OBJECTIVE: To examine the expression of chemokine receptor CCR10 on monocytes/macrophages in the joints of patients with rheumatoid arthritis (RA), and to investigate the role of chemokine CCL28 and its receptor CCR10 in the migration of RA monocytes and its mechanism. METHODS: The expression of CCR10 in synovial tissues from 8 RA patients, 4 osteoarthritis (OA) patients, and 4 normal controls was analyzed by immunohistochemistry, and cell staining was scored on a 0-5 scales. Flow cytometry was used to measure the percentage of CCR10 positive cells in CD14⁺ monocytes from peripheral blood of 26 RA patients and 20 healthy controls, as well as from synovial fluid of 15 RA patients. The chemotactic migration of monocytes from RA patients and healthy controls in response to CCL28 was evaluated using an in vitro Transwell system. Western blotting was conducted to assess phosphorylation of the extracellular signal-regulated kinase (ERK) and protein kinase B (Akt) pathways in RA monocytes upon CCL28 treatment. RESULTS: CCR10 was predominantly expressed in RA synovial lining cells and sublining macrophages, endothelial cells, and lymphocytes. CCR10 expression was significantly increased on lining cells and sublining macrophages in RA synovial tissue compared with OA and normal synovial tissue (both P < 0.01). The patients with RA had markedly elevated expression of CCR10 on peripheral blood CD14⁺ monocytes compared with the healthy controls [(15.6±3.0)% vs. (7.7±3.8)%, P < 0.01]. CCR10 expression on synovial fluid monocytes from the RA patients was (32.0±15.0)%, which was significantly higher than that on RA peripheral blood monocytes (P < 0.01). In vitro, CCL28 caused significant migration of CD14⁺ monocytes from peripheral blood of the RA patients and the healthy controls at concentrations ranging from 10-100 μg/L (all P < 0.01). The presence of neutralizing antibody to CCR10 greatly suppressed CCL28-driven chemotaxis of RA monocytes (P < 0.01). Stimulation of RA monocytes with CCL28 induced a remarkable increase in phosphorylation of ERK and Akt (both P < 0.05). ERK inhibitor (U0126) and phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002) strongly reduced the migration of RA monocytes in response to CCL28 (both P < 0.01). CONCLUSION RA patients had increased CCR10 expression on peripheral blood, synovial fluid, and synovial tissue monocytes/macrophages. CCL28 ligation to CCR10 promoted RA monocyte migration through activation of the ERK and PI3K/Akt signaling pathways. The CCL28-CCR10 pathway could participate in monocyte recruitment into RA joints, thereby contributing to synovial inflammation and bone destruction.
Humans ; Monocytes/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Endothelial Cells/metabolism* ; Phosphatidylinositol 3-Kinases/metabolism* ; Arthritis, Rheumatoid ; Synovial Membrane ; Chemokines, CC/metabolism* ; Synovial Fluid ; Osteoarthritis ; Receptors, CCR10/metabolism*

OBJECTIVETo compare and estimate the diagnostic value and characteristic of different diagnostic methods (blood laboratory test, histological analysis, synovial fluid cytological test and microbiological examination) in detecting the presence of periprosthetic joint infection.

METHODSData of 52 patients underwent hip or knee joint revision in Peking University People's Hospital Arthritis Clinic and Research Center between July 2013 and March 2015 were analyzed retrospectively. For each patient, results of blood laboratory tests(peripheral-blood white blood cell, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), interleukin-6 (IL-6) and high-sensitivity C-reactive protein (Hs-CRP)), histological analysis, synovial fluid white cell count (SWCC), microbiological examinations (synovial fluid, tissue and prosthetic joint sonication fluid) were collected. Data were analyzed by t-test, independent sample median test or χ(2) test, respectively. Sensitivity, specificity, positive predictive value, negative predictive value and accuracy for each method were calculated and compared by receiver operating characteristic curve.

RESULTSThere were 30 female and 22 male patients. Twenty-one patients (40.4%) were diagnosed as PJI. The levels of CRP, ESR, IL-6 and Hs-CRP in patients with PJI were higher than that in aseptic failure patients (Z=23.084, 13.499, 5.796, 17.045, all P<0.05). The sensitivities of CRP, ESR, IL-6 and Hs-CRP were 90.5%, 81.0%, 95.0% and 90.0%. The sensitivities of histological analysis and SWCC were 55.0% and 70.6%, while they had high specificity as 89.7% and 85.7%. The sensitivity of sonication fluid culture was 90.0%, which was higher than that of tissue culture (71.4%) and synovial fluid culture (65.0%) (χ(2) = 5.333, 6.400, all P<0.05).

CONCLUSIONSThe tests of CRP, ESR, IL-6 and Hs-CRP have good value in detecting PJI preoperatively. Histological analysis and SWCC have high specificity, which could help to exclude PJI. Sonication fluid culture has a higher sensitivity than tissue culture and synovial fluid culture.

Arthroplasty, Replacement, Hip ; Arthroplasty, Replacement, Knee ; Blood Sedimentation ; C-Reactive Protein ; metabolism ; Female ; Humans ; Interleukin-6 ; blood ; Knee Joint ; Male ; Prosthesis-Related Infections ; diagnosis ; ROC Curve ; Retrospective Studies ; Sensitivity and Specificity ; Synovial Fluid ; cytology

BACKGROUNDPeriprosthetic joint infection (PJI) is the main cause of failure following total joint arthroplasty. Until now, the diagnosis of PJI is still confronted with technical limitations, and the question of whether synovial fluid biomarker, C-reactive protein (CRP), can provide high value in the diagnosis of PJI remains unanswered and, therefore, was the aim of the study.

METHODSFirst, we conducted a systematic review on CRP in the diagnosis of PJI by searching online databases using keywords such as "periprosthetic joint infection", "synovial fluid", and "C-reactive protein". Eligible studies providing sufficient data to construct 2 × 2 contingency tables were then selected based on the list of criteria and the quality of included studies was assessed subsequently. Finally, the reported sensitivity, specificity, diagnostic odds ratio (DOR), summary receiver operating characteristic (SROC) curve, and the area under the SROC (AUSROC) were pooled together and used to evaluate overall diagnostic performance.

RESULTSSeven studies were included in our review, six of which comprising a total of 456 participants were further investigated in our meta-analysis. The pooled sensitivity, specificity, and DOR were 0.92 (95% confidence interval [CI]: 0.86-0.96), 0.90 (95% CI: 0.87-0.93), and 101.40 (95% CI: 48.07-213.93), respectively. The AUSROC was 0.9663 (standard error, 0.0113).

CONCLUSIONSSynovial fluid CRP is a good biomarker for the diagnosis of PJI with high sensitivity and specificity.

Arthroplasty, Replacement, Hip ; adverse effects ; Arthroplasty, Replacement, Knee ; adverse effects ; Biomarkers ; metabolism ; C-Reactive Protein ; metabolism ; Female ; Humans ; Male ; Prosthesis-Related Infections ; diagnosis ; Synovial Fluid ; metabolism

Chemokine-like factor 1 (CKLF1) is a newly cloned chemotactic cytokine with CCR4 being its functional receptor. Recent evidence demonstrates a role of CKLF1 in arthritis. The aim of this study was to quantify the expression of CKLF1 as well as assess the correlation between CKLF1 and plasma acute-phase markers. Synovium was obtained from 16 osteoarthritis (OA), 15 rheumatoid arthritis (RA) and 10 ankylosing spondylitis (AS) patients undergoing total joint arthroplasty, with other 11 patients treated for meniscal tears during sport accidents serving as normal controls. Levels of CKLF1 and CCR4 mRNA were detected by qRT-PCR, and the expression of CKLF1 was investigated by immunohistochemistry staining, subsequently analyzed with semiquantitative scores. Plasma acute-phase markers of inflammation were determined by ELISA. CKLF1 was found with a particularly up-regulated expression in synovim from AS and RA patients, and CCR4 mRNA levels increased in RA patients, not in OA or AS patients. Elevated levels of plasma markers of inflammation including CRP, ESR and D-dimer were observed in RA. Further, significantly positive correlations between relative expression levels of CKLF1 and CRP/ESR in RA patients and a positive correlation between CKLF1 and ESR in AS patients were found. There was no detectable correlation between CKLF1 and plasma D-dimer. This study confirms an increased but different level of CKLF1 in RA, OA and AS patients, all significantly higher than that in controls. Additionally, the significant positive correlations between CKLF1 levels and CRP/ESR in RA and between CKLF1 and ESR suggest that CKLF1 might contribute to the inflammation state and clinical symptoms in these rheumatic diseases. Further studies are required to investigate the utility of targeting specific CKLF1 for symptom control or disease modification in RA and AS.
Adult ; Arthritis, Rheumatoid ; metabolism ; Biomarkers ; metabolism ; Case-Control Studies ; Chemokines ; genetics ; metabolism ; Female ; Humans ; MARVEL Domain-Containing Proteins ; genetics ; metabolism ; Male ; Middle Aged ; Osteoarthritis ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Receptors, CCR4 ; genetics ; metabolism ; Spondylitis, Ankylosing ; metabolism ; Synovial Fluid ; metabolism

Objective To investigate the expression of miRNA-140 in chondrocytes and synovial fluid of osteoarthritis (OA) patients, and explore the relationship between the miRNA-140 expression and OA severity.Methods This study enrolled 30 OA patients who underwent total knee arthroplasty for chondrocytes sampling and 30 OA patients who underwent intra-articular injection for synovial fluid sampling. All OA patients were grouped into mild [Kellgren and Lawrence (KL) grade 1-2], moderate (KL grade 3) and severe (KL grade 4), with 10 in each subgroups for each sampling purposes. 7 non-OA patients and 10 patients with knee injury were collected for cartilage and synovial fluid sampling respectively as control groups. Chondrocytes were isolated from the cartilage tissue and cultured in vitro. Quantitative real time PCR for miRNA-140 in chondrocytes and synovial fluid were performed, and the U6 snRNA was used as internal control. The expression difference of miRNA-140 among groups and correlation between the expression and the KL grade of OA were analysed using one-way ANOVA and Spearman test respectively. Results The expression of miRNA-140 in chondrocytes of knees in OA patients was reduced than that in normal knees, and the between-group difference was statistically significant (F=305.464, P<0.001). miRNA-140 could be detected in synovial fluid of both normal knees and OA knees, its relative expression level was reduced in synovial fluid of OA group compared with normal group, and the between-group difference was statistically significant as well (F=314.245, P<0.001). The relative expression level of miRNA-140 in both chondrocytes and synovial fluid were negatively correlated with the KL grade of OA(r=-0.969, P<0.001; r=-0.970, P<0.001). Conclusion miRNA-140 could be detected in chondrocytes and synovial fluid of OA patients, and its expression was negatively correlated with the severity of OA.
Adult ; Aged ; Cells, Cultured ; Chondrocytes ; metabolism ; Female ; Humans ; Knee Joint ; metabolism ; Male ; MicroRNAs ; analysis ; Middle Aged ; Osteoarthritis, Knee ; metabolism ; Real-Time Polymerase Chain Reaction ; Synovial Fluid ; metabolism

OBJECTIVETo explore the effect of Bushen Gujin Recipe (BGR) on serum and synovial expression of interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-α) in knee osteoarthritis (KOA) model rabbits.

METHODSTotally 36 8-month-old healthy male New Zealand white rabbits were randomly divided into 4 groups, i.e., the normal control group, the model group, the Western medicine group (Meloxicam, at the daily dose of 6 mg/kg), and the TCM group (BGR, at the daily dose of 53 g/kg), 9 in each group. Modeling was performed in all rabbits except those in the normal control group by using Hulth A method. All medication was performed for 8 consecutive weeks. Contents of IL-1 and TNF-α were detected using ELISA from serum, partial synovial tissue of the front knee joint, cartilage and subchondral bone of the medial femoral condyle.

RESULTSThe joint space became narrowed in the Western medicine group, ranging between the model group and the TCM group. The articular surface was rough with obvious osteophytes. The joint space was slightly narrower in the TCM group; the articular surface was slightly rough with mild osteophytes. Compared with the normal control group, contents of IL-1 and TNF-α in serum and synovial increased in the model group (P < 0.01). Compared with the model group, contents of IL-1 and TNF-α in serum and the synovial fluid decreased in the two treatment groups (P < 0.01). There was no statistical difference in contents of IL-1 and TNF-α between the Western medicine group and the TCM group.

CONCLUSIONBGR promoted the synthesis of cartilage matrix and carti- lage repair through inhibiting the secretion of IL-1 and TNF-α, and prolonging cartilage degeneration.

Animals ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Interleukin-1 ; metabolism ; Knee Joint ; Male ; Osteoarthritis, Knee ; drug therapy ; metabolism ; Rabbits ; Synovial Fluid ; Synovial Membrane ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

BACKGROUND/AIMS: To investigate the rate of detection of monosodium urate (MSU) crystals in the synovial fluid (SF) of patients with acute gouty arthritis and factors associated with false-negative results. METHODS: A total of 179 patients with acute gouty arthritis who had undergone SF crystal examination were identified from the data warehouse of two university hospitals. Clinical and laboratory data were obtained from the medical records. RESULTS: The overall rate of detection of MSU crystals was 78.8%. In univariate analyses, the only significant differences between the variables of crystal-negative and crystal-positive patients were a lower C-reactive protein level (p = 0.040) and fewer patients undergoing emergent surgery in the crystal-positive group (p = 4.5 x 10(-6)). In logistic regression analyses, MSU crystal-negative results were significantly associated with the interval from arthritis onset to crystal examination (p = 0.042), and this was the most significant risk factor for arthroscopic surgery (p = 2.1 x 10(-4)). Seventeen patients who underwent arthroscopic surgery had a significantly longer hospital stay (p = 0.007) and a significant delay in gout treatment (p = 8.74 x 10(-5)). The distribution of crystal-negative patients differed significantly between the SF samples that were evaluated by both the laboratory medicine and the rheumatology departments (p = 1.2 x 10(-14)), and the kappa value was 0.108. CONCLUSIONS: Although several clinical features were associated with detection failure, SF MSU crystal identification was critically dependent on the observer. Considering the impact on the treatment outcomes, implementation of a quality control program is essential.
Acute Disease ; Aged ; Arthritis, Gouty/diagnosis/*metabolism/*surgery ; Arthroscopy ; Biological Markers/metabolism ; Crystallization ; False Negative Reactions ; Female ; Hospitals, University ; Humans ; Length of Stay ; Logistic Models ; Male ; Microscopy, Polarization ; Middle Aged ; Observer Variation ; Predictive Value of Tests ; Reproducibility of Results ; Republic of Korea ; Retrospective Studies ; Synovial Fluid/*metabolism ; Time Factors ; Time-to-Treatment ; Treatment Outcome ; Uric Acid/*metabolism

Effect of interleukin-6 receptor (IL-6R) antibody on polymethyl methacrylate (PMMA) bone cement-mediated expression of osteoprotegerin (OPG) and receptor activator of nuclear factor-kappaB ligand (RANKL) in synovial fibroblasts was investigated. Synovial tissue obtained from total knee arthroplasty was digested and cultured. Inverted microscope was employed to observe the synovial cells and immunocytochemistry (SABC method) staining was used to identify synovial fibroblasts. This experiment was divided into three groups according to different culture media: PMMA group (75 μg/mL PMMA bone cement particles), IL-6R antibody group (10 ng/mL IL-6R antibody+75 μg/mL PMMA bone cement particles), and control group (no IL-6R antibody or PMMA bone cement particles). Influence of IL-6R antibody and PMMA on proliferation of synovial fibroblasts was measured by cell counting kit-8 (CCK-8). ELISA method was used to measure OPG and RANKL levels in culture solution. Fluorescence quantitative real-time PCR (FQ-PCR) was used to detect the expression of OPG and RANKL mRNA. After three consecutive passages, more than 95% of the primary synovial cells became long spindle fibroblast-like cells. SABC staining results showed that the fibroblast-like cells were negative for anti-CD68 antibody and positive for anti-vimentin antibody, with brown madder stained. CCK-8 test demonstrated that the absorbance (A) value at 450 nm was significantly lower in IL-6R antibody group than in PMMA group and control group (P<0.01), but there was no statistically significant difference in A value at 450 nm between the control group and PMMA group (P>0.05). Results of ELISA indicated that the expression of OPG was significantly higher in IL-6R antibody group than in PMMA group and control group (P<0.01). The expression of RANKL was inhibited (P<0.05), and the ratio of OPG/RANKL was significantly increased in IL-6R antibody group as compared with PMMA group and control group. There was no significant difference in the expression of OPG between control group and PMMA group (P>0.05), but the expression of RANKL was higher in PMMA group than in control group (P<0.05), and there was a significant difference in the ratio of OPG/RANKL between them (P<0.05). Results of FQ-PCR revealed the expression of RANKL mRNA was significantly inhibited (P<0.01) and the expression of OPG mRNA was significantly increased (P<0.01) in IL-6R antibody group as compared with PMMA group and control group. The expression of RANKL mRNA was higher in PMMA group than in control group (P<0.05), but the expression of OPG mRNA had no significant difference between them (P>0.05). IL-6R antibody could significantly increase the expression of OPG, but inhibit the expression of RANKL, which might provide a theoretical basis of molecular biology for the prevention and treatment of aseptic loosening of prosthesis.
Antibodies ; administration & dosage ; immunology ; Bone Cements ; Fibroblasts ; immunology ; Gene Expression ; drug effects ; Humans ; Osteoprotegerin ; biosynthesis ; genetics ; Polymethyl Methacrylate ; administration & dosage ; Prostheses and Implants ; RANK Ligand ; biosynthesis ; genetics ; metabolism ; Receptors, Interleukin-6 ; immunology ; metabolism ; Synovial Fluid ; immunology ; metabolism

OBJECTIVETo investigate the role of natural killer-22 (NK-22) cells in the synovial fluid in the proliferation of fibroblast-like synoviocytes (FLS) in patients with rheumatoid arthritis (RA) and explore the possible signal pathway involved.

METHODSNK-22 cells in the SF of RA patients were sorted by flow cytometry. NK-22 cells were cultured for two weeks and the purity was detected by flow cytometry before stimulation with 20 ng/ml phorbol 12-myristate 13-acetate and 0.5 µmol/L ionomycin for 4 h. The level of interleukin-22 (IL-22) in the culture medium supernatant was then measured with ELISA. The proliferation of FLS in the presence of the culture supernatant of NK-22 cells was assessed with MTT assay at 24, 48 and 72 h, and the effect of IL-22 antibody on FLS proliferation was also observed. Real-time PCR and Western blotting were used to detect Stat3 mRNA and p-Stat3 protein levels, respectively, in the FLS exposed to rhIL-22 and AG490.

RESULTSNK-22 cells were successfully sorted by flow cytometry with a purity exceeding 90%. The levels of IL-22 in the supernatant of NKp44(+)NK cell culture averaged 1273.42∓254.48 pg/ml. The FLS proliferated rapidly 24, 48, and 72 h after the addition of culture supernatant of NK-22 cells (P<0.05). IL-22 antibody obviously inhibited the proliferation of FLS induced by NK-22 cell culture supernatant (P<0.05). Exposure of the FLS to rhIL-22 obviously increased cellular Stat3 expression levels, which were significantly lowered by the addition of AG490 (P<0.05).

CONCLUSIONNK-22 cells in the SF of RA patients can produce high concentrations of IL-22 to promote the proliferation of FLS through the STAT3 signal pathway.

Arthritis, Rheumatoid ; metabolism ; Cell Proliferation ; Cells, Cultured ; Fibroblasts ; cytology ; Humans ; Interleukins ; metabolism ; Killer Cells, Natural ; cytology ; metabolism ; STAT3 Transcription Factor ; metabolism ; Signal Transduction ; Synovial Fluid ; cytology ; Synovial Membrane ; cytology

Direct plating of synovial fluid (SF) on agar-based media often fails to identify pathogens in septic arthritis (SA). We developed a PCR assay for the simultaneous detection of Kingella kingae and Staphylococcus aureus from SF to evaluate molecular detection in SF and to estimate the incidence of K. kingae in SA in North America. The assay was based on detection of the cpn60 gene of K. kingae and the spa gene of S. aureus in multiplex real-time PCR. K. kingae was identified in 50% of patients between 0 and 5 yr of age (n=6) but not in any patients >18 yr old (n=105). Direct plating of SF on agar-based media failed to detect K. kingae in all samples. The PCR assay was inferior to the culture-based method for S. aureus, detecting only 50% of culture-positive cases. Our findings suggest that K. kingae is a common pathogen in pediatric SA in North America, in agreement with previous reports from Europe. PCR-based assays for the detection of K. kingae may be considered in children with SA, especially in those with a high degree of clinical suspicion.
Adult ; Arthritis, Infectious/diagnosis/*microbiology ; Bacterial Proteins/genetics ; Child ; Child, Preschool ; DNA, Bacterial/*analysis/metabolism ; Humans ; Infant ; Infant, Newborn ; Kingella kingae/*genetics/isolation & purification ; *Real-Time Polymerase Chain Reaction ; Staphylococcus aureus/*genetics/isolation & purification ; Synovial Fluid/*microbiology