Calcium Crystals Research Articles

The role of ank in catabolic events of articular chondrocytes

s / Osteoarthritis and Cartilage 22 (2014) S57–S489 S133 210 THE ROLE OF ANK IN CATABOLIC EVENTS OF ARTICULAR CHONDROCYTES T. Kirsch y, T. Minashima y, K. Campbell y, S. Hadley z, Y. Zhang x. yNYU Sch. of Med., New York, NY; zBrigham Women’s Hosp., Harvard Med. Sch., Boston, MA, USA; xCtr. for Joint Surgery, Southwest Hosp., Third Military Med. Univ., Chong Qing, China Purpose: The progressive ankylosis protein (ANK) is a transmembrane protein that transports intracellular pyrophosphate (PPi) to the extracellular milieu. Lack of functional ANK in ank/ank mice or ank knockout mice leads to pathological mineralization of articular cartilage, ligaments, menisci and tendons resulting in arthritic changes because of the reduced amounts of extracellular PPi, a main mineralization inhibitor. We and others, however, have shown that ANK is expressed at low levels in healthy articular cartilage, and ANK expression levels drastically increase during osteoarthritis (OA) initiation and progression. These findings raise the possibility that ANK plays more complex and unknown functions in articular cartilage and OA pathology than only preventing basic calcium crystal formation. Therefore, the purpose of this study was to determine the role of ANK in catabolic events of articular chondrocytes. Methods: The interactions of ANK with Myb-binding protein 1a (MYBBP1a) and sphingosine kinase 1 (SPHK1)were identified using yeast two-hybrid screening and co-immunoprecipitation. To determine the role of these interactions in catabolic events of articular chondrocytes, ank/ank and wild type (WT) mouse chondrocytes or femoral heads were treated with interleukin-1beta (IL-1b) in the absence or presence of the SPHK inhibitor, SKI-II. Catabolic marker mRNA levels were analyzed by real time PCR; proteoglycan loss using safranin O staining and MMP-13 immunostaining were determined in femoral head explants; NF-kB activity was determined by transfecting chondrocytes with a NF-kBspecific luciferase reporter and analyzing nuclear translocation of p65 by immunoblotting; MYBBP1a nuclear or cytoplasmic amounts were determined by immunohistochemistry and immunoblotting. Results: Yeast two-hybrid screening and co-immunoprecipitation experiments revealed that the N-terminal region of ANK interacts with SPHK1, whereas the C-terminal region interacts withMYBBP1a. MYBBP1a isa cytoplasmicprotein that canshuttle into thenucleuswhere it represses NF-kB transcriptional activity. Consequently, the loss of ANK/MYBBP1a interactions resulted in increased MYBBP1a nuclear amounts and decreasedNF-kB activity in IL-1b-treated ank/ank chondrocytes compared to IL-1b-treated WT cells. The loss of ANK/SPHK1 interaction resulted in decreased SPHK1 activity in IL-1b-treated ank/ank chondrocytes compared to IL-1b-treated WT cells. Activation of NF-kB-dependent transcriptional activity by IL-1b involves translocation of NF-kB family members, in particular p65 (RelA) and p50, from the cytoplasm to the nucleus. The increase of the nuclear p65 amount and the decrease of the cytoplasmic p65 amount in IL-1b-treated ank/ank chondrocytes were markedly reduced compared to IL-1b-treatedWTcells. The SPHK inhibitor, SKI-II reduced the amount of nuclear p65 and increased the amount of cytoplasmicp65 in IL-1b-treatedWTchondrocytes butnot in IL-1b-treated ank/ank chondrocytes. Transfection with ank expression vector reduced nuclearMYBBP1a amounts and fully restored SPHK andNF-kB activities in IL-1b-treated ank/ank chondrocytes, whereas transfection with P5L mutant ank that doesnot interactwith SPHK1or F376del ank thatdoes not interact with MYBBP1a increased SPHK activity or reduced nuclear MYBBP1a, respectively, and consequently either transfection with P5L or F376del ank only partially restored NF-kB activity. Finally, lack of ANK/ MYBBP1a and SPHK1 interactions in ank/ank chondrocytes resulted in decreased catabolic marker mRNA levels, proteoglycan loss, and MMP-13 immunostaining in IL-1b-treated articular chondrocytes or femoral heads. Conclusion: This study shows that ANK interacts via the N-terminal and C-terminal cytoplasmic regions with SPHK1 and MYBBP1a, respectively. The ANK/MYBBP1a interaction results in the inhibition of the nuclear translocation of MYBBP1a. Nuclear MYBBP1a acts as a corepressor of NF-kB transcriptional activity. The ANK/SPHK1 interaction is a major regulator of SPHK1 activity. Both interactions stimulate NF-kB activity and ultimately catabolic events in IL-1b-treated articular chondrocytes. Since IL-1b and NF-kB play major roles in OA pathogenesis, our findings suggest that increased ANK expression in OA cartilage may be a major contributor to cartilage destruction. Future experiments are needed to fully understand the mechanisms of the interactions between ANK, MYBBP1a and SPHK1 in OA pathology and develop strategies to interfere with these interactions as a potential novel therapy for the treatment of OA. 211 OSX-CRE DIRECTED ABLATION OF SITE-1 PROTEASE IN MICE RESULTS IN KYPHOSIS AND SCOLIOSIS D. Patra, E. DeLassus, L.J. Sandell.Washington Univ. Sch. of Med., St. Louis,

Diagnosis and Clinical Manifestations of Calcium Pyrophosphate and Basic Calcium Phosphate Crystal Deposition Diseases

Basic calcium phosphate and pyrophosphate calcium crystals are the 2 main calcium-containing crystals that can deposit in all skeletal tissues. These calcium crystals give rise to numerous manifestations, including acute inflammatory attacks that can mimic alarming and threatening differential diagnoses, osteoarthritis-like lesions, destructive arthropathies, and calcific tendinitis. Awareness of uncommon localizations and manifestations such as intraspinal deposition (eg, crowned dens syndrome, tendinitis of longus colli muscle, massive cervical myelopathy compression) prevents inappropriate procedures and cares. Coupling plain radiography, ultrasonography, computed tomography, and synovial fluid analysis allow accurate diagnosis by directly or indirectly identifying the GRAAL of microcrystal-related symptoms.

Clinical implications of pathogenic calcium crystals

The role of intraarticular calcifications has been challenged over the past years, with respect to cartilage destruction, especially in osteoarthritis. Main themes reviewed in this article will discuss prevalence of articular cartilage calcifications, mechanisms of cartilage calcifications, and mostly the pathogenic role of both calcium pyrophosphate and basic calcium phosphate crystals. A direct pathogenic role of both calcium crystals has been depicted, in cartilage as a crystal-induced stress, or via acute or chronic crystal-induced synovitis.

CP-117 Severe calcinosis cutis successfully treated with a topical W/O emulsion of 10% sodium thiosulfate

BackgroundCalcinosis is caused by accumulation of calcium salts in the tissues, with subcutaneous nodules, atrophy and ulceration over the affected area. Treatment with intravenous sodium thiosulfate has been used, since...

Nonpharmacologic and Pharmacologic Management of CPP Crystal Arthritis and BCP Arthropathy and Periarticular Syndromes

Calcium crystal arthritis is often unrecognized, poorly managed, and few effective therapies are available. The most common types of calcium crystals causing musculoskeletal syndromes are calcium pyrophosphate (CPP) and basic calcium phosphate (BCP). Associated syndromes have different clinical presentations and divergent management strategies. Acute CPP arthritis is treated similarly to acute gouty arthritis, whereas chronic CPP and BCP arthropathy may respond to strategies used for osteoarthritis. Calcific tendonitis is treated with a variety of interventions designed to dissolve BCP crystals. A better understanding of the causes and larger well-planned trials of current therapies will lead to improved care.

The Structural Consequences of Calcium Crystal Deposition

Calcium pyrophosphate dihydrate and basic calcium phosphate (BCP) crystals are the most common calcium-containing crystals associated with rheumatic disease. Clinical manifestations of calcium crystal deposition include acute or chronic inflammatory and degenerative arthritides and certain forms of periarthritis. The intra-articular presence of BCP crystals correlates with the degree of radiographic degeneration. Calcium crystal deposition contributes directly to joint degeneration. Vascular calcification is caused by the deposition of calcium hydroxyapatite crystals in the arterial intima. These deposits may contribute to local inflammation and promote further calcification, thus aggravating the atherosclerotic process. Calcium crystal deposition results in substantial structural consequence in humans.

Characterisation of calcium crystals in Abelia spp. using X-ray diffraction and electron microscopy

SummaryThe localisation, chemical composition, and morphology of calcium (Ca) oxalate crystals in the leaves and stems of Abelia mosanensis and of A. × grandiflora were analysed using a variable pressure-scanning electron microscope (VP-SEM) equipped with an X-ray diffraction system, a low temperature SEM (LT-SEM), and a transmission electron microscope (TEM). Foliar analyses of macro- and micro-elements were performed on the leaves and stems of A. mosanensis. A greater number of Ca oxalate crystals were observed in A. mosanensis than in A. × grandiflora. Three morphologically distinguishable types of crystals were observed: the prismatic crystals found inside the chloroplast, multifaceted star-like spherical and bladed aggregate crystals (druses) inside the vacuoles of the mesophyll cells, and small angular crystals (sand crystals) inside the cuticle. Semi-solid crystals that may drip and accumulate to become a solid Ca oxalate crystal were observed by LT-SEM, which indicated the growth of druses of Ca oxalate crystals. The growth of prismatic crystals and of druses were evident through the formation of crystalline lamellae. Micro-analysis indicated that the crystals were Ca oxalate and contained magnesium in A. mosanensis, or silicon in A. × grandiflora. Abelia stems with low foliar calcium concentrations showed no Ca oxalate crystal formation. This is the first report, to our knowledge, on the presence and possible growth of crystals of different morphologies and chemical compositions in Abelia.

Leaf anatomical traits corroborate the leaf economic spectrum: a case study with deciduous forest tree species

Deciduous forests are seasonal systems that occur scattered throughout the Neotropics. Many aspects about these ecosystems have been studied. However, there is a lack of information about leaf structure and its possible functional and adaptive aspects. Here we examined leaf anatomy and specific leaf area (SLA) in 13 dominant tree species of dry forests in central Brazil, identifying structural patterns of these species. As the system is seasonal, with species presenting a deciduous behavior, we expect to find tree species with a set of mesomorphic leaf traits. The studied dry forest trees share similar leaf structure, with one-layered adaxial and abaxial epidermis and a well-developed mesophyll, high values of SLA, and the presence of calcium crystals and mucilaginous cells. Higher values of SLA and the high investment achieved in the mesophyll (especially in parenchyma tissue) are probably related to the accomplishment of a high performance of carbon and nutrients gain during the limited wet season.

In vitro Dissolution of Calcium Oxalate Stones with Ethylenediaminetetraacetic Acid and Snake Venom Thrombin-Like Enzyme

Objective: The aim of this study was to determine the feasibility of using snake venom thrombin-like enzyme (SVTLE) and/or ethylenediaminetetraacetic acid (EDTA) to dissolve calcium oxalate stones in vitro. Methods: Seven calcium oxalate stones were incubated with various chemolytic agents [EDTA, Tris-HCl/EDTA (TE) buffer or SVTLE diluted in TE buffer]. The pH, calcium concentration, stone weight and stone surface integrity were recorded, as well as related pathological changes to bladder mucosae. Results: Compared to all other solutions, those containing SVTLE and buffered EDTA had higher concentrations of mobilized calcium and caused significantly more stone weight loss, stone fragility and gaps in the calcium crystals. Also, there were no adverse pathological effects on rabbit bladder mucosae from any of the solutions. Conclusions: The data indicate that buffered EDTA and SVTLE can be used to dissolve calcium oxalate stones and, at the concentrations used here, do not damage tissue.

Modeling the Nucleation and Growth of Calcium Carbonate Crystals in Cooling Crystallizer Unit

This paper covers the nucleation and growth of calcium carbonate crystals in a cooling crystallizer unit. In nucleation process, small particles are generated in a new phase inside the homogenous supersaturated phase, this situation occurs by cooling or warming. The system was modeled to assess the effect of temperature and mixer rotation on the size of produced crystals. Experiments were performed at 9 different temperatures and 5 different mixer rotations and the effect of these two parameters on the size of carbonate calcium crystals were studied. The results demonstrated that temperature decrease would lead to increase of the crystals diameter. Moreover, the increase in mixer rotation causes decrease in the crystals diameter. Based on these findings, the procedure of cooling the crystallizer was modeled in the meta-stable Region. Furthermore, optimum rotation of the mixer and the optimum measure of the seeds were calculated for the discrete cooling crystallizer. Finally, the model that defines the relation between the mixer rotation and the crystals’ diameter, the number of crystals and diameter of particles, as well as the number of particles and mixer rotation was determined to be developed.

Association between low cortical bone mineral density, soft-tissue calcification, vascular calcification and chondrocalcinosis: a case-control study

ObjectivesTo examine the association between bone mineral density (BMD), soft-tissue calcification, vascular calcification and chondrocalcinosis (CC).MethodsA case-control study within the Genetics of Osteoarthritis and Lifestyle (GOAL) database (n=3170). All GOAL...

Quiz Page August 2013: Kidney Failure in a Transplant From an Identical Twin

Quiz Page August 2013: Kidney Failure in a Transplant From an Identical Twin

AB0999 Detection of calcium crystals in knee osteoarthritis synovial fluid. A comparison between ordinary light and scanning electronic microscopy

BackgroundOsteoarthritis (OA) is the most common joint disease and the knee is frequently involved. OA is characterized by a progressive loss of articular cartilage, osteophyte formation, thickening of the subchondral...

Ulva flexuosa subsp. pilifera (Chlorophyta, Ulvophyceae) from the Wielkopolska region (West Poland): a new observation on the ultrastructure of vegetative cells

Abstract Ulva flexuosa subsp. pilifera previously known from northern Poland, from the channel near Szczecin and ponds near Łódź, has recently been found in the Malta Reservoir in the Wielkopolska (West Poland) region. Specimens collected in the Wielkopolska region were examined in detail, also under a transmission electron microscope (TEM). The morphometric analysis of Ulva thalli (both young and mature specimens) was performed in order to study the differences in the ultrastructure of vegetative cells. Rectangular cells in young thalli measured from 32.21–55.81 μm to 20.24–35.12 μm, and they formed clear longitudinal rows, while cells in the mature specimens ranged from 25.09–47.66 μm to 18.90–31.56 μm. This study indicates that vegetative cells of the mature thalli show tendency towards distortions of both the longitudinal and transverse cells arrangement. This distortion is determined by the development of possible carbonate calcium crystals on the thalli surface. The ultrastructural analysis (TEM) confirmed that the structure and placement of thylakoids is genus/species specific.

Angiophagy

Cell viability in the brain is critically dependent on uninterrupted blood flow through the microvasculature. However, microvessels, because of their small diameter and low-flow velocity, are prone to occlusion by blood clots, fragments of atheromatous plaques, or other circulating debris. Furthermore, microvascular occlusions can occur after large-vessel thrombosis because of blood stasis or due to clot fragmentation and distal microembolization. Microvascular occlusion could thus be a frequent phenomenon throughout life that may play important roles in the pathogenesis of cerebral ischemia and reperfusion, age-related vascular cognitive impairment, and cognitive decline after surgical procedures, such as coronary artery bypass graft. Given the propensity of microvessels to occlusion, it is likely that robust mechanisms have evolved to ensure microvascular patency. The main mechanism that prevents microvascular occlusion is constituted by the combined effects of the fibrinolytic system and hemodynamic pressure, which lead to embolus dissolution and washout. However, the fibrinolytic system is limited to breaking down fibrin-based blood clots and is unable to disrupt other materials, such as calcium and cholesterol crystals and cell debris, which are present in complex thrombi and may occlude microvessels throughout life. Using an experimental mouse model of internal carotid embolization with fluorescently labeled microemboli (10–60 µm), we were able to study the outcome of individual microvascular occlusions in the mouse brain. We tracked individual microvessels using high-resolution confocal imaging in histological brain preparations and by time-lapse 2 photon microscopy of the meninges and superficial cortex in living mice. Using these methods, we recently found a novel mechanism of microvascular recanalization, which we termed “angiophagy.” This mechanism involves the engulfment of emboli by the endothelium followed by their translocation through the vessel wall into the perivascular space. Angiophagy is a robust mechanism that leads to blood flow reestablishment within hours of the embolus extrusion.1 Embolus extravasation …

178 ER STRESS IN VASCULAR CALCIFICATION

Vascular calcification is a pathological process of deposition of calcium and phosphate crystals in the form of hydroxyapatite (HAp) in the blood vessel wall. Its presence in the vessel wall...

Cervical spine instability from tumoral calcinosis

Dear Editor, In the present case, we describe the clinical presentation, surgical management, and long-term follow-up of a rare case of tumoral calcinosis-induced cervical spine instability. Tumoral calcinosis is a rare disease characterized by the deposition of calcium crystals in peri-articular soft tissue. It can be familial or associated with other conditions, typically metabolic or other illnesses that result in metabolic disturbances [5]. It most commonly affects large joints, such as the hip and shoulders, but has been described in smaller joints including the spine.When tumoral calcinosis occurs, it appears as multiloculated, solid calcifications that generally cause local swelling, pain, and restricted mobility of the adjacent joint. Surgical resection of these calcifications is associated with a variable rate of recurrence. In patients with associated metabolic abnormalities, recurrence rates up to 33 % have been reported [2]. Some reports also suggest that incomplete resection is associated with higher rates of recurrence [3, 4, 6]. While tumoral calcinosis occurs around joints, erosion into adjacent bone or joint is a rare complication. Tumoral calcinosis of the cervical spine with bone erosion and instability has only been reported once in the literature [1]. A uremic patient on chronic dialysis presented with C1-2 instability from tumoral calcinosis. We present a rare case of significant cervical spine instability caused by tumoral calcinosis-induced bone erosion in a patient without any metabolic disturbances or personal/family history of calcific disease. To the best of our knowledge, this represents the first case of cervical spine instability caused by tumoral calcinosis without any of the typical associated conditions.

Quality of Opuntia robusta and its use in development of mayonnaise-like product.

The aim of this study was to characterize Opuntia robusta parenchyma and mucilage as foodstuffs. Solute absorption of Opuntia robusta parenchyma was studied, mucilage was used to develop a mayonnaise-like product as substitute emulsifier alternative to egg yolk and oil substitute. Shelf life of mayonnaise-like product was evaluated. Scanning electron microscopy demonstrated that oxalate calcium crystal were present in the Opuntia robusta parenchyma and mucilage with druses morphology; whereas that in the mayonnaise-like product were not observed calcium oxalate crystals, due probably to the product pH is acidic. The heat treatment of parenchyma at 50°C, 65°Brix, had highest influence on the solute absorption (about 35%). Results revealed that best mixture was the formulated with 26.79% oil, 62.50% mucilage and 10.71% whey protein. Mucilage was the component showing the highest effect on all textural characteristics of mayonnaise-like product. The storage temperature had highest effect on the textural characteristics of mayonnaise-like product than storage time. With the right combination of mucilage of Opuntia, oil and whey protein, it is possible to develop a low-fat mayonnaise with functional properties similar to those of commercial mayonnaise.

Workshop report: 4th European crystal network meeting

The 4th workshop of the European Crystal Network took place recently in Paris. Approximately 40 biomedical scientists took part in this closed meeting, whose aim was to facilitate scientific exchange and foster collaborations between centers involved in research in crystal-induced inflammation and crystal deposition diseases. Rheumatology was well represented at the meeting, but there were also participants from other medical disciplines (genetics, epidemiology, nephrology) and basic scientists with diverse interests (crystallography, biochemistry, inflammation, physics). Within the rheumatic diseases, a major focus is gout and hyperuricemia, but the role of calcium crystals in disease is also gaining interest. Recent advances in genetics, inflammation research and the biochemical pathways that mediate calcium crystal deposition were some of the highlights of this year’s meeting. New clinical and therapeutic aspects of hyperuricemia and gout were also discussed. This report summarizes the subjects that were presented with reference to gout and hyperuricemia. Other presentations on the mechanisms of tissue calcification and their possible role in osteoarthritis are not summarized here.

Accumulation of Calcium Crystal in Rat (Rattus norvegicus) Kidney

The aim of the research was to know the effect of tuber Porang powder (Amorphophallus muelleri) to calcium crystal accumulation on kidneys. The research used 2,5-months-old male rats that were orally given tuber Porang powder from three different locations (Klangon, Sumber Baru, and Sumber Bendo) with doses of 6 mg/100 gr BW/day for 3 months. The distribution of calsium crystal accumulation on kidneys was observed under an Olympus microscope CX 31 at 400x magnification. The data were analysed using ANOVA employing SPSS 16 for Windows. The result of the study showed that consumption of tuber Porang powder induced calcium crystal accumulation in rat kidneys. The use of tuber Porang powder from different locations was found to generate different impacts on the accumulation of calcium crystal. The lowest calcium crystal accumulation in kidneys was the ones given Klangon Porang powder, which was equal to 8,19±2,17 spot/ area. In short, the consumption of tuber Porang powder caused the accumulation of crystal calcium in rat kidneys.