Calcified Zone Research Articles

Treatment with recombinant lubricin attenuates osteoarthritis by positive feedback loop between articular cartilage and subchondral bone in ovariectomized rats

Osteoarthritis (OA) is a most commonly multifactorial degenerative joint disease along with the aging population, particularly in postmenopausal women. During the onset of OA, articular cartilage and subchondral bone act in concert as a functional unit. This present study is to investigate the effects of early or late treatment with recombinant lubricin on the onset of osteoarthritis (OA) in ovariectomized (OVX) rats. We found that both early and late recombinant lubricin treatments attenuated the onset of OA by positive feedback loop between articular cartilage and subchondral bone, although late treatment contributed to a lesser effect compared with early treatment. Specifically, treatment with recombinant lubricin protected articular cartilage from degeneration, demonstrated by lower proteoglycan loss, lower OARSI scores, less calcification cartilage zone and reduced immunostaining for collagen X (Col X) and matrix metalloproteinase (MMP-13) but increased the expression of lubricin, in comparison with vehicle-treated OVX rat group. Further, chondroprotective effects of lubricin normalized bone remodeling in subchondral bone underneath. It's suggested that treatment with recombinant lubricin inhibited the elevation of TRAP and Osterix positive cells in OVX rats and led to the normalization of subchondral bone microarchitectures with the suppression of subsidence of bone volume ratio (BV/TV) and trabecular thickness (Tb.Th) and the increase of trabecular separation (Tb.Sp) in vehicle-treated OVX rats. What's more, the normalization of subchondral bone in turn attenuated the articular cartilage erosion by inhibiting vascular invasion from subchondral bone to calcified cartilage zone, exemplified by inhibiting the elevation of CD31 positive cells in calcified cartilage and angiography in subchondral bone. Together, these results shed light that both early and late recombinant lubricin treatments attenuate the onset of OA by balancing the interplay between articular cartilage and subchondral bone in OVX rats, while also providing a further rationale for its therapeutic targeting to postmenopausal OA and suggesting that treatment timing is a pivotal factor for better effect acquisition.

Mönckeberg's arteriosclerosis: vascular calcification complicating microvascular surgery

Mönckeberg's arteriosclerosis is often an incidental finding, identified either clinically or on plain radiography. It can occasionally be associated with diabetes mellitus or chronic kidney disease. It differs from the more common atherosclerosis in that the tunica intima remains largely unaffected and the diameter of the vessel lumen is preserved. Despite such vessels appearing hard and pulseless throughout their affected length, they deliver relatively normal distal perfusion, indeed there is often a bounding pulse at the end of the calcified zone. They appear unremarkable on magnetic resonance angiography but visibly calcified on plain radiography. Mönckeberg's arteriosclerosis has a prevalence of<1% of the population, but when it does occur it can cause consternation at the prospect of using these vessels for microvascular anastamosis. We report our experience of deliberately using these vessels in an osseocutaneous radial forearm free flap reconstruction. Although there are some technical considerations to bear in mind, we would suggest that unlike vessels affected by atherosclerosis, anastomosis of arteries affected by Mönckeberg's arteriosclerosis has little or no impact on free flap survival.

A developmentally inspired combined mechanical and biochemical signaling approach on zonal lineage commitment of mesenchymal stem cells in articular cartilage regeneration.

Articular cartilage is organized into multiple zones including superficial, middle and calcified zones with distinct cellular and extracellular components to impart lubrication, compressive strength, and rigidity for load transmission to bone, respectively. During native cartilage tissue development, changes in biochemical, mechanical, and cellular factors direct the formation of stratified structure of articular cartilage. The objective of this work was to investigate the effect of combined gradients in cell density, matrix stiffness, and zone-specific growth factors on the zonal organization of articular cartilage. Human mesenchymal stem cells (hMSCs) were encapsulated in acrylate-functionalized lactide-chain-extended polyethylene glycol (SPELA) gels simulating cell density and stiffness of the superficial, middle and calcified zones. The cell-encapsulated gels were cultivated in a medium supplemented with growth factors specific to each zone and the expression of zone-specific markers was measured with incubation time. Encapsulation of 60 × 10(6) cells per mL hMSCs in a soft gel (80 kPa modulus) and cultivation with a combination of TGF-β1 (3 ng mL(-1)) and BMP-7 (100 ng mL(-1)) led to the expression of markers for the superficial zone. Conversely, encapsulation of 15 × 10(6) cells per mL hMSCs in a stiff gel (320 MPa modulus) and cultivation with a combination of TGF-β1 (30 ng mL(-1)) and hydroxyapatite (3%) led to the expression of markers for the calcified zone. Further, encapsulation of 20 × 10(6) cells per mL hMSCs in a gel with 2.1 MPa modulus and cultivation with a combination of TGF-β1 (30 ng mL(-1)) and IGF-1 (100 ng mL(-1)) led to up-regulation of the middle zone markers. Results demonstrate that a developmental approach with gradients in cell density, matrix stiffness, and zone-specific growth factors can potentially regenerate zonal structure of the articular cartilage.

Anisotropic temperature sensitive chitosan-based injectable hydrogels mimicking cartilage matrix.

In this study, chitosan-based hydrogels were formulated with material similarities to three of the four zones of articular cartilage. Gelatin, hyaluronic acid (HA), and β-tricalcium phosphate for the superficial, radial, and calcified zones, were blended in different amounts and tested for formation of uniform solution, gelability, and rheological characteristics. Confined compression in two configurations (series and parallel to anisotropy), and cyclical tests were performed at the physiological conditions. In vivo gelation and systemic effects were evaluated in male BALB/c mice subcutaneous model. At day 5, hydrogels were harvested along with the adjoining skin and analyzed by histology. Formulations that produced solutions after pH adjustments were selected for each zone. Anisotropic hydrogels were formed by mixing solutions from each zone, which showed uniform gradation. Addition of HA improved structural integrity relative to other formulations. When hydrogels were in series, combined hydrogel modulus was the average of all zones while that in parallel orientation was half of that series orientation. Cyclical tests demonstrated repeatable strength and durability. All formulations were injectable into the subcutaneous region. H/E stained tissues showed minimal invasion of inflammatory cells in radial and calcified zones. Structural integrity of the hydrogel is suggested to be the resultant of the presence of HA.

Histological changes caused by continuous ultrasonic waves of 01 MHz on the epiphyseal disk of murine tibia

AIMS: To evaluate the histological changes in the epiphyseal disk of the tibia in growing mice during exposure to ultrasonic waves of 1MHz frequency.&#x0D; &#x0D; METHODS: Experimental study of quantitative and randomized nature. For this purpose, 16 Swiss albino mice of the Webster strain, during growth, aged 3 weeks and weighing 10-15 g were randomly divided into two groups of eight animals each and exposed to ultrasonic waves in the right tibia. Groups D1 (exposed to a dose of 0.1 W/cm2) and D2 (exposed dose of 0.5 W/cm2) were assigned. The control group, designated C, was represented by 16 samples relating to the contralateral side of each animal of groups D1 and D2. Procedures occurred in an application/day, totaling 10 applications without interruption. The samples were analyzed histological by hematoxylin-eosin staining techniques, Alcian blue and Von Kossa stains, in order to measure the area of the head versus the tibia epiphysial disc, beyond the thickness of the proliferative and hypertrophic zones. For statistical analysis we used the Friedman test with Bonferroni correction.&#x0D; &#x0D; RESULTS: The bone tissue was influenced by ultrasonic waves, as well as the epiphyseal plate, where the group had the largest parameter of the total area larger than the other disc. The calcified zone was not influenced by the ultrasonic waves, but the proliferative zone was sensitive to this therapy, especially at the lowest dose. No pathological changes were found in the treated and control groups.&#x0D; &#x0D; CONCLUSIONS: With the parameters used in this study positive action of ultrasound was observed in the cartilage and bone tissue formation, not being possible to determine what was the intensity level of the application of ultrasound statistically more significant.

The N-Terminal Cleavage of Chondromodulin-I in Growth-Plate Cartilage at the Hypertrophic and Calcified Zones during Bone Development

Chondromodulin-I (ChM-I) is a 20–25 kDa anti-angiogenic glycoprotein in cartilage matrix. In the present study, we identified a novel 14-kDa species of ChM-I by immunoblotting, and purified it by immunoprecipitation with a newly raised monoclonal antibody against ChM-I. The N-terminal amino acid sequencing indicated that it was an N-terminal truncated form of ChM-I generated by the proteolytic cleavage at Asp37-Asp38. This 14-kDa ChM-I was shown by the modified Boyden chamber assay to have very little inhibitory activity on the VEGF-A-induced migration of vascular endothelial cells in contrast to the intact 20–25 kDa form of ChM-I (ID50 = 8 nM). Immunohistochemistry suggested that 20–25 kDa ChM-I was exclusively localized in the avascular zones, i.e. the resting, proliferating, and prehypertrophic zones, of the cartilaginous molds of developing long bone, whereas the 14-kDa form of ChM-I was found in hypertrophic and calcified zones. Immunoblotting demonstrated that mature growth-plate chondrocytes isolated from rat costal cartilage actively secrete ChM-I almost exclusively as the intact 20–25 kDa form into the medium in primary culture. Taken together, our results suggest that intact 20–25 kDa ChM-I is stored as a component of extracellular matrix in the avascular cartilage zones, but it is inactivated by a single N-terminal proteolytic cleavage in the hypertrophic zone of growth-plate cartilage.

0374 Leptin and its receptor in human calcific aortic valve: expression and functional implications

Calcific aortic valve disease (CAVD) affects 2% to 6% of population over 65 years in industrialized countries. CAVD, assimilated to an “atherosclerosis-like” disease, results from dysregulated mechanisms such as fibrosis, inflammation and calcification. These mechanisms could be enhanced by the osteoblastic differentiation of the valvular interstitial cells (VIC), the most prevalent cell type in the human aortic valves. Leptin, known as the product of obesity gene, has recently been linked to aortic valve calcification in ApoE -/- mice. Our hypothesis is that leptin could play an important role in the calcifying processes implicated in CAVD via direct effects on VIC phenotype in humans. Expression of leptin and its receptor was analysed by RT-qPCR, immunohistochemistry and ELISA in 74 human calcific aortic valves and in 10 normal valves. VIC isolated from calcific and normal aortic valves were also analysed. The effects of leptin on the expression of osteoblastic markers such as osteopontin, osteonectin, alkaline phosphatase, BMP-2 and BMP-7 in cultured VIC were investigated. The presence of leptin and its long, functional, receptor isoform (ObR-L) was observed in human aortic valves with an overexpression of leptin in calcified vs non-calcified valvular zones (114.52 pg/mg (47.96- 234.55) vs 66.95 pg/mg (42.76-137.15), p<0.05). Moreover, ObR-L and leptin were constitutively expressed in VIC with a higher ObR-L expression in VIC isolated from pathological vs normal valves (p<0.05). Finally, stimulation of VIC by leptin at physiological concentrations (20 to 100 ng/mL) significantly increased osteonectin (pro-calcifying) and decreased BMP-7 (anti-calcifying) expression levels. Taken together, these novel findings suggest that leptin could be involved in the process of human valve calcification by modifying VIC phenotype.

Dystrophic Calcification and Accentuated Localized Argyria After Fractionated Carbon Dioxide Laser Therapy of Hypertrophic Scars

Fractionated, ultrapulsed carbon dioxide (CO2) laser therapy is a powerful tool for the treatment of scars. Common adverse effects of this therapeutic modality have been previously documented. We describe 2 unreported adverse effects of ultrapulsed CO2 laser treatment of mature scars in a patient previously treated with silver-impregnated dressings. A teenage survivor of toxic epidermal necrolysis presented with faint but diffuse dyschromia clinically and histologically consistent with localized argyria secondary to silver-impregnated dressings used years earlier. The patient was subsequently treated with fractionated CO2 for her scarring, but her hyperpigmentation worsened with each treatment. A subsequent biopsy specimen revealed a zone of dystrophic calcification with adjacent pseudo-ochronotic fibers that were not appreciated on biopsy specimens taken before CO2 laser treatment, suggesting unique complications not previously reported. We present 2 unique complications secondary to ultrapulsed, fractionated CO2 laser treatment in a patient previously treated with silver-impregnated dressings: (1) the appearance of pseudo-ochronotic fibers in areas of worsening pigmentation and (2) evidence of dystrophic calcification limited to columns of fractionated laser ablation. Therefore, a history of argyria or treatment with silver-impregnated dressings should be considered before treatment with fractionated CO2 lasers.

Toward regeneration of articular cartilage

Articular cartilage is classified as permanent hyaline cartilage and has significant differences in structure, extracelluar matrix components, gene expression profile, and mechanical property from transient hyaline cartilage found in the epiphyseal growth plate. In the process of synovial joint development, articular cartilage originates from the interzone, developing at the edge of the cartilaginous anlagen, and establishes zonal structure over time and supports smooth movement of the synovial joint through life. The cascade actions of key regulators, such as Wnts, GDF5, Erg, and PTHLH, coordinate sequential steps of articular cartilage formation. Articular chondrocytes are restrictedly controlled not to differentiate into a hypertrophic stage by autocrine and paracrine factors and extracellular matrix microenvironment, but retain potential to undergo hypertrophy. The basal calcified zone of articular cartilage is connected with subchondral bone, but not invaded by blood vessels nor replaced by bone, which is highly contrasted with the growth plate. Articular cartilage has limited regenerative capacity, but likely possesses and potentially uses intrinsic stem cell source in the superficial layer, Ranvier's groove, the intra-articular tissues such as synovium and fat pad, and marrow below the subchondral bone. Considering the biological views on articular cartilage, several important points are raised for regeneration of articular cartilage. We should evaluate the nature of regenerated cartilage as permanent hyaline cartilage and not just hyaline cartilage. We should study how a hypertrophic phenotype of transplanted cells can be lastingly suppressed in regenerating tissue. Furthermore, we should develop the methods and reagents to activate recruitment of intrinsic stem/progenitor cells into the damaged site.

Vigorous periosteal reaction secondary to copper deficiency in an infant on total parenteral nutrition

Sir, We are using this forum to share our recent experience with a 5-month-old child who had dramatic radiographic findings, in order to alert health care workers, including pediatric radiologists, about the increased likelihood of seeing copper deficiency cases in view of the scarcity of parenteral copper. Depending on the supply available, parenteral copper has been frequently omitted from total parenteral nutrition formulations in the last year or so [1]. A 5-month-old boy was transferred to our hospital for intestinal rehabilitation and evaluation for possible intestinal transplantation. He had been born at a gestational age of 34 weeks secondary to preterm labor. Prenatal US was concerning for polyhydramnios and intestinal obstruction. At birth he was found to have multiple small bowel atresias. After multiple surgeries for resections with reanastomosis, he remained dependent on parenteral nutrition and intolerant of enteral feeds. The child had been hospitalized since birth. The original chest/abdomen radiograph at our hospital showed vigorous periosteal reaction of humeri and femurs (sparing the region of the metaphysis) (Fig. 1) along with periosteal reaction of some ribs and the scapulae. Our original differential diagnoses included diffuse osteomyelitis, hypertrophic pulmonary osteoarthropathy, scurvy, vitamin D deficiency and child abuse. A bone survey then revealed osteoporosis (for example, the zone of provisional calcification of knee secondary centers could be seen better than the bone inside) and multiple metaphyseal abnormalities appearing as spurs (Fig. 2) and similar in some locations to “bucket handle” classic metaphyseal lesions of child abuse (Fig. 3). The pattern was somewhat suggestive of scurvy, a disease rarely seen in current practice. The other diagnoses considered were Menkes syndrome (kinky hair syndrome) and copper deficiency from inadequate intake. Vitamin C levels were normal, eliminating scurvy as a diagnosis. Recognition of the national shortage of intravenous micronutrients and trace elements led to evaluation of intake of copper in this boy with intestinal failure who had been dependent on total parenteral nutrition his entire life. It was discovered that because of the national shortage of parenteral copper, he had not received any copper at the referring hospital. A serum copper level showed a very low level of <10 μg/dL (normal 75–165 μg/dL), a very low ceruloplasmin level <4 mg/dL (normal 18–37 mg/dL), and alkaline phosphatase of 1,324 IU/L (normal 82–383 IU/L). Vitamin D, calcium and phosphorus levels were normal. He was anemic (hemoglobin 8.3 g/dL) on admission with normal white blood cell count. In the absence of any enteral feeds and knowledge of absence of copper in the total parenteral nutrition, the radiologic changes were attributed to acquired copper deficiency. Radiologic findings in this boy are consistent with prior reports of profound infantile copper deficiency [2, 3]. The exuberant extent of the periosteal reaction may well relate to the many weeks of normal in-patient handling of the baby without realizing he was bleeding subperiosteally. Copper deficiency leads to reduction of the activity of numerous copper-containing enzymes that are necessary for the structure and function of bone, skin, hair, blood vessels and the nervous system [4]. Children with deficient copper A. E. Oestreich (*) Department of Radiology, Cincinnati Children’s Hospital Medical Center Radiology 5031 CCHMC, 3333 Burnet Ave., Cincinnati, OH 45229-3039, USA e-mail: alan.oestreich@cchmc.org

Discordant radiologic and histological dimensions of the zone of provisional calcification in fetal piglets

Studies have shown that the fracture plane of the classic metaphyseal lesion (CML) of infant abuse occurs in the region of the primary spongiosa, encompassing a radiodense fracture fragment customarily referred to as the "zone of provisional calcification" or ZPC. However, the zone of provisional calcification is defined differently in the pathology and the imaging literature, potentially impeding efforts to understand the fundamental morphological features of the classic metaphyseal lesion. We systematically correlated micro-CT data with histology in piglets to explore the differing definitions of the zone of provisional calcification and to elucidate the anatomical basis for divergent definitions. The distal tibias of five normal fetal piglets were studied postmortem. The specimens were resected and imaged with digital radiography (50μm resolution) and micro-CT (45μm(3) isotropic resolution). Image processing techniques were applied to the micro-CT data for visualization and data analysis. The resected tissue specimens were then processed routinely and the light microscopic features were correlated with the imaging findings. The longitudinal dimension of the radiologic zone of provisional calcification is greater than the histological ZPC, and these dimensions are statistically distinct (P < 0.0002). The radiologic zone of provisional calcification consists of two adjoining mineralized discoid regions that span the chondro-osseous junction-a thick discoid region that encompasses the densest region of the primary spongiosa, and a thin discoid region (corresponding to the histological ZPC) that is situated in the base of the physis adjacent to the metaphysis. The correlation of the normal histology and micro-CT appearance of this dynamic and complex region provides an anatomical foundation upon which a deeper appreciation of the morphology of the classic metaphyseal lesion can be built.

Layering PLGA-based electrospun membranes and cell sheets for engineering cartilage-bone transition

It is now widely acknowledged that implants that have been designed with an effort towards reconstructing the transition between tissues might improve their functionality and integration in vivo. This paper contributes to the development of improved treatment for articular cartilage repair by exploring the potential of the combination of electrospinning technology and cell sheet engineering to create cartilage tissue. Poly(lactic-co-glycolic acid) (PLGA) was used to create the electrospun membranes. The focus being on the cartilage-bone transition, collagen type I and hydroxyapatite (HA) were also added to the scaffolds to increase the histological biocompatibility. Human mesenchymal stem cells (hMSCs) were cultured in thermoresponsive dishes to allow non-enzymatic removal of an intact cell layer after reaching confluence. The tissue constructs were created by layering electrospun membranes with sheets of hMSCs and were cultured under chondrogenic conditions for up to 21 days. High viability was found to be maintained in the multilayered construct. Under chondrogenic conditions, reverse-transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry have shown high expression levels of collagen type X, a form of collagen typically found in the calcified zone of articular cartilage, suggesting an induction of chondrocyte hypertrophy in the PLGA-based scaffolds. To conclude, this paper suggests that layering electrospun scaffolds and cell sheets is an efficient approach for the engineering of tissue transitions, and in particular the cartilage-bone transition. The use of PLGA-based scaffold might be particularly useful for the bone-cartilage reconstruction, since the differentiated tissue constructs seem to show characteristics of calcified cartilage.

The expression of glycophorin A and osteoprotegerin is locally increased in carotid atherosclerotic lesions of symptomatic compared to asymptomatic patients

The aim of this study was to evaluate in detail the histopathological characteristics of endarterectomized carotid atherosclerotic lesions in symptomatic versus asymptomatic patients. Twenty carotid lesions, 10 from asymptomatic and 10 from symptomatic patients who underwent carotid endarterectomy were classified according to histomorphological features. Samples were analyzed for intraplaque localization and for the expression of proteins associated with inflammation, such as CD68, interleukin (IL)-1β, tumor necrosis factor-α (TNF-α), pentraxin-3 (PTX-3), nuclear factor-κB (NF-κB), C-reactive protein (CRP) and transforming growth factor-β (TGF-β), as well as for proteins associated with vascular remodelling, such as matrix-metalloproteinase-9 (MMP-9), glycophorin A (GYPA), osteoprotegerin (OPG), vascular cell adhesion molecule-1 (VCAM-1), endothelin-1 (ET-1), vascular endothelial growth factor (VEGF) and vascular smooth muscle cell actin (VSMA). Corresponding expression scores were compared between the symptomatic and asymptomatic patients and evaluated statistically. The expression of all 14 evaluated markers was significantly elevated in the border zone adjacent to the mixed plaque compared with the unaffected control area of the same sample (p<0,016). The expression scores of GYPA and OPG were significantly higher in the border zones around the calcified (GYPA, p=0.035; OPG, p=0.043) and mixed (GYPA, p<0.001; OPG, p=0.007) plaque zones of symptomatic patients compared to asymptomatic patients. No difference in expression scores was observed for any of the analyzed inflammatory marker proteins between the border zones of symptomatic and asymptomatic patients. In conclusion, the increased expression of GYPA, indicating intraplaque hemorrhage, and OPG, indicating the transdifferentiation of vascular cells, in carotid atherosclerotic lesions may be associated with an increased risk of plaque instability.

Mechanical evaluation of a tissue-engineered zone of calcification in a bone–hydrogel osteochondral construct

The objective of this study was to test the hypothesis that mechanical properties of artificial osteochondral constructs can be improved by a tissue-engineered zone of calcification (teZCC) at the bone–hydrogel interface. Experimental push-off tests were performed on osteochondral constructs with or without a teZCC. In parallel, a numerical model of the osteochondral defect treatment was developed and validated against experimental results. Experimental results showed that the shear strength at the bone–hydrogel interface increased by 100% with the teZCC. Numerical predictions of the osteochondral defect treatment showed that the shear stress at the bone–hydrogel interface was reduced with the teZCC. We conclude that a teZCC in osteochondral constructs can provide two improvements. First, it increases the strength of the bone–hydrogel interface and second, it reduces the stress at this interface.

Spatial periodicity in growth plate shear mechanical properties is disrupted by vitamin D deficiency

The growth plate is a highly organized section of cartilage in the long bones of growing children that is susceptible to mechanical failure as well as structural and functional disruption caused by a dietary deficiency of vitamin D. The shear mechanical properties of the proximal tibial growth plate of rats raised either on normal or vitamin D and calcium deficient diets were measured. A sinusoidal oscillating shear load was applied to small excised growth plate specimens perpendicular to the direction of growth while imaging the deformation in real time with a fast confocal microscope. Local deformations and shear strains were quantified using image correlation. The proliferative zone of the growth plate bores the majority of the shear strain and the resting, hypertrophic and calcification zones deformed less. Surprisingly, we regularly observed discontinuous deformations in the proliferative zone in both groups that resembled cell columns sliding past one another in the direction of growth. These discontinuities manifested as regions of concentrated longitudinal shear strain. Furthermore, these shear strain concentrations were spaced evenly in the proliferative zone and the spacing between them was similar across growth plate regions and across control specimens. In contrast to the healthy controls, the vitamin D deficient growth plate exhibited larger variations in the size and orientation of cellular columns in the proliferative and hypertrophic zones. High strains were observed between columns, much as they were in the controls. However, the regular spacing of shear strain concentrations was not preserved, echoing the observation of decreased structural organization.

Bisphosphonate therapy for chronic recurrent multifocal osteomyelitis

This patient was diagnosed with chronic multifocal recurrent osteomyelitis of the ankle approximately 4 years prior to the presented images. Multiple treatments were administered, most notably monthly intravenous pamidronate, a therapy that was continued for 17 months, until recrudescent disease prompted a change in therapy. At the time of the presented images, the patient had not received bisphosphonate therapy for 2 years. Bisphosphonates are a class of medications used primarily in adult patients with decreased bone mineral density. When the drug is incorporated into the hydroxyapatite matrix of nascent bone, osteoclastic activity is inhibited, and as a result, bone mineral density increases. Also used in patients with expansile bone lesions, the decrease in osteoclastic activity serves to slow the progression of enlargement, demonstrating an analgesic effect. Finally, it is thought that bisphosphonates engender a direct anti-inflammatory response by altering the inflammatory cascade [1]. In children, bisphosphonates have been used for the treatment of conditions resulting in low bone mineral density, such as osteogenesis imperfecta [2]. Furthermore, several recent studies have demonstrated the effectiveness of bisphosphonate therapy as a second-line treatment for CRMO-associated pain [3]. Most CRMO treatment regimens consist of daily infusions of IV pamidronate for 1–3 days, repeated every 3 months [1, 3, 4]. Pain relief is usually rapid, within days of initiation, and resolution of inflammation by MR imaging follows in up to 80 % of patients [2]. Radiologically, the cyclical inhibition of osteoclast activity in the skeletally immature patient gives rise to dense metaphyseal bands migrating longitudinally from the zone of provisional calcification into the diaphysis, as previously described [5, 6]. Additionally, undertubulation of the affected areas, as well a “bone within a bone” appearance can be seen [7]. The presence of metaphyseal bands engenders differential considerations such as lead poisoning and hypervitaminosis D, as well as causes of standard growth recovery lines (times of malnutrition, severe illness, etc.). As none of the causes of metaphyseal bands are common, the interpreting radiologist may be inspired to suggest further testing when confronted with this unusual finding. However, the recognition of focal undertubulation (most striking at the distal femur in this case) helps cinch the diagnosis. Recognition of these findings can be clinically important, as the abrupt change of bone mineral density at the dense bands has been implicated as a stress riser, similar to the interface between a prosthesis and native bone [8]. The potential increase in fracture risk presented by these bands may prompt a change in the duration of therapy.

Lesions at the junction between uncalcified and calcified cartilage zones during the development of osteoarthritis

Lesions at the junction between uncalcified and calcified cartilage zones during the development of osteoarthritis

Platelets express and release osteocalcin and co‐localize in human calcified atherosclerotic plaques

Although vascular-calcification mechanisms are only partially understood, the role of circulating calcifying cells and non-collagenous bone matrix proteins in the bone-vascular axis is emerging. In spite of the fact that platelets represent a cellular interface between hemostasis, inflammation and atherosclerosis, and have a myeloid precursor, a possible involvement in the modulation of vascular calcification has rarely been investigated. We investigated if osteocalcin (OC) is released by platelets and described OC expression in patients with carotid artery occlusive disease. Expression and release of OC were determined by Western blot, immunofluorescence, fluorescence-activated cell sorting (FACS) and ELISA in human resting and activated platelets and megakaryocytes. Co-localization of platelet aggregates, macrophages, OC and calcifications was studied in carotid endarterectomy specimens and normal tissues. Human platelets expressed OC and co-localized with CD63 in δ-granules. Upon activation with an endogenous mechanism, platelets released OC in the extracellular medium. Expression of OC in megakaryocytes suggested lineage specificity. The OC count in circulating platelets and the released amount were significantly higher in patients with carotid artery occlusive disease than in healthy controls (P < 0.0001) in spite of similar serum levels. In atherosclerotic plaques, OC strongly overlapped with CD41+ platelets in the early stage of calcification, but this was not seen in normal tissues. CD68+OC+ cells were present at the periphery of the calcified zone. Given the active role played by platelets in the atherosclerotic process, the involvement of OC release from platelets in atherosclerotic lesions and the impact of genetic and cardiovascular risk factors in mediating bone-marrow preconditioning should be investigated further.

Aortic endovascular repair modeling using the finite element method

Aim: The goal is to simulate different stages of the endovascular procedure in the preoperative phase. Methods: We have developed a numerical model of the endovascular treatment of abdominal aortic aneurysms (AAA) using finite element analysis (FEA), we took into account the geometry of the biological region reconstructed from scans, a local characterization of the guidewire/catheter mechanical properties, a mapping of material properties depending on the degree of calcification, a real behavior of the vascular walls, and a projection of the aorta environment. Results: Our results present the endovascular system navigation from the femoral artery to the neck of the aneurysm, predict the deformation of femoral, iliac and aortic arteries while driving flexible and stiff endovascular devices, and detect stress concentration due to tortuous and calcified artery zones. A given group of patients with very angulated and calcified arteries were validated, based on a tuning between clinical data and our endovascular simulation. Conclusion: Our model allows controlling with accuracy the delivery system rise during surgery, predicting the feasibility of the surgery with reliability as well as choose the best guide for each patient, taking into account the risk of rupture of calcified areas in the case of high angulations, and using planning simulated images with deformation of artery walls to propose stent sizing.

Osteochondral Interface Generation by Rabbit Bone Marrow Stromal Cells and Osteoblasts Coculture

Physiological osteochondral interface regeneration is a significant challenge. This study aims to investigate the effect of the coculture of chondrogenic rabbit bone marrow stromal cells (rBMSCs) with rabbit osteoblasts in a specially designed two-dimensional (2D)-three-dimensional (3D) co-interface culture to develop the intermediate osteochondral region in vitro. The 2D-3D coculture system was set up by first independently culturing chondrogenic rBMSCs on a scaffold and osteoblasts in cell culture plates, and subsequently placed in contact and cocultured. As control, samples not cocultured with osteoblasts were used. The regulatory effects exerted by osteoblasts on chondrogenic rBMSCs were quantified by real-time polymerase chain reaction. To study the effect of coculture on cells located in different parts of the scaffold, samples were separated into two parts and significantly different gene expression patterns were found between them. In comparison with the control group, a significant moderate downregulation of chondrogenic marker genes, such as Collagen II and Aggrecan was observed. However, the Sox-9 and Collagen I expression increased. More importantly, chondrogenic rBMSCs in the coculture system were shown to form the osteochondral interface layer by expressing calcified cartilage zone specific extracellular matrix marker Collagen X and the hypertrophic chondrocyte marker MMP-13, which were not observed in the control group. Specifically, only the chondrogenic rBMSC layer in contact with the osteoblasts expressed Collagen X and MMP-13, indicating the positive influence of the coculture upon interface formation. Biochemical analyses, histology results, and immunohistochemical staining further supported this observation. In conclusion, this study revealed that specific regulatory stimulations from osteoblasts in the 2D-3D interface coculture system could induce the formation of ostochondral interface for the purpose of osteochondral tissue engineering.