B.J.E. de Lange-Brokaar

Synovial inflammation, immune cells and their cytokines in osteoarthritis: a review.

OBJECTIVE Although osteoarthritis (OA) is considered a non-inflammatory condition, it is widely accepted that synovial inflammation is a feature of OA. However, the role of immune cells and their cytokines in OA is largely unknown. This narrative systematic review summarizes the knowledge of inflammatory properties, immune cells and their cytokines in synovial tissues (STs) of OA patients. DESIGN Broad literature search in different databases was performed which resulted in 100 articles. RESULTS Of 100 articles 33 solely investigated inflammation in OA ST with or without comparison with normal samples; the remaining primarily focussed on rheumatoid arthritis (RA) ST. Studies investigating different severity stages or cellular source of cytokines were sparse. OA ST displayed mild/moderate grade inflammation when investigated by means of haematoxylin and eosin (H&E) staining. Most frequently found cells types were macrophages, T cells and mast cells (MCs). Overall the number of cells was lower than in RA, although the number of MCs was as high as or sometimes even higher than in RA ST. Cytokines related to T cell or macrophage function were found in OA ST. Their expression was overall higher than in normal ST, but lower than in RA ST. Their cellular source remains largely unknown in OA ST. CONCLUSION Inflammation is common in OA ST and characterized by immune cell infiltration and cytokine secretion. This inflammation seems quantitatively and qualitatively different from inflammation in RA. Further research is needed to clarify the role of inflammation, immune cells and their cytokines in the pathogenesis of OA.

Inflammatory Cells in Patients with Endstage Knee Osteoarthritis: A Comparison between the Synovium and the Infrapatellar Fat Pad

Objective. To get a better understanding of inflammatory pathways active in the osteoarthritic (OA) joint, we characterized and compared inflammatory cells in the synovium and the infrapatellar fat pad (IFP) of patients with knee OA. Methods. Infiltrating immune cells were characterized by flow cytometry in 76 patients with knee OA (mean age 63.3, 52% women, median body mass index 28.9) from whom synovial tissue (n = 40) and IFP (n = 68) samples were obtained. Pain was assessed by the visual analog scale (VAS; 0–100 mm). Spearman rank correlations and linear regression analyses adjusted for sex and age were performed. Results. Macrophages and T cells, followed by mast cells, were the most predominant immune cells in the synovium and IFP, and were equally abundant in these tissues. Macrophages and T cells secreted mostly proinflammatory cytokines even without additional stimulation, indicating their activated state. Accordingly, most CD4+ T cells had a memory phenotype and contained a significant population of cells expressing activation markers (CD25+, CD69+). Interestingly, the percent of CD69+ T cells was higher in synovial than IFP CD4+ T cells. Preliminary analyses indicated that the number of synovial CD4+ T cells were associated with VAS pain (β 0.51, 95% CI 0.09–1.02, p = 0.02). Conclusion. Our data suggest that the immune cell composition of the synovium and the IFP is similar, and includes activated cells that could contribute to inflammation through secretion of proinflammatory cytokines. Moreover, preliminary analyses indicate that synovial CD4+ T cells might associate with pain in patients with endstage OA of the knee.

Degree of synovitis on MRI by comprehensive whole knee semi-quantitative scoring method correlates with histologic and macroscopic features of synovial tissue inflammation in knee osteoarthritis

OBJECTIVE To evaluate the association between synovitis on contrast enhanced (CE) MRI with microscopic and macroscopic features of synovial tissue inflammation. METHOD Forty-one patients (mean age 60 years, 61% women) with symptomatic radiographic knee OA were studied: twenty underwent arthroscopy (macroscopic features were scored (0-4), synovial biopsies obtained), twenty-one underwent arthroplasty (synovial tissues were collected). After haematoxylin and eosin staining, the lining cell layer, synovial stroma and inflammatory infiltrate of synovial tissues were scored (0-3). T1-weighted CE-MRIs (3 T) were used to semi-quantitatively score synovitis at 11 sites (0-22) according to Guermazi et al. Spearmans rank correlations were calculated. RESULTS The mean (SD) MRI synovitis score was 8.0 (3.7) and the total histology grade was 2.5 (1.6). Median (range) scores of macroscopic features were 2 (1-3) for neovascularization, 1 (0-3) for hyperplasia, 2 (0-4) for villi and 2 (0-3) for fibrin deposits. The MRI synovitis score was significantly correlated with total histology grade [r = 0.6], as well as with lining cell layer [r = 0.4], stroma [r = 0.3] and inflammatory infiltrate [r = 0.5] grades. Moreover, MRI synovitis score was also significantly correlated with macroscopic neovascularization [r = 0.6], hyperplasia [r = 0.6] and villi [r = 0.6], but not with fibrin [r = 0.3]. CONCLUSION Synovitis severity on CE-MRI assessed by a new whole knee scoring system by Guermazi et al. is a valid, non-invasive method to determine synovitis as it is significantly correlated with both macroscopic and microscopic features of synovitis in knee OA patients.

Association of pain in knee osteoarthritis with distinct patterns of synovitis

To determine possible patterns of synovitis on contrast‐enhanced magnetic resonance imaging (CE‐MRI) and its relation to pain and severity in patients with radiographic knee osteoarthritis (OA).

Adipocytes Modulate the Phenotype of Human Macrophages through Secreted Lipids

Previous studies have shown accumulation and an enhanced proinflammatory profile of macrophages in adipose tissue of obese mice, indicating the presence of an interaction between adipocytes and macrophages in this tissue. However, the consequences of this interaction in humans are yet incompletely understood. In this study, we explored the modulating effects of adipocytes on the phenotype of macrophages in humans and studied the possible molecular pathways involved. Adipocyte-conditioned media (ACM) treatment of macrophages for 48 h strongly reduced the LPS-induced IL-12p40 secretion by macrophages, whereas the production of TNF-α and other cytokines remained largely unaffected. This effect was independent of the source of adipocytes. Interestingly, the level of inhibition correlated directly with body mass index (BMI) of the adipocyte donor. Because adipocytes release many different cytokines, adipokines, and lipids, we have separated the protein and lipid fractions of ACM, to obtain insight into the molecular nature of the soluble mediators underlying the observed effect. These experiments revealed that the inhibitory effect resided predominantly in the lipid fraction. Further studies revealed that PGE2 and linoleic and oleic acid were potent inhibitors of IL-12p40 secretion. Interestingly, concentrations of these ACM-derived lipids increased with increase in BMI of the adipocyte donor, suggesting that they could mediate the BMI-dependent effects of ACM. To our knowledge, these results provide first evidence that obesity-related changes in adipose tissue macrophage phenotype could be mediated by adipocyte-derived lipids in humans. Intriguingly, these changes appear to be different from those in murine obesity.

Degree of synovitis on MRI is correlated with histological and macroscopic features of synovial tissue inflammation in knee osteoarthritis

Background and objectives Synovitis is often present on MRI of OA knees and is an important determinant of pain. To better understand the nature of synovitis seen on MRI the authors compared microscopic and macroscopic features of synovial tissue inflammation with synovitis grade on contrast enhanced (CE) MRI. Methods and methods Twenty-two patients (mean age 61±7 years, 73% women, mean BMI 30±5 kg/mm2) with symptomatic radiographic knee OA attending the rheumatology outpatient clinic were included. Arthroscopy of the index knee was performed and macroscopic features (neovascularisation, villi, fibrin and hyperplasia) were scored (0–4). Furthermore, 15–20 synovial biopsies per knee were obtained. After H&E staining, synovial tissue samples were microscopically scored on features: synovial lining layer hyperplasia/enlargement, activation of resident cells/stroma and degree of inflammatory infiltrates. Each feature was scored from 0–3 and a total sum score per patient was devised. Mean total scores (0–9) by three observers were used. Saggital and axial T1-weighted CE MRI images (3T) were used to semi quantitatively score synovitis at 11 different sites according to Guermazi et al, ranging from 0–22.1 Self reported pain was assessed by visual analogue scale (VAS, 0–100). Pearson correlations adjusted for age were used for correlation between total histology synovitis score and total MRI score. Spearman ρ correlations were used for correlation between total histology score and macroscopic features. Both were calculated by SPSS 16.0. Results The mean (SD) synovitis score on MRI was 7.8 (3.9), representing a mild synovitis and mean (SD) histology score was 2.1 (1.5). Median (range) score of macroscopic features (0–4) were 2.0 (1.0–4.0) for neovascularisation, 1.0 (0.0–3.0) for hyperplasia, 2.0 (0.0–4.0) for villi and 2.0 (0.0–3.0) for fibrin. Synovitis score on MRI correlated significantly with microscopic synovitis score (r=0.5, p=0.019] and macroscopic neovascularisation score (r=0.6, p=0.002) and hyperplasia (r=0.4, p=0.40). Furthermore statistically significant correlation between microscopic synovitis score and macroscopic neovascularisation (r=0.5, p=0.012) existed. No significant correlations with VAS pain were seen. Conclusions Synovitis severity on T1 weighted CE MRI images is significantly correlated with both macroscopic and microscopic features of synovitis in patients with knee OA. No association with severity of pain was seen. Therefore, CE MRI evaluation is a reliable, non invasive way to determine synovitis severity in OA patients.

SAT0610 Magnetic Resonance Imaging Outcomes in Patients with Chronic or Recurrent Gonarthritis Treated with Intra-Articular Infliximab or Corticosteroid Injections

Background Intra-articular corticosteroid injection can be used to treat arthritis. It is unknown whether and which signs of joint inflammation on magnetic resonance imaging (MRI) change after such treatment. Objectives To evaluate changes in signs of synovial inflammation by MRI. Methods In the RIA study (Remicade Intra Articularly), a prospective, randomized, double-blind trial to compare the efficacy of intra-articular (ia) injection with infliximab (IFX) or methylprednisolone (MP), 23 patients with active inflammatory recurrent gonarthritis despite ≥1 previous ia corticosteroid injection underwent T1 contrast enhanced MRI of the affected knee before and 4 weeks after the study medication. MRIs were scored using the MRI Osteoarthritis Knee Score (MOAKS), scoring Hoffa synovitis 0-3, and the Knee Osteoarthritis Scoring System (KOSS), scoring joint effusion 0-3. Outcomes of two consecutive MRIs were compared between the randomization groups. Results Two consecutive MRIs were available for 20 knees of 18 patients, which were treated with ia IFX (14 interventions) or with MP (12 interventions). There were no differences in age, number of disease modifying antirheumatic drugs (DMARDs) and number of previous ia corticosteroid injections between the IFX and MP treated patients. Median (IQR) Hoffa synovitis scores and effusion scores were similar prior to IFX injections or MP injections. Four weeks after an injection the Hoffa synovitis scores and effusion scores had decreased significantly in IFX injected knees (from 2.5 (1.8-3.0) to 2.0 (1.0-2.3), p=0.021 and from 2.5 (2.0-3.0) to 1.0 (1.0-3.0), p=0.007, respectively) but not significantly in MP injected knees (from 2.0 (2.0-2.0) to 1.5 (1.0-2.0), p=0.157 and from 3.0 (2.0-3.0) to 2.0 (1.0-3.0), p=0.102, respectively). Clinical evaluation 6 months after the injections showed recurrence of clinical arthritis in all IFX injected knees, and in 50% of MP injected knees. Median (IQR) Hoffa synovitis scores and effusion scores before injection were similar in MP injected patients who did or did not had recurrence of clinical arthritis 2.0 (2.0-3.0) and 2.0 (1.0-2.0) p=0.080 in patients with recurrence, respectively, and 3.0 (3.0-3.0) and 2.5 (1.8-3.0) p=0.617 in patients with no recurrence, respectively. MP injected patients who did or did not have a recurrence of clinical arthritis also showed no difference in improvement in Hoffa synovitis scores and effusion scores after injection (delta Hoffa synovitis score 0.0 (-1.5-0.5) p=0.414 and 0 (-1-0) p=0.157, respectively, delta effusion score 0.0 (-1.0-0.0), p=0.157 and 0.0 (-0.5-0.0) p=0.317, respectively. Conclusions In patients with chronic or recurrent gonarthritis treated with ia IFX or MP, there is a statistically significant improvement in MRI Hoffa synovitis and effusion scores after ia IFX injections but not after ia MP injections. This may represent short term efficacy which could be greater for IFX than for MP. However, improvement in MRI scores was not associated with long term clinical efficacy: after 6 months all IFX injected knees and 50% of MP injected knees showed clinical recurrence of arthritis. Disclosure of Interest G. Akdemir: None declared, A. van der Bijl: None declared, B. de Lange-Brokaar: None declared, T. Huizinga: None declared, C. Allaart Grant/research support from: The study was sponsored by Schering-Plough BV and Centocor, Inc., M. Kloppenburg: None declared

AB0041 Number of Mast Cell Are Higher in End Stage Knee Osteoarthritis

Background Mast cells could be important in osteoarthritis (OA), as it is the only type of immune cell whose numbers were shown to be as high or sometimes higher compared to rheumatoid arthritis (RA). To assess their role in OA, we investigated whether the number and activation status of mast cells varies with the disease stage in OA. Objectives To compare number of mast cells and their degranulation status in synovial tissue between different stages of OA. Methods 56 patients with symptomatic knee OA attending the rheumatology or orthopaedic outpatient clinic were included. 22 patients with mild to established knee OA (mean (±SD) age 60 (7) yrs, 73% women, median (range) BMI 29 (24-44) kg/mm2, median (range) 2(1-4) KL grade)) underwent arthroscopy of the knee and synovial biopsies were obtained. 34 patients with end-stage knee OA (mean (±SD) age 63 (9) yrs, 59% women, median (range) BMI 29 (21-50) kg/mm2, median (range) KL grade 3.0 (1-4)) underwent arthroplasty and synovial tissues were collected. After haematoxylin and eosin staining, samples were microscopically scored on following features: synovial lining layer hyperplasia (0-3), activation of resident cells/stroma (0-3) and inflammatory infiltrates (0-3). Mean total scores (0-9) by 3 observers was used. Synovial tissue biopsies were investigated by double immunofluorescence for both CD117 and tryptase. Number of total mast cells CD117+ and degranulated (CD117+tryptase- or CD117+with visible granules outside cell) and non-degranulated mast cells (CD117+tryptase+ with no granules seen outside cell) were counted in 10 sub sequential high-power fields (HPF) at two locations in synovial tissue by 3 blinded observers. Mean number of mast cells by 3 observers per patient per 10 HPF were used for analysis. Mann-Witney U and Kruskal-Wallis tests were used for comparisons. Results A mean (±SD) total histology grade of 2.5 (1.6) was observed for all patients. Mast cells were readily observed and were mainly located in the sublining layer. Median (range) of mast cells per patient per 10 HPF was 21 (1-116) and was significantly higher in the end-stage (26 (1-116)) than in the mild to established (12 (1-48)) knee OA (p-value=0.007)) (Fig. 1a). Furthermore, total number of mast cells significantly increased with increasing KL grade (p-value=0.001). A relatively small proportion of the mast cells was degranulated in both groups (Fig. 1b). A trend towards a higher percentage of degranulated mast cells in mild to established OA compared to end-stage OA was observed, although it did not reach significance (Fig. 1b) Conclusions Our data indicate that there is an accumulation of mast cells in end-stage OA compared to mild/established OA. Furthermore a less activated state of mast cells was observed in end-stage OA. This suggests that mast cells might have different biological functions in different phases of the disease. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.4290

OP0025 Different Patterns of Synovitis Present in OA Patients Associate Differentially with Pain

Background Synovitis is prevalent in knee OA and is an important determinant of pain. Objectives To better understand the nature of synovitis and its association with pain we investigated patterns of synovitis on contrast enhanced (CE) MRI and its relation to pain and radiographic severity. Methods 91 patients (mean (SD) age 62 (7.5) years, 68% woman, BMI median (IQR) 29 (26-31) kg/mm2, median (IQR) Kellgren-Lawrence score 3 (2-4)) with symptomatic knee OA attending the rheumatology or orthopedic outpatient clinic were included. 55 patients underwent arthroscopy and 36 arthroplasty. Sagittal and axial T1-weighted CE MRI images (3T) were used to semi-quantitatively score synovitis at 11 sites (total range 0-22) according to Guermazi et al.1 (Ann Rheum Dis 2011). Self-reported pain was assessed by visual analogue scale (VAS, 0-100), knee injury and osteoarthritis outcome score (KOOS (subscale pain),0-100) and measure of intermittent and constant osteoarthritis pain (ICOAP, intermittent pain (0-100), constant pain (0-100)). Principal component analysis (PCA) with varimax rotation and Keiser Normalization was used to investigate patterns of synovitis for all patients. A factor was said to load significantly on a component when loading exceeded 0.4. Subsequently, different patterns were associated with pain measures in linear regression analysis adjusted for gender and age using SPSS 20.0. Log transformations were used when appropriate. Results A mild synovitis was observed (median (IQR) 7.0 (5-10)). Mean (SD) KOOS pain was 51.8 (23.3). Median (IQR) VAS was 53.0 (32-70) and ICOAP constant pain 35.0 (15.0-55.0) and ICOAP intermittent pain 45.8 (25-60.4). PCA resulted in extraction of 3 components (Eigen value > 1), together explaining 53.7% of variance. Component 1 was characterized by synovitis at 7 sites with mainly medial parapatellar involvement associated with KOOS pain, ICOAP constant pain and radiographic severity, not with VAS and ICOAP intermittent pain. Component 2 was characterized by synovitis at site adjacent to the anterior cruciate ligament, medial parameniscal site, intercondylar site and suprapatellar site, but did not associate with any of the pain measures nor with radiographic severity. Component 3, characterized by synovitis at 3 sites (mainly characterized by synovitis loose body site), was also associated with radiographic severity. Conclusions Different patterns of synovitis in knee OA were observed. Our results suggest that a certain synovitis pattern is associated with pain, providing important insights into mechanisms underlying osteoarthritic knee pain. References Guermazi A, Roemer FW, Hayashi D, Crema MD, Niu J, Zhang Y et al. Assessment of synovitis with contrast-enhanced MRI using a whole-joint semiquantitative scoring system in people with, or at high risk of, knee osteoarthritis: the MOST study. Ann Rheum Dis 2011;70:805-11. Disclosure of Interest None Declared

Adipocytes Modulate the Phenotype of Macrophages Through Secreted Lipids

Background Adipose tissue secretes a wide range of soluble factors that can influence whole body metabolism. Previous studies have shown an accumulation of macrophages and an enhanced pro-inflammatory profile of these cells in adipose tissue of obese mice. Modulation of macrophages by soluble mediators released by adipocytes has been proposed as a possible mechanism underlying these changes. In humans, an increased number of macrophages in adipose tissue of obese individuals have been observed, although no clear change in macrophages phenotype could be established. Moreover, no information exists about the interaction between macrophages and adipocytes in humans. Objective In the present study, we explored the possibility that adipocytes modulate the phenotype of macrophages and studied the possible molecular pathways involved in this modulation. Results Treatment of macrophages with adipocyte-conditioned medium (ACM) resulted in a strong reduction in IL-12p40 secretion upon LPS stimulation, whereas TNFα and other cytokines remained largely unaffected. This effect was independent of the source of ACM. Interestingly, the inhibition increased with increase in Body Mass Index (BMI) of the adipocyte donor. Therefore, it was hypothesised that the effect is mediated by a soluble factor whose release is correlated to the BMI of the adipocyte donor. To this end, we measured several cytokines, adipokines and lipids present in ACM. Among these, the release of several free fatty acids (FA) and PGE2 correlated with the BMI of the adipocyte donor. Further tests indicated that oleic and linoleic acid, as well as PGE2 were able to inhibit IL12p40 secretion, whereas palmitic acid could not. Upon separation of ACM protein and lipid fractions, we confirmed that inhibition of IL12p40 resides mainly in the ACM lipid fraction. Conclusions These results provide first evidence that obesity-related changes in macrophage phenotype could be mediated by adipocytes in humans. These effects are mainly mediated through lipids released by adipocytes. Intriguingly, modulation appears different than in murine obesity, indicating that the immunomodulatory effects of obesity could be different in humans and mice.