Objective
To investigate the usefulness of point‐of‐care hand and wrist joint ultrasound (US) examination in patients with established rheumatoid arthritis (RA).
Methods
Fifty‐one RA patients were ...evaluated using clinical disease activity measures and gray‐scale and power Doppler (PD) US. Agreement between US and clinical findings and its impact on physicians' confidence and clinical decision were assessed.
Results
Agreement between intraarticular PD signal and joint swelling (JS) was moderate (82%; κ = 0.44). Agreement between PD signal and joint tenderness to palpation (TTP) was fair (75%; κ = 0.24). The greatest agreement between PD signal and clinical findings was seen in the 5th metacarpophalangeal (MCP) joint (96% JS, 88% TTP) and the poorest agreement was seen in the wrist (69% JS, 65% TTP) and 2nd (75% JS, 72% TTP) and 3rd (82% JS, 72% TTP) MCP joints. The presence of PD signal in nonswollen and/or nontender joints accounted for most of the disagreement in the wrists, while the opposite was true for the 2nd/3rd MCP joints. Agreement between sonographic synovial thickening and clinical findings was poor. Total sonographic synovial hypertrophy or PD score correlated significantly with physician‐recorded, but not patient‐recorded, clinical outcomes. US increased both physicians' confidence in their clinical decision (P < 0.0005, irrespective of Clinical Disease Activity Index score) and patients' confidence in physicians' medical decisions (88.4% of the cases). US modified biologic agent and/or disease‐modifying antirheumatic drug (DMARD) use in 7 individual cases, but it did not affect the overall treatment plan (P > 0.15) or DMARD (P < 0.062) or biologic agent (P > 1.0) use in this group of RA patients.
Conclusion
PD examination of the wrist and 2nd/3rd MCP joints might be feasible and clinically meaningful in evaluation of disease activity in patients with established RA. US examination of the hand/wrist joints in RA increases physicians' confidence in their clinical decision and can help to individualize DMARD and biologic agent use.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Rheumatoid arthritis is a prototypical autoimmune disease that causes joint inflammation and destruction
. There is currently no cure for rheumatoid arthritis, and the effectiveness of treatments ...varies across patients, suggesting an undefined pathogenic diversity
. Here, to deconstruct the cell states and pathways that characterize this pathogenic heterogeneity, we profiled the full spectrum of cells in inflamed synovium from patients with rheumatoid arthritis. We used multi-modal single-cell RNA-sequencing and surface protein data coupled with histology of synovial tissue from 79 donors to build single-cell atlas of rheumatoid arthritis synovial tissue that includes more than 314,000 cells. We stratified tissues into six groups, referred to as cell-type abundance phenotypes (CTAPs), each characterized by selectively enriched cell states. These CTAPs demonstrate the diversity of synovial inflammation in rheumatoid arthritis, ranging from samples enriched for T and B cells to those largely lacking lymphocytes. Disease-relevant cell states, cytokines, risk genes, histology and serology metrics are associated with particular CTAPs. CTAPs are dynamic and can predict treatment response, highlighting the clinical utility of classifying rheumatoid arthritis synovial phenotypes. This comprehensive atlas and molecular, tissue-based stratification of rheumatoid arthritis synovial tissue reveal new insights into rheumatoid arthritis pathology and heterogeneity that could inform novel targeted treatments.
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GEOZS, IJS, IMTLJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZAGLJ
Objective
In gout, autoinflammatory responses to urate crystals promote acute arthritis flares, but the pathogeneses of tophi, chronic synovitis, and erosion are less well understood. Defining the ...pathways of epigenomic immunity training can reveal novel pathogenetic factors and biomarkers. The present study was undertaken to seminally probe differential DNA methylation patterns utilizing epigenome‐wide analyses in patients with gout.
Methods
Peripheral blood mononuclear cells (PBMCs) were obtained from a San Diego cohort of patients with gout (n = 16) and individually matched healthy controls (n = 14). PBMC methylome data were processed with ChAMP package in R. ENCODE data and Taiji data analysis software were used to analyze transcription factor (TF)–gene networks. As an independent validation cohort, whole blood DNA samples from New Zealand Māori subjects (n = 13 patients with gout, n = 16 control subjects without gout) were analyzed.
Results
Differentially methylated loci clearly separated gout patients from controls, as determined by hierarchical clustering and principal components analyses. IL23R, which mediates granuloma formation and cell invasion, was identified as one of the multiple differentially methylated gout risk genes. Epigenome‐wide analyses revealed differential methylome pathway enrichment for B and T cell receptor signaling, Th17 cell differentiation and interleukin‐17 signaling, convergent longevity regulation, circadian entrainment, and AMP‐activated protein kinase signaling, which are pathways that impact inflammation via insulin‐like growth factor 1 receptor, phosphatidylinositol 3‐kinase/Akt, NF‐κB, mechanistic target of rapamycin signaling, and autophagy. The gout cohorts overlapped for 37 (52.9%) of the 70 TFs with hypomethylated sequence enrichment and for 30 (78.9%) of the 38 enriched KEGG pathways identified via TFs. Evidence of shared differentially methylated gout TF‐gene networks, including the NF‐κB activation–limiting TFs MEF2C and NFATC2, pointed to osteoclast differentiation as the most strongly weighted differentially methylated pathway that overlapped in both gout cohorts.
Conclusion
These findings of differential DNA methylation of networked signaling, transcriptional, innate and adaptive immunity, and osteoclastogenesis genes and pathways suggest that they could serve as novel therapeutic targets in the management of flares, tophi, chronic synovitis, and bone erosion in patients with gout.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Our aim was to investigate the collagenolytic potential and localization of matrix metalloproteinase-2 (MMP-2) in relation to its regulatory proteins membrane type MT1-MMP and tissue inhibitor of ...metalloproteinases-2 (TIMP-2) in rheumatoid arthritis (RA). For this purpose, we have used purification of MMP-2, MMP-8, MMP-9 and interstitial type I, II, and III collagens; SDS-PAGE/densitometric collagenase activity assay; zymography; Western blotting; reverse transcriptase polymerase chain reaction;
in situ hybridization; and immunofluorescence, ABC, ABC-APAAP double immunostainings. MMP-2 degraded human type II collagen almost as effectively as MMP-8, whereas MMP-9 did not cleave type II collagen. In synovial tissue, MT1-MMP, TIMP-2 and MMP-2 were found in synovial lining in fibroblast- and macrophage-like cells, in stromal cells and in vascular endothelium. MT1-MMP, TIMP-2 and MMP-2 were strongly expressed in the pannocytes of the invasive pannus at the interface, but staining was weak and/or there were few positive cells both “above” and “below” the soft-to-hard tissue (cartilage and/or bone) interface. Rheumatoid synovial tissue extract contained proteolytically active 62/59 kDa MMP-2 and 43 kDa MTI-MMP, but no free TIMP-2. These results indicate that components of the ternary MT1-MMP/TIMP-2/MMP-2 complex are coexpressed in the normal synovial lining and in its pathological extension on the hyaline articular cartilage. MMP-2 may participate in the remodeling of the normal lining and also seems to be localized/focalized to pannocytes at a site critical for tissue destruction in arthritis.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
To assess salivary gland ultrasonography (US) as a diagnostic tool for secondary Sjögren syndrome (sSS) in patients with rheumatoid arthritis (RA).
Salivary gland US images from 30 patients with RA ...were graded using a validated semiquantitative scoring system. Sicca symptoms, oral health, and RA disease activity were assessed.
US changes consistent with SS were found in 40% of patients. Patients with higher US scores had more sicca symptoms as well as higher RA activity and poorer oral health.
Salivary gland US may aid the diagnosis of sSS in patients with RA.
Type IV collagenases/gelatinases (matrix metalloproteinases MMP-2 and MMP-9) in labial salivary glands (LSG) and saliva in Sjögren's syndrome (SS) and healthy controls were studied. Zymograms and ...Western blots disclosed that SS saliva contained 92/82 kD MMP-9/type IV collagenase duplex. Specific activity measurement disclosed 53.1 ± 9.8 U/mg protein MMP-9 in SS compared to 16.5 ± 2.6 U/mg in healthy controls (p = 0.01). MMP-2 did not differ between SS and controls. In SS salivary glands, MMP-2 and MMP-9 were also expressed, in addition to stromal fibroblasts and occasional infiltrating neutrophils, respectively, in acinar end piece cells. In addition, an effective proMMP-9 activator, human trypsin-2 (also known as tumor-associated trypsin-2 or TAT-2), was found in acinar end piece cells and in saliva. Interestingly, proteolytically processed MMP-9 was found in saliva (
vide supra), and
in vivo activated MMP-9 was significantly higher in SS than in controls (p = 0.002). LSGs, particularly in SS, were characterized ultrastructurally by areas containing small cytoplasmic vesicles in the basal parts of the epithelial cells associated with areas of disordered and thickened basal lamina. Based on our results, we conclude here that SS saliva contains increased concentrations of MMP-9, which is of glandular origin in part. Pro MMP-9 is to a large extent proteolytically activated. This is probably mediated by the most potent pro MMP-9 activator found
in vivo thus far, namely trypsin-2. Therefore, the MMP-9/trypsin-2 cascade may be resposible for the increased remodelling and/or structural destruction of the basement membrane scaffolding in salivary glands in SS. Due to the role of basal lamina as an important molecular sieve and extracellular matrix-cell signal, these pathological changes may contribute to the pathogenesis of the syndrome.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Abstract ▪625▪This icon denotes a clinically relevant abstract
There is little objective information about intra-articular bleeding patterns in progressive hemophilic arthropathy. When hemophiliacs ...present with joint pain or suspected tissue bleeding, clinical decisions are routinely based on subjective symptoms and findings including pain, decreased range of motion (ROM), warmth and swelling. These presentations are non-specific and may occur with bleeding or inflammation in hemophilic arthropathy. We hypothesized that routine clinical assessment is unreliable for diagnosis and management of joint pain in adult hemophiliacs, and that introduction of rapid, objective musculoskeletal ultrasound (MSKUS) imaging might differentiate intra-articular bleeds vs joint inflammation, and intra-muscular bleeds vs other regional pain syndromes.
Rapid MSKUS as ‘point of care' testing at our Center in assessment of adult hemophiliacs with symptomatic joint or muscle pain. We used the GE Logiq e BT11 US-module with high frequency 8–13 MHz linear transducer and real time spatial compound imaging capability (resolution 135 um, comparable to T3MRI). Standardized imaging protocols (Querol F et al; Haemophilia 2012; 18:3;215–226) served to obtain gray scale (B-mode) and power Doppler examination. This technique can detect joint effusions as small as physiological amounts of joint fluid.
Fifteen spontaneous, non-traumatic painful episodes in 11 adults with hemophilia were evaluated (6 ankle, 7 knee, 2 muscle). Twelve of the 15 episodes were reported by the patient as bleeding and 3 as arthritis. Of 10 episodes of patient-perceived joint bleeding, only 3 were confirmed by US. One muscle bleed with a 2 inch increase in thigh circumference, indurated swelling and pain was rediagnosed as meralgia paresthetica without bleeding by US evaluation. The other 7 episodes of patient-perceived joint bleeding were due to synovitis (synovial thickening/swelling, synovial and joint fat pad hyperemia on power Doppler, and with or without simple effusion). In contrast, 2 of 3 perceived arthritis flares were reclassified as bleeds. Physician assessment of pain etiology was incorrect in 8 of 15 instances. Swelling, warmth and loss of ROM were present in 3 of 6 confirmed bleeding episodes, and in 6 of 9 confirmed non-bleeding episodes. In 3 of 6 confirmed bleeding episodes, simultaneous plasma factor activities were 5%, 8%, 28%. US findings led to a change of treatment in 14 of 15 episodes. Intensified clotting factor infusions for acute treatment or initiation/intensification of prophylaxis were started in 6 instances. Discontinuation or decrease of clotting factor and/or addition of joint steroid injections, anti-inflammatories or pain management were initiated in 8 instances. Patients reported major improvements in pain, swelling or ROM in 11 of 14 instances when management changed.
We report significant discrepancies between MSKUS findings and patient/physician perceived classification of (sub)acute pain as bleeding, arthritis or intra-muscular bleed. Patients and physicians misclassified pain etiology for the majority of episodes based on symptoms and physical findings. Swelling, warmth and loss of ROM were present in approximately half of either confirmed bleeding or arthritic episodes, demonstrating that these findings are clinically not reliable. MSKUS-confirmed bleeding was present in half of the patients at plasma factor levels usually thought to protect against spontaneous bleeding. MSKUS changed treatment decisions for almost all episodes with prompt symptom relief in most patients. Results from this pilot study indicate that our understanding of symptoms and progression of adult hemophilic arthropathy, including protective clotting factor levels, is rudimentary. The current practice of prescribing clotting factor or conservative measures based on patient and physician perception alone is inadequate and may compromise outcomes for the rapidly growing adult population with hemophilia. Prospective studies evaluating MSKUS for the correct diagnosis of bleeding vs arthritis and more precisely directed therapy in hemophilic arthropathy is urgently needed.
No relevant conflicts of interest to declare.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The hypothesis of the present work was that the pannus tissue overlying the articular hard tissues has an aggressive phenotype and contains the newly discovered collagenase-3 and its endogenous ...inducers and activators. We therefore analyzed the eventual presence of collagenase-3 and its regulation at the pannus-cartilage junction. Collagenase-3 mRNA (
in situ hybridization) and enzyme protein (ABC and immunofluorescence staining) were found in the pannocytes in the pannus-hard tissue junction. Inflammatory round cells associated with the critical interface contained TNF-α and IL-1β. These cytokines induced collagenase-3 secretion in cultured rheumatoid synovial fibroblasts. Procollagenase-3 activators, stromelysin-1, 72 kDa type IV collagenase/gelatinase and membrane-type 1-MMP, were also found in the pannus-hard tissue junction. Active collagenase-3 was inhibited with alendronate (IC
50=500-750 μM). Collagenase-3, due to its substrate profile and local synthesis in a milieu favoring its activation, might play a major role in the degradation of cartilage type II and bone type I collagens. Alendronate, at concentrations attainable in vivo, is able to inhibit collagenase-3. This might offer an option to control collagenase-3-mediated tissue destruction in rheumatoid arthritis.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Objective. To show the eventual presence and extent of production of matrix metalloproteinase 13 (MMP‐13, or collagenase 3) in rheumatoid synovial tissue samples and extracts, and to assess the ...inhibition characteristics of recombinant MMP‐13.
Methods. Immunohistochemical avidin‐biotin‐peroxidase complex staining/morphometry was used to analyze MMP‐13‐positive cells in situ. Neutral salt extraction of synovial tissue, electrophoresis of the extract in different buffer systems, and Western blotting were also used. The inhibitory properties of doxycycline, clodronate, pamidronate, and D‐penicillamine for recombinant enzyme were determined with a soluble type II collagen assay.
Results. MMP‐13 was detected in fibroblast‐ and macrophage‐like mononuclear cells in the synovial lining and stroma and in vascular endothelial cells. The overall expression of MMP‐13 in these cells in the synovial stroma was high in rheumatoid arthritis (86 ± 12%) compared with osteoarthritis (17 ± 5%) patient samples (P = 0.0027). In a high‐pH native electrophoresis gel, immunoreactivity to anti‐MMP‐1 and anti‐MMP‐13 were clearly separated, with anti‐MMP‐13‐immunoreactive material migrating faster than anti‐MMP‐1‐immuno‐reactive material. Finally, in contrast to MMP‐1 and MMP‐8, MMP‐13 was found to be relatively resistant to the inhibitory effects of doxycycline and clodronate in vitro.
Conclusion. Due to its localization in synovial tissue, its substrate profile, increased expression, and relative resistance to known MMP inhibitors, MMP‐13 is suggested to play a major role in the pathogenesis of tissue destruction in rheumatoid arthritis.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Objective
To assess the clinical and histologic effects of an intraarticular application of low‐dose (noncytotoxic) liposomal clodronate in established antigen‐induced monarthritis (AIA) in rabbits.
...Methods
AIA was monitored by assessments of joint swelling, C‐reactive protein levels, and radiographic changes in 17 NZW rabbits for 8 weeks during the course of weekly intraarticular injections of liposomal clodronate (0.145 mg/injection, low dose) or “empty” liposomes. The contralateral knee was injected with liposome buffer alone as the control. End‐point analyses included macroscopic joint examination, immuno‐ and TUNEL staining, Safranin O staining/microspectrophotometry, and tumor necrosis factor α (TNFα) convertase enzyme (TACE) inhibition assay.
Results
Liposomal clodronate–treated rabbits showed a reduction and delay in joint swelling during the first 3 injections. Expression of matrix‐bound (solubilized) TNFα, lining cell hyperplasia, and levels of RAM‐11+ macrophages were low in the synovium of the liposomal clodronate treatment group, but the proportion of apoptotic lining cells was not affected. The radiologic score was low at the end of weeks 2 and 4, but at 8 weeks, no difference, compared with controls, was found in pannus formation or in the extent of joint erosion; also, joint swelling was higher than before initiation of treatment. Injections of liposomal clodronate prevented cartilage proteoglycan loss, which was significant in the superficial zone only. TACE activity was not inhibited by clodronate.
Conclusion
Liposomal clodronate had temporary antiinflammatory and antierosive effects on established AIA in rabbits. Over the long‐term, the loss of cartilage proteoglycans was halted. This observed treatment effect may be related to the inhibition of TNFα production and processing in the synovium.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
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