Computational model of the dual action of PTH - Application to a rat model of osteoporosis.

This paper presents a pharmacokinetic/pharmacodynamic (PK/PD) model of the action of PTH(1-34) on bone modelling and remodelling, developed for quantitatively investigating the dose- and administration pattern-dependency of the bone tissue response to this drug. Firstly, a PK model of PTH(1-34) was developed, accounting for administration via subcutaneous injections. Subsequently, the PK model was coupled to a (mechanistic) bone cell population model of bone modelling and remodelling, taking into account the effects of PTH(1-34) on the differentiation of lining cells into active osteoblasts, on the apoptosis of active osteoblasts, and on proliferation of osteoblast precursors, as well as on the key regulatory pathways of bone cell activities. Numerical simulations show that the coupled PK/PD model is able to distinguish between continuous and intermittent administration patterns of PTH(1-34), in terms of yielding both catabolic bone responses (if drug administration is carried out continuously) and anabolic bone responses (if drug administration is carried out intermittently). The model also features a non-linear relation between bone gain and drug dose (as known from experiments); doubling the dose from 80 µg/kg/day to 160 µg/kg/day induced a 1.3-fold increase of the bone volume-to-total volume ratio. Furthermore, the model presented in this paper confirmed that bone modelling represents an essential mechanism of the anabolic response of bone to PTH(1-34) administration in rat models, and that the large amount of bone formation observed in such models cannot be explained via remodelling alone.

Monocyte Chemoattractant Protein-1 (MCP-1/CCL2) Drives Activation of Bone Remodelling and Skeletal Metastasis.

PURPOSE OF REVIEW: The purpose of this review is to explore the role of monocyte chemoattractant protein-1 (MCP-1 or CCL2) in the processes that underpin bone remodelling, particularly the action of osteoblasts and osteoclasts, and its role in the development and metastasis of cancers that target the bone. RECENT FINDINGS: MCP-1 is a key mediator of osteoclastogenesis, being the highest induced gene during intermittent treatment with parathyroid hormone (iPTH), but also regulates catabolic effects of continuous PTH on bone including monocyte and macrophage recruitment, osteoclast formation and bone resorption. In concert with PTH-related protein (PTHrP), MCP-1 mediates the interaction between tumour-derived factors and host-derived chemokines to promote skeletal metastasis. In breast and prostate cancers, an osteolytic cascade is driven by tumour cell-derived PTHrP that upregulates MCP-1 in osteoblastic cells. This relationship between PTHrP and osteoblastic expression of MCP-1 may drive the colonisation of disseminated breast cancer cells in the bone. There is mounting evidence to suggest a pivotal role of MCP-1 in many diseases and an important role in the establishment of comorbidities. Coupled with its role in bone remodelling and the regulation of bone turnover, there is the potential for pathological relationships between bone disorders and bone-related cancers driven by MCP-1. MCP-1's role in bone remodelling and bone-related cancers highlights its potential as a novel anti-resorptive and anti-metastatic target.

CCL2 and CCR2 are Essential for the Formation of Osteoclasts and Foreign Body Giant Cells.

Osteoclasts are multinucleated cells responsible for bone resorption. They are derived from the fusion of cells in the monocyte/macrophage lineage. Monocytes and macrophages can also fuse to form foreign body giant cells (FBGC). Foreign body giant cells are observed at the interface between a host and a foreign body such as implants during a foreign body reaction. Macrophages are attracted to the site of bone resorption and foreign body reactions by different cytokines. Chemokine (C-C) ligand-2 (CCL2) is an important chemotactic factor and binds to a receptor CCR2. In this study we investigated the importance of CCL2 and the receptor CCR2 in the formation of osteoclasts and FBGC. CCL2 mRNA was more highly expressed in giant cell culture than macrophages, being 9-fold and 16-fold more abundant in osteoclasts and FBGC respectively. Significantly fewer osteoclasts and FBGC were cultured from the bone marrow of CCL2 and CCR2 knockout mice, when compared to wild type. Not only were the number of giant cells reduced but there was a significant reduction in the number of nuclei and the size of these cells in the cultures of CCL2 and CCR2 knockout mice. Formation of osteoclasts and FBGC were recovered in cultures by addition of exogenous CCL2 to the media containing marrow cells from CCL2-/- mice. We conclude that CCL2 and its receptor CCR2 are important for the formation of osteoclasts and FBGC and absence of these genes causes inhibition of osteoclast and FBGC formation.

Pentosan Polysulfate: a Novel Glycosaminoglycan-Like Molecule for Effective Treatment of Alphavirus-Induced Cartilage Destruction and Inflammatory Disease.

UNLABELLED: Arthritogenic alphaviruses such as Ross River virus (RRV) and chikungunya virus (CHIKV) cause large-scale epidemics of severe musculoskeletal disease and have been progressively expanding their global distribution. Since its introduction in July 2014, CHIKV now circulates in the United States. The hallmark of alphavirus disease is crippling pain and inflammation of the joints, a similar immunopathology to rheumatoid arthritis. The use of glycans as novel therapeutics is an area of research that has increased in recent years. Here, we describe the promising therapeutic potential of the glycosaminoglycan (GAG)-like molecule pentosan polysulfate (PPS) to alleviate virus-induced arthritis. Mouse models of RRV and CHIKV disease were used to characterize the extent of cartilage damage in infection and investigate the potential of PPS to treat disease. This was assessed using histological analysis, real-time PCR, and fluorescence-activated cell sorting (FACS). Alphaviral infection resulted in cartilage destruction, the severity of which was alleviated by PPS therapy during RRV and CHIKV clinical disease. The reduction in cartilage damage corresponded with a significant reduction in immune infiltrates. Using multiplex bead arrays, PPS treatment was found to have significantly increased the anti-inflammatory cytokine interleukin-10 and reduced proinflammatory cytokines, typically correlated with disease severity. Furthermore, we reveal that the severe RRV-induced joint pathology, including thinning of articular cartilage and loss of proteoglycans in the cartilage matrix, was diminished with treatment. PPS is a promising new therapy for alphavirus-induced arthritis, acting to preserve the cartilage matrix, which is damaged during alphavirus infection. Overall, the data demonstrate the potential of glycotherapeutics as a new class of treatment for infectious arthritis. IMPORTANCE: The hallmark of alphavirus disease is crippling pain and joint arthritis, which often has an extended duration. In the past year, CHIKV has expanded into the Americas, with approximately 1 million cases reported to date, whereas RRV continues to circulate in the South Pacific. Currently, there is no licensed specific treatment for alphavirus disease, and the increasing spread of infection highlights an urgent need for therapeutic intervention strategies. Pentosan polysulfate (PPS) is a glycan derivative that is orally bioavailable, has few toxic side effects, and is currently licensed under the name Elmiron for the treatment of cystitis in the United States. Our findings show that RRV infection damages the articular cartilage, including a loss of proteoglycans within the joint. Furthermore, treatment with PPS reduced the severity of both RRV- and CHIKV-induced musculoskeletal disease, including a reduction in inflammation and joint swelling, suggesting that PPS is a promising candidate for drug repurposing for the treatment of alphavirus-induced arthritis.

Bindarit, an inhibitor of monocyte chemotactic protein synthesis, protects against bone loss induced by chikungunya virus infection.

UNLABELLED: The recent global resurgence of arthritogenic alphaviruses, in particular chikungunya virus (CHIKV), highlights an urgent need for the development of therapeutic intervention strategies. While there has been significant progress in defining the pathophysiology of alphaviral disease, relatively little is known about the mechanisms involved in CHIKV-induced arthritis or potential therapeutic options to treat the severe arthritic symptoms associated with infection. Here, we used microcomputed tomographic (µCT) and histomorphometric analyses to provide previously undescribed evidence of reduced bone volume in the proximal tibial epiphysis of CHIKV-infected mice compared to the results for mock controls. This was associated with a significant increase in the receptor activator of nuclear factor-κB ligand/osteoprotegerin (RANKL/OPG) ratio in infected murine joints and in the serum of CHIKV patients. The expression levels of the monocyte chemoattractant proteins (MCPs), including MCP-1/CCL2, MCP-2/CCL8, and MCP-3/CCL7, were also highly elevated in joints of CHIKV-infected mice, accompanied by increased cellularity within the bone marrow in tibial epiphysis and ankle joints. Both this effect and CHIKV-induced bone loss were significantly reduced by treatment with the MCP inhibitor bindarit. Collectively, these findings demonstrate a unique role for MCPs in promoting CHIKV-induced osteoclastogenesis and bone loss during disease and suggest that inhibition of MCPs with bindarit may be an effective therapy for patients affected with alphavirus-induced bone loss. IMPORTANCE: Arthritogenic alphaviruses, including chikungunya virus (CHIKV) and Ross River virus (RRV), cause worldwide outbreaks of polyarthritis, which can persist in patients for months following infection. Previous studies have shown that host proinflammatory soluble factors are associated with CHIKV disease severity. Furthermore, it is established that chemokine (C-C motif) ligand 2 (CCL2/MCP-1) is important in cellular recruitment and inducing bone-resorbing osteoclast (OC) formation. Here, we show that CHIKV replicates in bone and triggers bone loss by increasing the RANKL/OPG ratio. CHIKV infection results in MCP-induced cellular infiltration in the inflamed joints, and bone loss can be ameliorated by treatment with an MCP-inhibiting drug, bindarit. Taken together, our data reveal a previously undescribed role for MCPs in CHIKV-induced bone loss: one of recruiting monocytes/OC precursors to joint sites and thereby favoring a pro-osteoclastic microenvironment. This suggests that bindarit may be an effective treatment for alphavirus-induced bone loss and arthritis in humans.

Integrating gross pathology into teaching of undergraduate medical science students using human cadavers.

Human cadavers offer a great opportunity for histopathology students for the learning and teaching of tissue pathology. In this study, we aimed to implement an integrated learning approach by using cadavers to enhance students' knowledge and to develop their skills in gross tissue identification, handling and dissection techniques. A total of 35 students enrolled in the undergraduate medical science program participated in this study. A 3-hour laboratory session was conducted that included an active exploration of cadaveric specimens to identify normal and pathological tissues as well as tissue dissection. The majority of the students strongly agreed that the integration of normal and morbid anatomy improved their understanding of tissue pathology. All the students either agreed or strongly agreed that this laboratory session was useful to improve their tissue dissection and instrument handling skills. Furthermore, students from both cohorts rated the session as very relevant to their learning and recommended that this approach be added to the existing histopathology curriculum. To conclude, an integrated cadaver-based practical session can be used effectively to enhance the learning experience of histopathology science students, as well as improving their manual skills of tissue treatment, instrument handling and dissection.

EphrinB2 signaling in osteoblasts promotes bone mineralization by preventing apoptosis.

Cells that form bone (osteoblasts) express both ephrinB2 and EphB4, and previous work has shown that pharmacological inhibition of the ephrinB2/EphB4 interaction impairs osteoblast differentiation in vitro and in vivo. The purpose of this study was to determine the role of ephrinB2 signaling in the osteoblast lineage in the process of bone formation. Cultured osteoblasts from mice with osteoblast-specific ablation of ephrinB2 showed delayed expression of osteoblast differentiation markers, a finding that was reproduced by ephrinB2, but not EphB4, RNA interference. Microcomputed tomography, histomorphometry, and mechanical testing of the mice lacking ephrinB2 in osteoblasts revealed a 2-fold delay in bone mineralization, a significant reduction in bone stiffness, and a 50% reduction in osteoblast differentiation induced by anabolic parathyroid hormone (PTH) treatment, compared to littermate sex- and age-matched controls. These defects were associated with significantly lower mRNA levels of late osteoblast differentiation markers and greater levels of osteoblast and osteocyte apoptosis, indicated by TUNEL staining and transmission electron microscopy of bone samples, and a 2-fold increase in annexin V staining and 7-fold increase in caspase 8 activation in cultured ephrinB2 deficient osteoblasts. We conclude that osteoblast differentiation and bone strength are maintained by antiapoptotic actions of ephrinB2 signaling within the osteoblast lineage.

Differential expression of chemokines, chemokine receptors and proteinases by foreign body giant cells (FBGCs) and osteoclasts.

Osteoclasts and foreign body giant cells (FBGCs) are both derived from the fusion of macropahges. These cells are seen in close proximity during foreign body reactions, therefore it was assumed that they might interact with each other. The aim was to identify important genes that are expressed by osteoclasts and FBGCs which can be used to understand peri-implantitis and predict the relationship of these cells during foreign body reactions. Bone marrow macrophages (BMM) were treated with receptor activator of nuclear factor kappa B ligand (RANKL) to produce osteoclasts. Quantitative PCR (qPCR) was used to identify the genes that were expressed by osteoclasts and FBGCs compared to macrophage controls. TRAP staining was used to visualise the cells while gelatine zymography and western blots were used for protein expression. Tartrate-resistant acid phosphatase (TRAP), matrix metallo proteinase 9 (MMP9), nuclear factor of activated T cells 1 (NFATc1), cathepsin K (CTSK) and RANK were significantly lower in FBGCs compared to osteoclasts. Inflammation specific chemokines such as monocyte chemotactic protein (MCP1 also called CCL2), macrophage inflammatory protein 1 alpha (MIP1α), MIP1ß and MIP1γ, and their receptors CCR1, CCR3 and CCR5, were highly expressed by FBGCs. FBGCs were negative for osteoclast specific markers (RANK, NFATc1, CTSK). FBGCs expressed chemokines such as CCL2, 3, 5 and 9 while osteoclasts expressed the receptors for these chemokines i.e. CCR1, 2 and 3. Our findings show that osteoclast specific genes are not expressed by FBGCs and that FBGCs interact with osteoclasts during foreign body reaction through chemokines.

Validating real-time three-dimensional echocardiography against cardiac magnetic resonance, for the determination of ventricular mass, volume and ejection fraction: a meta-analysis.

INTRODUCTION: Real-time three-dimensional echocardiography (RT3DE) is currently being developed to overcome the challenges of two-dimensional echocardiography, as it is a much cheaper alternative to the gold standard imaging method, cardiac magnetic resonance (CMR). The aim of this meta-analysis is to validate RT3DE by comparing it to CMR, to ascertain whether it is a practical imaging method for routine clinical use. METHODS: A systematic review and meta-analysis method was used to synthesise the evidence and studies published between 2000 and 2021 were searched using a PRISMA approach. Study outcomes included left ventricular end-systolic volume (LVESV), left ventricular end-diastolic volume (LVEDV), left ventricular ejection fraction (LVEF), left ventricular mass (LVM), right ventricular end-systolic volume (RVESV), right ventricular end-diastolic volume (RVEDV) and right ventricular ejection fraction (RVEF). Subgroup analysis included study quality (high, moderate), disease outcomes (disease, healthy and disease), age group (50 years old and under, over 50 years), imaging plane (biplane, multiplane) and publication year (2010 and earlier, after 2010) to determine whether they explained the heterogeneity and significant difference results generated on RT3DE compared to CMR. RESULTS: The pooled mean differences for were - 5.064 (95% CI - 10.132, 0.004, p > 0.05), 4.654 (95% CI - 4.947, 14.255, p > 0.05), - 0.783 (95% CI - 5.630, 4.065, p > 0.05, - 0.200 (95% CI - 1.215, 0.815, p > 0.05) for LVEF, LVM, RVESV and RVEF, respectively. We found no significant difference between RT3DE and CMR for these variables. Although, there was a significant difference between RT3DE and CMR for LVESV, LVEDV and RVEDV where RT3DE reports a lower value. Subgroup analysis indicated a significant difference between RT3DE and CMR for studies with participants with an average age of over 50 years but no significant difference for those under 50. In addition, a significant difference between RT3DE and CMR was found in studies using only participants with cardiovascular diseases but not in those using a combination of diseased and healthy participants. Furthermore, for the variables LVESV and LVEDV, the multiplane method shows no significant difference between RT3DE and CMR, as opposed to the biplane showing a significant difference. This potentially indicates that increased age, the presence of cardiovascular disease and the biplane analysis method decrease its concordance with CMR. CONCLUSION: This meta-analysis indicates promising results for the use of RT3DE, with limited difference to CMR. Although in some cases, RT3DE appears to underestimate volume, ejection fraction and mass when compared to CMR. Further research is required in terms of imaging method and technology to validate RT3DE for routine clinical use.

Utility of Cadaveric Porcine Heads for Teaching Oral Surgical Procedures in an Australian Dental School: A Pilot Study.

Background/Objectives: Cadaveric models have traditionally been a mainstay of dental and medical education worldwide since their inception. In Australia, educators at dental schools were among the first to use cadaveric porcine heads in formal teaching in oral surgery. This practice has since fallen out of favour in most modern dental curricula. The aim of this pilot study was to determine the utility of cadaveric porcine models for oral surgery training from a student perspective (Griffith University, Gold Coast, Australia). Methods: Thirty participants who were all third-year dental students attended a two-hour session comprising a 30 min lecture followed by a 90 min practical workshop. The lecture outlined the steps and supervision of students during the practical and was provided by a consultant maxillofacial surgeon. At the conclusion of the workshop, participants were asked to anonymously complete a printed questionnaire with eight questions related to their experience. Results: Prior to the workshop, two-thirds (61%) of participants felt that they had been taught the surgical procedure for raising mucoperiosteal flaps adequately in their dental school curriculum during their third year, although only 43% of students had assisted specialty residents in raising a mucoperiosteal flap and 14% reported having performed the procedure themselves. Almost all students (96%) agreed that the porcine model was useful for their dental education and that they would practice the exercise using the model again if provided with the opportunity. The questionnaire had a 93.33% completion rate. Conclusions: This pilot study indicates that porcine heads present a useful, low-cost adjunct in the learning of basic oral surgical procedures.

Foreign body giant cells and osteoclasts are TRAP positive, have podosome-belts and both require OC-STAMP for cell fusion.

Macrophages have the ability to fuse and form multinucleated giant cells such as Osteoclast (OCs) and FBGCs. Osteoclast stimulatory transmembrane protein (OC-STAMP) is an important cell surface protein involved in the formation of OCs. This study sought to determine if OC-STAMP also regulates formation of FBGCs using expression analysis and subsequent inhibition studies. qPCR and Western blot analysis showed that OC-STAMP expression is significantly higher in FBGCs compared to control monocytes (P < 0.05). Four days following cell culture, OCs were positive for TRAP and F-actin ring formation, but FBGCs were not. In contrast, FBGCs were positive for TRAP and showed podosome belts comprised of F-actin on Day 8. FBGCs were subsequently plated onto dentine, but despite presenting some morphologic features of OCs (OC-STAMP expression, TRAP reactivity, and podosome belts) they failed to resorb bone. To evaluate a role for OC-STAMP in FBGCs, we inhibited this cell surface protein with anti-OC-STAMP antibody and observed that cell fusion and podosome belt formation was inhibited in both OCs and FBGCs. Our data support the hypothesis that OC-STAMP is a regulatory molecule for FBGCs; and that they are functionally distinct from OCs, despite similarities in gene expression profile, podosome belt formation, and TRAP expression.

DNA nanotherapy for pre-neoplastic cervical lesions.

OBJECTIVE: This study aims to test the hypothesis that targeted nanoparticle delivery of DNA encoding HPV16-regulated diphtheria toxin (DT-A) will result in the death of HPV16-infected cells. MATERIALS AND METHODS: Plasmid constructs containing a HPV16 Long Control Region (LCR) DNA sequence upstream of DT-A or luciferase reporter (Luc) DNA sequences were used to formulate poly(ß-amino ester) nanoparticles. The effect on tumor growth of HPV/DT-A-nanoparticle injection directly into HPV16(+) CaSki human cervical cancer cell-derived xenografts in mice was determined. To evaluate the ability of the HPV16 LCR regulatory sequence to activate gene expression specifically in HPV16-infected cells, mice underwent bioluminescent optical imaging following intraperitoneal injection of HPV/Luc-nanoparticles. The use of Lutrol F127, a thermal-sensitive gel, to target delivery of nanoparticles and subsequent gene expression to cervical epithelial cells was evaluated in ex vivo cultures of mouse cervix and following intravaginal delivery of nanoparticle/gel in mice, as well as in ex vivo cultures of surgical LEEP samples. RESULTS: The selected HPV16 LCR regulatory sequence activates gene expression in both HPV16-infected cells and non-infected cells. However, in the cervix, it is specifically active in epithelial cells. Following exposure of cervical cells to HPV/DT-A-nanoparticles mixed with Lutrol F127 gel, DT-A is expressed and cells die. CONCLUSIONS: An HPV16 DNA sequence that targets gene expression specifically to HPV16-infected cells remains to be discovered. Topical application of a Lutrol F127 thermal gel/nanoparticle mix is illustrative of how to restrict exposure of cells to therapeutic nanoparticles, thereby allowing for targeted DNA delivery to cervical pre-cancerous lesions.

Association of change in fat and lean mass with incident cardiovascular events for women in midlife and beyond: A prospective study using dual-energy x-ray absorptiometry (DXA).

OBJECTIVE: To determine whether changes in fat and lean mass over time, quantified using dual-energy x-ray absorptiometry (DXA), are related to incident cardiovascular events. Previous studies using surrogate anthropometric methods have had inconsistent findings. STUDY DESIGN: Prospective, longitudinal observational study of women aged 40 to 80 randomly selected from the electoral roll and stratified into decades: 40-49, 50-59, 60-69 and 70-79 years. MAIN OUTCOME MEASURES: Changes in anthropometric measurements (body mass index and waist-to-hip ratio) and DXA-quantified fat mass and lean mass between the first and fifth years of the study. Incident cardiovascular events recorded from the sixth to the 12th year. RESULTS: In total 449 participants (87.9 %) were analyzed. A 10 % or greater decrease in total fat mass index was associated with a 67 % lower likelihood of any cardiovascular event (OR = 0.33, 95%CI 0.15-0.71); no association was observed for an increase. A 10 % or greater decrease in abdominal fat mass index was associated with a 62 % lower likelihood of incident stroke (OR = 0.38, 95%CI 0.16-0.91); no association was observed for an increase. A 10 % or greater decrease in appendicular lean mass index resulted in increased odds ratio of 2.91 for incident peripheral artery events (OR = 2.91, 95%CI 1.18-7.20). CONCLUSIONS: Reducing fat mass for women in midlife and beyond may decrease the risk of cardiovascular events. An increase in fat mass may not contribute to additional cardiovascular events. A reduction in limb muscle mass may provide an independent marker for cardiometabolic risk and peripheral artery disease. No independent association was found using anthropometric measurements and incident cardiovascular events.

MCP-1 expression in breast cancer and its association with distant relapse.

BACKGROUND: Distant relapse of breast cancer complicates management of the disease and accounts for 90% of breast cancer-related deaths. Monocyte chemoattractant protein-1 (MCP-1) has critical roles in breast cancer progression and is widely accepted as a pro-metastatic chemokine. METHODS: This study explored MCP-1 expression in the primary tumour of 251 breast cancer patients. A simplified 'histoscore' was used to determine if each tumour had high or low expression of MCP-1. Patient breast cancers were retrospectively staged based on available patient data. p < 0.05 was used to determine significance and changes in hazard ratios between models were considered. RESULTS: Low MCP-1 expression in the primary tumour was associated with breast cancer-related death with distant relapse in ER- breast cancers (p < 0.01); however, this was likely a result of most low MCP-1-expressing ER- breast cancers being Stage III or Stage IV, with high MCP-1 expression in the primary tumour significantly correlated with Stage I breast cancers (p < 0.05). Expression of MCP-1 in the primary ER- tumours varied across Stage I, II, III and IV and we highlighted a switch in MCP-1 expression from high in Stage I ER- cancers to low in Stage IV ER- cancers. CONCLUSION: This study has emphasised a critical need for further investigation into MCP-1's role in breast cancer progression and improved characterisation of MCP-1 in breast cancers, particularly in light of the development of anti-MCP-1, anti-metastatic therapies.

Effects of PTH glandular and external dosing patterns on bone cell activity using a two-state receptor model-Implications for bone disease progression and treatment.

Temporal aspects of ligand specificity have been shown to play a significant role in the case of pulsatile hormone secretion, as exemplified by parathyroid hormone (PTH) binding to its receptor (PTH1R), a G-protein-coupled receptor expressed on surfaces of osteoblasts and osteocytes. The latter binding reaction regulates intracellular signalling and subsequently modulates skeletal homeostasis via bone remodelling. PTH glandular secretion patterns dictate bone cellular activity. In healthy humans, 70% of PTH is secreted in a tonic fashion, whereas 30% is secreted in low-amplitude and high-frequency bursts occurring every 10-20 min, superimposed on the tonic secretion. Changes in the PTH secretion patterns have been associated with various bone diseases. In this paper, we analyse PTH glandular secretion patterns for healthy and pathological states and their link to bone cellular responsiveness (αR). We utilise a two-state receptor ligand binding model of PTH to PTH1R together with a cellular activity function which is able to distinguish various aspects of the stimulation signal including peak dose, time of ligand exposure, and exposure period. Formulating and solving several constrained optimisation problems, we investigate the potential of pharmacological manipulation of the diseased glandular secretion and via clinical approved external PTH injections to restore healthy bone cellular responsiveness. Based on the mean experimentally reported data, our simulation results indicate cellular responsiveness in healthy subjects is sensitive to the tonic baseline stimulus and it is 28% of the computed maximum responsiveness. Simulation results for pathological cases of glucocorticoid-induced osteoporosis, hyperparathyroidism, initial and steady state hypocalcemia clamp tests indicate αR values significantly larger than the healthy baseline (1.7, 2.2, 4.9 and 1.9-times, respectively). Manipulation of the pulsatile glandular secretion pattern, while keeping the mean PTH concentration constant, allowed restoration of healthy baseline values from these catabolic bone diseases. Conversely, PTH glandular diseases that led to maximum bone cellular responsiveness below the healthy baseline value can't be restored to baseline via glandular manipulation. However, external PTH injections allowed restoration of these latter cases.

Monocyte Chemotactic Protein-1 (MCP1) Accumulation in Human Osteoclast Precursor Cultures.

In vitro osteoclast methods require constant treatment with macrophage colony stimulating factor (M-CSF) to support precursor survival and addition of the differentiation agent receptor activator of NF-κB ligand (RANKL). Constant exposure to granulocyte macrophage colony stimulating factor (GM-CSF) suppresses human osteoclast formation in vitro. Addition of the chemokine monocyte chemotactic protein-1 (MCP1) to such cultures dramatically increases osteoclast formation and overcomes GM-CSF mediated suppression. We investigated the effect of M-CSF, GM-CSF and the combination of M-CSF and GM-CSF treatment on the expression of chemokines in human CD14+ cells in culture. Of assayed chemokines, MCP1 was the most abundant in terms of mRNA transcript and protein in M-CSF treated cultures and was suppressed by GM-CSF. MCP1 protein accumulated up to 50 ng/mL in culture medium, greatly exceeding other assayed chemokines. C-C chemokine receptor-2 (CCR2) is the receptor for MCP1: the formation of osteoclast-like cells was inhibited by constant exposure to the CCR2 antagonist RS102895, in part by decreasing expression of RANK, the receptor for RANKL.

The effect of low-intensity whole-body vibration with or without high-intensity resistance and impact training on risk factors for proximal femur fragility fracture in postmenopausal women with low bone mass: study protocol for the VIBMOR randomized controlled trial.

BACKGROUND: The prevailing medical opinion is that medication is the primary (some might argue, only) effective intervention for osteoporosis. It is nevertheless recognized that osteoporosis medications are not universally effective, tolerated, or acceptable to patients. Mechanical loading, such as vibration and exercise, can also be osteogenic but the degree, relative efficacy, and combined effect is unknown. The purpose of the VIBMOR trial is to determine the efficacy of low-intensity whole-body vibration (LIV), bone-targeted, high-intensity resistance and impact training (HiRIT), or the combination of LIV and HiRIT on risk factors for hip fracture in postmenopausal women with osteopenia and osteoporosis. METHODS: Postmenopausal women with low areal bone mineral density (aBMD) at the proximal femur and/or lumbar spine, with or without a history of fragility fracture, and either on or off osteoporosis medications will be recruited. Eligible participants will be randomly allocated to one of four trial arms for 9 months: LIV, HiRIT, LIV + HiRIT, or control (low-intensity, home-based exercise). Allocation will be block-randomized, stratified by use of osteoporosis medications. Testing will be performed at three time points: baseline (T0), post-intervention (T1; 9 months), and 1 year thereafter (T2; 21 months) to examine detraining effects. The primary outcome measure will be total hip aBMD determined by dual-energy X-ray absorptiometry (DXA). Secondary outcomes will include aBMD at other regions, anthropometrics, and other indices of bone strength, body composition, physical function, kyphosis, muscle strength and power, balance, falls, and intervention compliance. Exploratory outcomes include bone turnover markers, pelvic floor health, quality of life, physical activity enjoyment, adverse events, and fracture. An economic evaluation will also be conducted. DISCUSSION: No previous studies have compared the effect of LIV alone or in combination with bone-targeted HiRIT (with or without osteoporosis medications) on risk factors for hip fracture in postmenopausal women with low bone mass. Should either, both, or combined mechanical interventions be safe and efficacious, alternative therapeutic avenues will be available to individuals at elevated risk of fragility fracture who are unresponsive to or unwilling or unable to take osteoporosis medications. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry (www. anzctr.org.au ) (Trial number ANZCTR12615000848505, https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id = 368962 ); date of registration 14/08/2015 (prospectively registered). Universal Trial Number: U1111-1172-3652.

Sponge swabs increase sensitivity of sterility testing of processed bone and tendon allografts.

Sterility testing is the final, and critical, step in quality control of tissue banking. It informs the decision whether to release the tissue allografts for clinical use, or not. The most common method for sterility testing of structural bone and tendon allografts is to swab using cotton tip streaks. This method provides low recovery efficiency; and therefore may pass allografts with low bioburden, providing false negatives. Our pilot data revealed organism recovery efficiencies of 60, 30 and 100% from cotton swab, membrane filtration and sponge swaps, respectively. Our aim was to develop a high sensitivity sterility test for structural bone and tendon allografts using a sponge sampling method. Eighty-one bone and tendon allograft samples were inoculated with organism suspensions (10(2) or less organisms per 0.1 mL) of Clostridium sporogenes, Staphylococcus aureus, Pseudomonas aeruginosa, Candida albicans, Bacillus subtilis, Aspergillus niger, Staphylococcus epidermidis and Micrococcus spp. Nasco sponges (4 × 8 cm) were used to aseptically sample the whole surface of allograft samples. The sponges were cut in half and cultured in either tryptone soya or fluid thioglycollate broths for 14 days. Positive culture samples were further examined for microbial morphology. The results showed that the sensitivity of the method, and negative predictive value, is 100% for all inoculated organisms incubated with thioglycollate. We conclude that this sponge sampling method should be applied as the standard for sterility testing of structural bone and tendon allografts.

Validation of 11 kGy as a radiation sterilization dose for frozen bone allografts.

A radiation sterilization dose (RSD) of 25 kGy is deleterious to bone allografts. This study aimed to establish a lower RSD for bone allografts using method 1 of International Standard Organisation 11137.2:2006. This provides a database to select an RSD corresponding to an allograft's bioburden, given that the bioburden's gamma resistance is equal to or less than the standard. This can be verified by irradiating 100 allografts at a dose selected to provide a sterility assurance level of 10(-2). The bioburden of our allografts was 0, which prescribed a verification dose of 1.3 kGy. After irradiating 100 allografts, sterility tests returned no positive cultures. We therefore validated an RSD of 11 kGy for allografts with that bioburden. According to the standard, this RSD provides a sterility assurance level of 10(-6) for bone allografts.