Calcium Pyrophosphate Dihydrate Crystals Research Articles

CT imaging features of calcium pyrophosphate dihydrate crystal deposition involving spine

PurposeTo explore the computed tomography (CT) imaging features of the spine involved by calcium pyrophosphate dihydrate crystal deposition disease (CPPD-CDD) in order to improve the understanding of this disease. Materials and methodsAmong 79 patients diagnosed with CPPD-CDD in our hospital, 51 patients with spinal involvement were selected, 9 patients without CT examination were excluded, and 42 patients were finally included as research objects. The CT images of the 42 patients were retrospectively analyzed. The analysis contents mainly included the distribution of lesions and the characteristic imaging findings of intervertebral disc, posterior ligament complex, sacroiliac joint and atlantoaxial joint. ResultsIn this paper, 42 patients with 11 males and 31 females were studied. The onset age ranged from 62 to 95 years. There were 264 lesions in 42 patients with spinal involvement, including 177 lumbar vertebrae, 43 cervical vertebrae, 27 thoracic vertebrae and 17 sacroiliac joints (14 bilateral and 3 unilateral). Characteristic imaging findings include: disc calcification (129 cases), posterior ligament complex calcification (107 cases), sacroiliac articular cartilage calcification (17 cases), and ligaments around the odontoid process calcification (11 cases). ConclusionsThe characteristic images of CPPD-CDD involving the spine are calcification of intervertebral disc, posterior ligament complex, sacroiliac joint surface and ligaments around odontoid process, and the lesions are frequent and coexisting, so it should be differentiated from gout, rheumatoid arthritis, ankylosing spondylitis, ossification of ligamentum flavum, basic calcium phosphate and infectious spondylodiscitis.

Crystal induced arthropathies-a comparative study of 40 patients with apatite rheumatism, chondrocalcinosis and primary synovial chondromatosis.

Introduction: Apatite rheumatism (AR), chondrocalcinosis (Ch-C), and primary synovial chondromatosis (prSynCh) are regarded as distinct clinical entities. The introduction of the non-staining technique by Bély and Apáthy (2013) opened a new era in the microscopic diagnosis of crystal induced diseases, allowing the analysis of MSU (monosodium urate monohydrate) HA (calcium hydroxyapatite), CPPD (calcium pyrophosphate dihydrate) crystals, cholesterol, crystalline liquid lipid droplets, and other crystals in unstained sections of conventionally proceeded (aqueous formaldehyde fixed, paraffin-embedded) tissue samples. The aim of this study was to describe the characteristic histology of crystal deposits in AR, Ch-C, and prSynCh with traditional stains and histochemical reactions comparing with unstained tissue sections according to Bély and Apáthy (2013). Patients and methods: Tissue samples of 4 with apatite rheumatism (Milwaukee syndrome), 16 with chondrocalcinosis, and 20 with clinically diagnosed primary synovial chondromatosis were analyzed. Results and conclusion: Apatite rheumatism, chondrocalcinosis, and primary synovial chondromatosis are related metabolic disorders with HA and CPPD depositions. The authors assume that AR and Ch-C are different stages of the same metabolic disorder, which differ from prSynCh in amorphous mineral production, furthermore in the production of chondroid, osteoid and/or bone. prSynCh is a defective variant of HA and CPPD induced metabolic disorders with reduced mineralization capabilities, where the deficient mineralization is replaced by chondroid and/or bone formation. The non-staining technique of Bély and Apáthy proved to be a much more effective method for the demonstration of crystals in metabolic diseases than conventional stains and histochemical reactions.

Osteostatin Mitigates Gouty Arthritis through the Inhibition of Caspase-1 Activation and Upregulation of Nrf2 Expression.

Gouty arthritis results from monosodium urate (MSU) crystal deposition in joints, initiating (pro)-interleukin (IL)-1β maturation, inflammatory mediator release, and neutrophil infiltration, leading to joint swelling and pain. Parathyroid hormone-related protein (107-111) C-terminal peptide (osteostatin) has shown anti-inflammatory properties in osteoblasts and collagen-induced arthritis in mice, but its impact in gouty arthritis models remains unexplored. We investigated the effect of osteostatin on pyroptosis, inflammation, and oxidation in macrophages, as well as its role in the formation of calcium pyrophosphate dihydrate crystals and MSU-induced gouty arthritis in mice models. Osteostatin ameliorated pyroptosis induced by lipopolysaccharide and adenosine 5'-triphosphate (LPS + ATP) in mice peritoneal macrophages by reducing the expression of caspase-1, lactate dehydrogenase release, and IL-1β and IL-18 secretion. Additionally, IL-6 and tumor necrosis factor-α (TNF-α) were also decreased due to the reduced activation of the NF-κB pathway. Furthermore, osteostatin displayed antioxidant properties in LPS + ATP-stimulated macrophages, resulting in reduced production of mitochondrial and extracellular reactive oxygen species and enhanced Nrf2 translocation to the nuclei. In both models of gouty arthritis, osteostatin administration resulted in reduced pro-inflammatory cytokine production, decreased leukocyte migration, and reduced caspase-1 and NF-κB activation. These results highlight the potential of osteostatin as a therapeutic option for gouty arthritis.

CPPD and Associated Crystals in Clinically Diagnosed Chondrocalcinosis: A Clinicopathological Study of 20 Patients

Introduction: Chondrocalcinosis (Ch-C), induced by calcium pyrophosphate dihydrate crystals (CPPD) [Ca2P2O7.2H2O] is considered as a distinct metabolic disease and a well-defined clinical entity, with characteristic clinical manifestations and symptoms. This study used previously found method by the author .i.e, non-staining approach to discover crystal deposits in patients with clinically diagnosed Ch-C, to find a possible correlation between coexisting crystals, and to assess the role of coexistent crystals in inflammatory cellular processes of joints. Patients and methods: There were forty (40) surgical specimens that had been traditionally processed from 16 patients who had been clinically diagnosed with Ch-C. Results: In unstained tissue sections CPPD crystals were found in 22 of 40 samples of all 16 patients. Beside the CPPD crystals calcium hydroxyapatite (HA) [Ca5(PO4)3(OH)] crystals were found in 26 tissue samples of all 16 patients, cholesterol (CC) [C27H46O] in 23 tissue samples of 12 patients, and crystalline liquid lipid droplets (CL) in 11 tissue samples of 5 patients. More or less amorphous deposits of calcium carbonate [CaCO3] and/or calcium phosphate [Ca3(PO4)2] were present alongside CPPD and the contemporaneous crystals. There was a significant and positive correlation between prevalence of CPPD and HA. The connection between inflammatory cellular infiltration and CPPD, HA, CC or CL crystals surrounded by mineral deposits was not significant. Conclusions: The non-staining approach is a practical, low-tech way to find CPPD, HA, CC, and other crystals (with or without CL). The positive and significant correlation between CPPD and HA support the earlier premise that CPPD and HA represent basically an identical metabolic disorder). CPPD and/or HA crystals can provoke inflammatory processes responsible for the clinical symptoms, but the variable amounts of amorphous minerals enclose (isolate) the crystals, and can reduce or eliminate the inflammatory reaction. The authors assume that CC (with or without CL) is an associated phenomenon (concomitant fat metabolic malady) without direct cause of inflammation, and is not responsible for clinical symptoms of crystal induced arthropathies.

Lateral meniscus with tears or with histologic calcification does not increase the risk of lateral osteoarthritis after medial unicompartmental arthroplasty.

Meniscal tears or histological meniscal calcifications (in the absence of radiological chondrocalcinosis) are frequent in osteoarthritis. Whether lateral meniscal lesions influence clinical outcomes after medial unicompartmental knee arthroplasty (UKA) is unknown. We analyzed 130 patients (130 knees) with medial unicompartmental knee arthroplasties between 2005 and 2015. These 130 knees had full articular cartilage thickness in the lateral compartment and no radiological chondrocalcinosis on preoperative radiographs. The lateral meniscus was analyzed with preoperative MRI and a biopsy of the anterior horn at the time of surgery. Synovial fluid was collected and analyzed for calcium pyrophosphate dihydrate crystal deposition (CPPD crystals). Lateral meniscal tears were untreated when detected on MRI or during surgery, with the hypothesis that these tears on the opposite compartment would remain asymptomatic in medial UKA. At average 10-year follow-up, patients were evaluated with clinical and radiographic outcome, with a focus on the risk of joint space narrowing of the lateral femorotibial compartment. CPPD crystals were present in the synovial fluid of 70 knees. Lateral meniscal tears were seen on MRI in 34 (49%) normal meniscuses of the 60 knees without CPPD crystals and in six other knees without histological meniscal calcification despite CPPD crystals. Histological calcification was present on 61 lateral meniscuses with 53 meniscal tears. The results showed no significant differences in the clinical outcomes between knees with lateral meniscal tears or lateral meniscal histological chondrocalcinosis or both lesions and those without these conditions. Additionally, radiographic progression of osteoarthritis in the opposite femorotibial compartment of the knee was not more frequent in patients with these meniscal issues. The tenyear cumulative survival rates, measured by the need for total knee arthroplasty, were 91% for knees without meniscal lesions and 92% for knees with these lesions. On this basis, treatment of meniscal tears of the lateral compartment and routine aspiration of the knee to assess for birefringent crystals in the planning of medial UKA do not appear necessary.

E062 Calcium pyrophosphate dihydrate crystal in operated rheumatoid arthritis of the knee

Abstract Background/Aims Calcium pyrophosphate dihydrate (CPPD) crystals are also one of the common types of crystals causing crystal-induced inflammatory arthritis. Several studies have elucidated important aspects of the molecular and cellular mechanisms of CPPD crystal formation. CPPD crystal deposition disease is a common arthritic disorder in mainly elderly patients. CPPD crystals are found in multiple joints (e.g. knee, wrist joints) in association with severe cartilage degeneration in osteoarthritis. However, there are few reports concerning about CPPD crystals in patients with rheumatoid arthritis (RA). In this study, I investigated the relationship between CPPD crystal and operated RA of the knee. Methods Seventy-seven total knee arthroplasties were performed for RA knees from January 2016 to September 2021. I defined the lower extremity alignment (LEA) base on the femorotibial angle (FTA) as valgus (FTA<170°), neutral (170°≤FTA<180°), and varus (FTA≥180°). I also defined the osteophyte formation degree (OFD) in the knee (grade 0-3). At the operation, joint fluids were collected from 70 knees (average age 71.6; male 8; female 72 body mass index (BMI) 24.4kg/m2,; C-reactive protein (CRP) 1.20mg/dL; erythrocyte sedimentation rate (ESR) 49.5mm/h; matrix metalloprotease (MMP)-3 289.5ng/dL; OFD (0/1/2/3) 0/38/25/7; FTA 177.9°; LEA: varus 35; neutral 22; valgus 13). We evaluated the relationship between CPPD crystals and age, gender, FTA, BMI, CRP, ESR, MMP-3, OFD and LEA. When I analyzed the risk factors using the logistic regression method, LEA were defined as valgus (FTA<175°) and varus (FTA≥176°). Results CPPD crystals were detected from 19 RA knees (27.1%). There were no significant differences between CPPD(+) and (-) groups about age, gender, FTA, BMI, CRP, ESR and MMP-3. The more severe osteophyte formation showed higher CPPD(+) rate tendency; however, there was no significance. CPPD(+) rates were 34.2% in varus knees, 23.1% in valgus knees and 18.2% in neutral knees, respectively. We reported that age, valgus knee, severe osteophyte and low BMI were the risk factors of CPPD(+) in operated osteoarthritis (OA) knees, previously. In this study, there was no difference between CPPD(+) and (-) groups about these factors. Especially, CPPD(+) rate in RA valgus knees was one third of that of OA valgus knees. Conclusion It may be suggested that CPPD production mechanism in RA knees is different from that in OA knees. Disclosure M. Nishikawa: None.

Management of temporomandibular joint diseases: a rare case report of coexisting calcium pyrophosphate crystal deposition and synovial chondromatosis

BackgroundThe coexistence of calcium pyrophosphate dihydrate crystal deposition (CPP) and synovial chondromatosis (SC) in the temporomandibular joint (TMJ) is rarely reported. CPP disease (CPPD) is complex arthritis synonymous with excessive pyrophosphate production and variable aberrations in mineral and organic phase metabolism of the joint cartilage, leading to local inundated CPP and crystal deposition of partially deciphered predispositions. Meanwhile, SC is a rare benign synovial joint proliferative disease of unclear etiology and has a low risk of malignant transformation. However, SC manifests severe joint disability and dysfunction because of connective tissue metaplasia of the synovial membrane, which forms cartilaginous nodules with or without calcifications or ossifications. These nodules often detach and form intra-articular loose bodies and very rarely within extraarticular spaces.Case presentationWe report the case of a 61-year-old man to expand the body of literature on these unusual coexisting arthropathies of the TMJ. The patient presented to our hospital in 2020 with complaints of pain in the right TMJ and trismus for over 6 months. Radiographic assessments of the TMJ provided a preoperative provisional diagnosis of SC. However, the histopathology of the open biopsy revealed tumor-like lesions comprising several deposits of rhomboid and rod-shaped crystals that displayed positive birefringence in polarized light, confirming a coexistence of CPPD. A second-stage operation was performed for the complete removal of the loose bodies and chalk-like lesions including synovectomy. No evidence of recurrence was recorded after a follow-up of nearly 1.5 years.ConclusionsIsolated CPPD and SC of the TMJ are prevalent in the literature however, monoarticular coexistence of these diseases is rare, due to the lack of consistency in the diagnostic criteria in clinical practice. Moreover, optimal treatment depends on several considerations. This report delineated the molecular etiopathology and underscored the need for continued deciphering of the causal mechanisms of coexisting CPPD and SC of the TMJ. In addition, the importance of confirmatory testing for accurate diagnosis, and appropriate management of these diseases were discussed.

Cartilage calcification in osteoarthritis: mechanisms and clinical relevance.

Pathological calcification of cartilage is a hallmark of osteoarthritis (OA). Calcification can be observed both at the cartilage surface and in its deeper layers. The formation of calcium-containing crystals, typically basic calcium phosphate(BCP) and calcium pyrophosphate dihydrate (CPP)crystals, is an active, highly regulated and complex biological process that is initiated by chondrocytes and modified by genetic factors, dysregulated mitophagy or apoptosis, inflammation and the activation of specific cellular-signalling pathways. The links between OA and BCP deposition are stronger than those observed between OA and CPP deposition. Here, we review the molecular processes involved in cartilage calcification in OA and summarize the effects of calcium crystals on chondrocytes, synovial fibroblasts, macrophages and bone cells. Finally, we highlight therapeutic pathways leading to decreased joint calcification and potential new drugs that could treat not only OA but also other diseases associated with pathological calcification.

Drugs Recommended in Adult Rheumatic Diseases, But Considered for Off-Label Use in Argentina

BackgroundOff-label (OL) drug use is the prescription of a drug for indications other than those authorised in its technical datasheet. The objective of this study was to identify drugs recommended in rheumatology but considered for off-label use in Argentina. MethodsA list of medications for certain selected rheumatic conditions was compiled. A drug was considered recommended if it was endorsed by a) at least one Argentine or Pan-American treatment guideline or consensus, or b) two international treatment guidelines, or c) one international treatment guideline and one selected textbook. Approval of these drugs for any condition in Argentina until December 31st, 2018 was explored, and medicines were divided into those with on-label indications and those considered for OL use. ResultsOne hundred and thirty-six medications were analysed in 13 clinical conditions. Sixty-seven OL recommendations (49%) were found, and several drugs had more than one. All the conditions included the recommendation of at least 1 OL drug except osteoporosis and rheumatoid arthritis. The frequency of OL recommendations for the following conditions was 100%: calcium pyrophosphate dihydrate crystal deposition disease, polymyalgia rheumatica, Sjögren syndrome, and systemic sclerosis. The drugs with the highest number of OL recommendations were methotrexate (in 7 conditions), and glucocorticoids and mycophenolate (in 4). There were 2 OL recommendations for rituximab and 1 for abatacept. ConclusionsAlmost all the rheumatic disorders analysed involved the recommendation of at least 1 OL medication, and in 4 conditions all the recommendations were OL. Most OL drugs recommended in rheumatology are neither biological nor small-molecule therapies.

Drugs Recommended in Adult Rheumatic Diseases, But Considered for Off-Label Use in Argentina.

Off-label (OL) drug use is the prescription of a drug for indications other than those authorised in its technical datasheet. The objective of this study was to identify drugs recommended in rheumatology but considered for off-label use in Argentina. A list of medications for certain selected rheumatic conditions was compiled. A drug was considered recommended if it was endorsed by a) at least one Argentine or Pan-American treatment guideline or consensus, or b) two international treatment guidelines, or c) one international treatment guideline and one selected textbook. Approval of these drugs for any condition in Argentina until December 31st, 2018 was explored, and medicines were divided into those with on-label indications and those considered for OL use. One hundred and thirty-six medications were analysed in 13 clinical conditions. Sixty-seven OL recommendations (49%) were found, and several drugs had more than one. All the conditions included the recommendation of at least 1 OL drug except osteoporosis and rheumatoid arthritis. The frequency of OL recommendations for the following conditions was 100%: calcium pyrophosphate dihydrate crystal deposition disease, polymyalgia rheumatica, Sjögren syndrome, and systemic sclerosis. The drugs with the highest number of OL recommendations were methotrexate (in 7 conditions), and glucocorticoids and mycophenolate (in 4). There were 2 OL recommendations for rituximab and 1 for abatacept. Almost all the rheumatic disorders analysed involved the recommendation of at least 1 OL medication, and in 4 conditions all the recommendations were OL. Most OL drugs recommended in rheumatology are neither biological nor small-molecule therapies.

CHARACTERIZATION OF THE CALCIFICATIONS IN THE POSTERIOR HORN OF HUMAN MENISCUS VIA RAMAN MICROSPECTROSCOPY

Purpose: Calcifications in joint soft tissues, such as meniscus and articular cartilage, are closely associated with osteoarthritis (OA) progression and could act as a potential target for disease-modifying agents in OA therapy. Basic calcium phosphate (BCP) and calcium pyrophosphate dihydrate (CPPD) crystals are the two most common forms of calcium crystals found in these tissues. Moreover, within the CPPD crystals, two different polymorphs - monoclinic and triclinic - have been identified in arthritic patients.

The utility of routine cultures, cell count, and crystal evaluation of aspirate from aseptic olecranon bursitis.

BackgroundAspiration of the olecranon bursa is a treatment option for acute olecranon bursitis (OB). Typically, the aspirate is sent for microbiologic analysis, cell count, and crystal analysis. This study investigates the utility of fluid aspirate analysis from patients with clinically diagnosed aseptic OB.MethodsIn this prospective study (IRB #i20-00986), patients presenting with acute aseptic OB were treated with aspiration as standard of care. Patients consented to participate in this study via phone. Patients with suspected infectious bursitis, open draining wound, and chronic OB were excluded. The aspirate was sent out for routine microbiologic analysis (aerobic and anaerobic cultures and Gram staining) and fluid analyses, including cell count with differential and crystal analysis. Nucleated and differential cell count was reported as absolute numbers per cubic millimeter and percentage, respectively. Compression wrap was applied after OB aspiration, and patients were asked to ice and take anti-inflammatory medications. Clinical follow-up was done after 6 weeks and at 3 months for resolution vs. recurrence of symptoms, and the mean time to resolution was reported.ResultsA total of 26 patients (28 cases) with aseptic OB were enrolled in this study. Two patients had bilateral OB. The mean time to aspiration after the onset of symptoms was 26.4 days. One patient had recurrence of swelling after the first aspiration and underwent repeat bursa aspiration. No organisms were isolated or reported on Gram staining on any of the aspirate samples. Two aspirates were reported positive for calcium pyrophosphate dihydrate crystals. No patient had monosodium urate crystals. All patients had resolution of swelling and symptoms without the development of postaspiration infection.ConclusionsThis study demonstrates limited clinical utility of routine microbiologic analysis (cell count, microbiologic, and crystal evaluation) of fluid aspirate from clinically diagnosed aseptic OB. Although 7% of fluid aspirates were positive for calcium pyrophosphate dihydrate crystals, it did not change the overall treatment.

Natural antibodies and CRP drive anaphylatoxin production by urate crystals

In gout, crystallization of uric acid in the form of monosodium urate (MSU) leads to a painful inflammatory response. MSU crystals induce inflammation by activating the complement system and various immune cell types, and by inducing necrotic cell death. We previously found that the soluble pattern recognition molecule C-reactive protein (CRP) recognizes MSU crystals, while enhancing complement activation. In the absence of CRP, MSU crystals still induced complement activation, suggesting additional CRP-independent mechanisms of complement activation. In the present study, we searched for additional MSU crystal-binding complement activators. We found that all healthy individuals, even unborn children, have MSU crystal-specific immunoglobulin M (IgM) in their blood. This indicates that innate IgM, also known as natural IgM, recognizes these crystals. In serum lacking IgM and CRP, MSU crystals showed negligible complement activation as assessed by the production of the anaphylatoxins C4a, C3a, and C5a (listed in order of production via the classical complement pathway). We show that IgM and CRP both activate the classical complement pathway on MSU crystals. CRP was more efficient at fixating active C1 on the crystals and inducing release of the most inflammatory anaphylatoxin C5a, indicating non-redundant functions of CRP. Notably, while CRP recognizes MSU crystals but not the related calcium pyrophosphate dihydrate (CPPD) crystals, natural IgM bound to both, suggesting common and distinct mechanisms of recognition of individual crystal types by complement activators.

Multilevel calcium pyrophosphate dihydrate deposition in cervical ligamentum flavum: clinical characteristics and imaging features

BackgroundInvolvement in cervical ligamentum flavum is a rare manifestation of the calcium pyrophosphate dihydrate deposition disease. Only few cases of this condition have been reported. We revealed eighteen cases of CPPD in cervical ligamentum flavum that diagnosed at a single medical center. In our case series, clinical characteristics and magnetic resonance imaging findings of patients are described.MethodsWe retrospectively reviewed the medical charts and imaging studies of the eighteen patients with pseudogout attack of the cervical ligamentum flavum. In addition, we discussed the differences between this disease and ossification of ligamentum flavum in image manifestations.ResultsThere were fourteen men and four women aged between 59 and 87 years. Diabetes mellitus and hypertension were the most common comorbidities. Myelopathy and neck pain were presented in most patients. C4–5 and C5–6 were attacked most frequently, and multiple- rather than single-level involvement could be observed in our series. “Acute on chronic phenomenon” was a specific magnetic resonance image finding in patients whose symptom durations were between 2 to 5 months. Compared to ossification of ligamentum flavum, calcium pyrophosphate dihydrate crystal deposition had different image signs, including morphology, side of the involved ligament, no continuity with the lamina, acute on chronic phenomenon, and presence of retro-odontoid mass.ConclusionsNodular calcifications in cervical ligamentum flavum raise highly suspicion for calcium pyrophosphate dihydrate deposition and must be diagnosed by histological examination and polarized light microscopy. This disease is different from ossification of ligamentum flavum, and it could be recognized by specific image features.

A Phage Display-Identified Short Peptide Capable of Hydrolyzing Calcium Pyrophosphate Crystals—The Etiological Factor of Chondrocalcinosis

Chondrocalcinosis is a metabolic disease caused by the presence of calcium pyrophosphate dihydrate crystals in the synovial fluid. The goal of our endeavor was to find out whether short peptides could be used as a dissolving factor for such crystals. In order to identify peptides able to dissolve crystals of calcium pyrophosphate, we screened through a random library of peptides using a phage display. The first screening was designed to select phages able to bind the acidic part of alendronic acid (pyrophosphate analog). The second was a catalytic assay in the presence of crystals. The best-performing peptides were subsequently chemically synthesized and rechecked for catalytic properties. One peptide, named R25, turned out to possess some hydrolytic activity toward crystals. Its catalysis is Mg2+-dependent and also works against soluble species of pyrophosphate.

Accuracy of synovial fluid analysis compared to histology for the identification of calcium pyrophosphate crystals: an ancillary study of the OMERACT US Working Group - CPPD subgroup.

The aim of this study was to evaluate the accuracy of synovial fluid analysis in the identification of calcium pyrophosphate dihydrate crystals compared to microscopic analysis of joint tissues as the reference standard. This is an ancillary study of an international, multicentre cross-sectional study performed by the calcium pyrophosphate deposition disease (CPPD) subgroup of the OMERACT Ultrasound working group. Consecutive patients with knee osteoarthritis (OA) waiting for total knee replacement surgery were enrolled in the study from 2 participating centres in Mexico and Romania. During the surgical procedures, synovial fluid, menisci and hyaline cartilage were collected and analysed within 48 hours from surgery under transmitted light microscopy and compensated polarised light microscopy for the presence/absence of calcium pyrophosphate crystals. All slides were analysed by expert examiners on site, blinded to other findings. A dichotomic score (absence/ presence) was used for scoring both synovial fluid and tissues. Microscopic analysis of knee tissues was considered the gold standard. Sensitivity, specificity, accuracy, positive and negative predictive values of synovial fluid analysis in the identification of calcium pyrophosphate crystals were calculated. 15 patients (53% female, mean age 68 yo ± 8.4) with OA of grade 3 or 4 according to Kellgren-Lawrence scoring were enrolled. 12 patients (80%) were positive for calcium pyrophosphate crystals at the synovial fluid analysis and 14 (93%) at the tissue microscopic analysis. The overall diagnostic accuracy of synovial fluid analysis compared with histology for CPPD was 87%, with a sensitivity of 86% and a specificity of 100%, the positive predictive value was 100% and the negative predictive value was 33%. In conclusion synovial fluid analysis proved to be an accurate test for the identification of calcium pyrophosphate dihydrate crystals in patients with advanced OA.

AB0629 ACCURACY OF SYNOVIAL FLUID ANALYSIS FOR THE IDENTIFICATION OF CALCIUM PYROPHOSPHATE CRYSTALS: AN ANCILLARY STUDY OF OMERACT CRITERION VALIDITY STUDY FOR ULTRASOUND IN CPPD

Background:Synovial fluid analysis (SFA) via compensated polarized light microscopy is still considered the gold standard for the identification and diagnosis of Calcium Pyrophosphate Deposition disease (CPPD)-related arthropathies[1], but very few studies have been published about its diagnostic accuracy.Objectives:The aim of this study was to evaluate the accuracy of SFA in the identification of calcium pyrophosphate dihydrate (CPP) crystals compared to microscopic analysis of joint tissues as the reference standard.Methods:This is an ancillary study of an international, multicentre cross-sectional study performed by the CPPD subgroup of the OMERACT Ultrasound working group[2]. Consecutive patients with knee osteoarthritis (OA) waiting for total knee replacement surgery were enrolled in the study from 2 participating centres, Mexico and Romania. During surgical procedures synovial fluid (SF), menisci and hyaline cartilage were collected and analysed within 48 hours after surgery under transmitted light microscopy and compensated polarised light microscopy for the presence/absence of CPP crystals. All slides were analysed by expert examiners on site, blinded to other findings. A dichotomic score (absence/presence) was used for scoring both SF and tissues. Microscopic analysis of knee tissues was considered the gold standard. Sensitivity, specificity, accuracy, positive and negative predictive values (PPV and NPV) of SFA in the identification of CPP crystals were calculated.Results:15 patients (53% female, mean age 68yo ± 8.4) with OA of grade 3 or 4 according to Kellgren-Lawrence scoring were enrolled. 12 patients (80%) were positive for CPP crystals at SFA and 14 (93%) at tissues microscopic analysis. Among 12 SFA positive patients, all were positive for CPP crystals in either medial or lateral meniscus, and 11 were positive in both; 10 patients were positive at the hyaline cartilage, and all 10 were also positive for at least one meniscus. Regarding the 3 SFA negative patients, only one had no crystals in the examined tissues, while the other 2 patients had CPP crystals in both menisci and hyaline cartilage. The overall diagnostic accuracy of SFA compared to histology analysis for CPPD was 87%, with a sensitivity of 86% and a specificity of 100%, the PPV was 100% and the NPV was 33% (Table 1).Table 1.sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and diagnostic accuracy of synovial fluid analysis compared to the reference standard. CI: Confidential Interval. SF: synovial fluid, in parentheses: numerators and denominators for all percentages provided.SensitivitySpecificityPPVNPVAccuracySF analysis86% (12/14)100% (1/1)100% (12/12)33% (1/3)87% (13/15)(0.65-0.99) CI 95%(0.0-0.25) CI 95%(0.65-0.99) CI 95%(0.0-0.25) CI 95%Conclusion:SFA demonstrated to be an accurate test for the identification of CPP crystals in patients with advanced OA. However, is not always feasible and carries some risks for the patient. Considering the availability of validated imaging techniques for the detection of CPPD, such as US, SFA could be used in those patients where imaging and clinical data are not definitely confirmatory of the disease.

Chondrocytes From Osteoarthritic and Chondrocalcinosis Cartilage Represent Different Phenotypes.

Basic calcium phosphate (BCP)-based calcification of cartilage is a common finding during osteoarthritis (OA) and is directly linked to the severity of the disease and hypertrophic differentiation of chondrocytes. Chondrocalcinosis (CC) is associated with calcium pyrophosphate dihydrate (CPPD) deposition disease in the joint inducing OA-like symptoms. There is only little knowledge about the effect of CPPD crystals on chondrocytes and the signaling pathways involved in their generation. The aim of this study was to investigate the chondrocyte phenotype in CC cartilage and the effect of CPPD crystals on chondrocytes. Cartilage samples of patients with CC, patients with severe OA, and healthy donors were included in this study. The presence of CC was evaluated using standard X-ray pictures, as well as von Kossa staining of cartilage sections. OA severity was evaluated using the Chambers Score on cartilage sections, as well as the radiological Kellgren–Lawrence Score. Patients with radiologically detectable CC presented calcification mainly on the cartilage surface, whereas OA patients showed calcification mainly in the pericellular matrix of hypertrophic chondrocytes. OA cartilage exhibited increased levels of collagen X and matrix metalloproteinase 13 (MMP13) compared with CC and healthy cartilage. This observation was confirmed by qRT-PCR using cartilage samples. No relevant influence of CPPD crystals on hypertrophic marker genes was observed in vitro, whereas BCP crystals significantly induced hypertrophic differentiation of chondrocytes. Interestingly, we observed an increased expression of p16 and p21 in cartilage samples of CC patients compared with OA patients and healthy controls, indicating cellular senescence. To investigate whether CPPD crystals were sufficient to induce senescence, we incubated chondrocytes with BCP and CPPD crystals and quantified senescence using β-gal staining. No significant difference was observed for the staining, but an increase of p16 expression was observed after 10 days of culture. Primary chondrocytes from CC patients produced CPPD crystals in culture. This phenotype was stabilized by mitomycin C-induced senescence. Healthy and OA chondrocytes did not exhibit this phenotype. BCP and CPPD crystals seem to be associated with two different chondrocyte phenotypes. Whereas BCP deposition is associated with chondrocyte hypertrophy, CPPD deposition is associated with cellular senescence.

Screening of Fv Antibodies with Specific Binding Activities to Monosodium Urate and Calcium Pyrophosphate Dihydrate Crystals for the Diagnosis of Gout and Pseudogout.

To date, medical diagnosis of gout and pseudogout has been performed by observing the crystals in the joint fluid of patients under a polarized microscope. Conventional diagnostic methods using a polarized microscope have disadvantages, such as time-consuming analysis, a high false negative rate, and difficulty in distinguishing gout with monosodium urate (MSU) crystals and pseudogout with calcium pyrophosphate dihydrate (CPPD) crystals in synovial fluids. In this study, a chromogenic assay for the diagnosis of gout and pseudogout, without the requirement of a polarized microscope and trained experts, was proposed using Fv antibodies with specific binding activities to MSU and CPPD crystals. The IgG VH chain Fv library with randomized complementarity-determining region 3 (CDR3) region was expressed on the outer membrane of Escherichia coli using autodisplay technology. The target Fv antibodies with binding activity to MSU and CPPD crystals were screened from the autodisplayed Fv library on the E. coli outer membrane, and five clones were selected. On the basis of the binding properties of the screened Fv antibodies, peptides with the selected clone of amino acid sequences of the CDR3 region (15 residues) were chemically synthesized. The binding properties of the synthetic peptides with amino acid sequences of CDR3 regions from the selected clones were analyzed using fluorescence imaging and flow cytometry, and the affinity constants (Kd) of each peptide for binding to MSU and CPPD crystals were calculated by fitting based on the isotherm model. A chromogenic assay configuration for gout and pseudogout was developed using synthetic peptides. In this chromogenic assay, synthetic peptides labeled with biotin and streptavidin-horseradish peroxidase (HRP) complex were used, and crystal detection was possible using a chromogenic reaction between HRP and a chromogenic substrate (TMB). Finally, gout and pseudogout were diagnosed by detecting MSU and CPPD crystals in the synovial fluid in the concentration range of 0-300 μg/mL.

Hemochromatosis Arthropathy in Heterozygous HFE H63D Mutation Without Iron Overload- An Entity Less Commonly Touched

Human homeostatic iron regulator protein gene (HFE gene) H63D mutations even when homozygous are rarely associated with iron overload. These mutations, independent of iron status, are associated with calcium pyrophosphate dihydrate crystal deposition disease (CPPD) leading to arthropathy even for heterozygotes. The arthropathy does not respond to iron depletion. We report a case of a 62-year-old male with chronic generalized arthralgias with no evidence of iron overload or elevated inflammatory markers with characteristic radiographic hook-like osteophytes suggestive of hemochromatosis arthropathy. Further, he was found to be a carrier of HFE H63D mutation. Recognition of the association can help guide goal directed management.