for maximum shoots number (4.33shoots) and callus induction 3.78. The best shoot length (3.08 cm) within 8 weeks from regeneration was achieved on MS medium fortified with 1.0 mg/L BAP + 0.5 mg/L NAA. The number of shoots formed per explants increased by re-culturing for six weeks on the same medium (4.5 mg/L BAP alone), since it produced (9 shoots). While the best shoot length (4cm) was obtained by sub culturing the explants on MS medium received 1.0 mg/L BAP + 0.5 mg/L NAA to MS without growth regulators after six weeks. The best callus fresh weight (9.32 g) and callus volume (7.76 cm3) were recorded with using TDZ at 3.0 mg/L that sub cultured on medium fortified with 4.5 mg/L BAP. In addition the highest dry weight (0.81 g) was obtained by re - culturing on the same medium that contained with 4.5 mg/L BAP.

Development of a novel murine femur fracture and fixation model

BackgroundAnimal models have been used for decades to simulate human fractures in the laboratory setting. Fracture models in mice are attractive because they offer a high volume, relatively low-cost method of investigating fracture healing characteristics. We report on the development of a novel murine femur fracture model that is rapid, reproducible and inexpensive. MethodsAs part of a pilot study to investigate the effects of smoking on fracture healing, fifteen 35–43 g twelve-week old female CD-1 mice underwent a novel surgical protocol using direct visualization of femur fracture creation and fixation. Following surgery, mice were sacrificed at 14 days, 28 days and 42 days. After sacrifice, the femora were analyzed using MicroCT and histology to evaluate progression of healing. ResultsOf the 14 mice that survived the surgical procedure (one succumbed to a complication of anesthesia), two lost reduction and did not heal. Histology demonstrated at 14 days 44.1% (SD±2.9%) of callus composed of cartilage. At 28 days there was 19.0% (SD±3.4%) of callus composed of cartilage. At 42 days there was 8.4% (SD±2.6%) callus composed of cartilage (p < 0.005). MicroCT demonstrated that from 14 to 42 days the average callus volume decreased from 101.6 mm3 to 68.2 mm3 while the relative bone volume of callus increased from 14 to 42 days (15%–31%) (p = 0.068). ConclusionsOur novel fracture and fixation model is an effective, rapid, reproducible and inexpensive method to simulate a fracture in a laboratory setting. Additionally, our model reliably creates a reproducible progression of radiographic and histological bone healing.

Do antiosteoporotic drugs improve bone regeneration in vivo?

Treatment of complex fractures in the elderly is a challenge for operative reconstruction due to degraded bone structure. Early peri-operative bone anabolic treatment could improve new bone formation, avoid implant loosening and accelerate fracture healing. To compare the osteoanabolic potential of different drugs after distraction osteogenesis, 168 female Sprague-Dawley rats underwent lengthening of the right femur using a monolateral external fixator. Animals were randomly divided into six groups: vehicle-injected group, PTH(1-34), raloxifen, strontium ranelate, alendronate and simvastatin. Histomorphometry, CT-scanning, DEXA- and biomechanical analysis were performed to evaluate new bone formation, callus volume, mineralisation and biomechanical strength. Expression of bone metabolic mediators and differentiation indicators of distracted and intact bone were examined by RT-PCR and western blot. Histological analysis showed significant increase of the bone mass after treatment with PTH(1-34), raloxifen and strontium ranelate (p = 0.02). Raloxifen increased bone mineral content (BMC) of the whole distracted femur significantly (p = 0.007). Callus volume was significantly larger in the PTH(1-34), raloxifen and simvastatin groups (p = 0.001) compared to control. Ultimate load of distracted new formed bone was increased in PTH(1-34) and raloxifen groups. It seems that PTH(1-34) and raloxifen have a stronger effect on bone where a repair response is activated. Strontium ranelate demonstrates similar effects to PTH regarding new bone formation but shows low values for mineralisation and biomechanical strength. This study suggests that peri-operative treatment of complex and/or osteoporotic fractures with PTH(1-34) and raloxifen might be useful as a stimulator of bone formation and mineralisation to shorten the consolidation time in humans.