J.C. Kwekkeboom

The infrapatellar fat pad of patients with osteoarthritis has an inflammatory phenotype

Objectives Obesity is a risk factor for the development of osteoarthritis (OA) in hands and knees. Adipose tissue can secrete different adipokines with powerful immunomodulatory effects. The infrapatellar fat pad (IFP) is an intra-articular organ in the vicinity of the synovium and cartilage. It is hypothesised that IFP-derived soluble factors could contribute to pathological processes in the knee joint. A study was therefore undertaken to compare the release of inflammatory mediators in the IFP and subcutaneous adipose tissue (ScAT) and to characterise the adipocytes and immune cell infiltrate in these tissues. Methods Paired IFP and ScAT samples were obtained from 27 patients with primary OA. The stromal vascular cell fraction (SVF) was isolated and characterised by fluorescence activated cell sorting. Cytokine and adipokine release in fat- and adipocyte-conditioned media was measured by luminex. Results IFP secreted higher levels of inflammatory mediators such as interleukin 6 (IL-6), adipsin, adiponectin and visfatin than ScAT. This could be due to differences in the phenotype of adipocytes and/or in the composition and phenotype of the SVF cells. IFP adipocyte-conditioned media showed a trend towards more IL-6 and adipsin than ScAT. Moreover, the SVF fraction of IFP contained more cells/g tissue, a lower percentage of T cells and a higher percentage of mast cells than ScAT. In addition, T cells had a predominantly pro-inflammatory phenotype while macrophages had a mixed pro- and anti-inflammatory phenotype in the IFP. Conclusion There are profound differences in secreted inflammatory factors and immune cell composition between the IFP and ScAT. These data indicate that IFP-derived soluble mediators could contribute to pathophysiological processes in the OA knee joint.

Are Baseline High Molecular Weight Adiponectin Levels Associated with Radiographic Progression in Rheumatoid Arthritis and Osteoarthritis

Objective. To investigate whether high molecular weight adiponectin (hmwAPN) mediates the associations of total adiponectin (totAPN) with radiographic progression in rheumatoid arthritis (RA) and hand osteoarthritis (HOA). Methods. Associations between baseline hmwAPN or totAPN levels with radiographic progression were determined using multivariate linear regression or generalized estimated equations. Results. In patients with RA, totAPN associated positively, whereas in patients with HOA it associated negatively with radiographic progression. In contrast, hmwAPN did not associate significantly with radiographic progression in either cohort. Conclusion. Our data indicate that the differential effects associated between totAPN and radiographic progression in either RA or HOA are not mediated by hmwAPN.

A1.11 T cells in the infrapatellar fat pad of osteoarthritis patients as a source of IL-6 in the joint

Background and objectives The infrapatellar fat pad (IFP) is an adipose tissue organ present in the knee next to the synovium and cartilage, thereby constituting a potential player in the pathological processes in the osteoarthritic joint. Obesity-associated changes occur in IFP and this supports the hypothesis that IFP could mediate the association between obesity and the development and progression of osteoarthritis (OA). Interestingly, these changes were observed in the stromal vascular fraction (SVF) rather than adipocytes. As this fraction contain many different types of immune cells, we characterised the SVF of the IFP in OA patients phenotypically and functionally. Materials and methods IFP samples were obtained from knee OA patients (N = 43) undergoing joint replacement surgery (58.1% women; mean (SD) age 66.4 years (10.9); mean (SD) BMI 29.2 kg/m2 (5.7)). The SVF was isolated and cells were characterised based on surface markers expression and cytokine production using flow cytometry. Results Characterisation of the SVF of IFP showed the presence of various immune cells in this tissue, whereby macrophages and T cells were most abundant. Interestingly, flow cytometry analyses of ex vivo cytokine production by different cells revealed a subpopulation of CD4+ T cells that were able to produce IL-6 without further stimulation. These IL-6 producing CD4+ T cells expressed CD69, indicating recent activation. Upon polyclonal stimulation, CD4+ T cells were able to secrete IFNγ, TNFα, and IL-4. However, IL-6 producing CD4+ T cells did not secrete these cytokines. Furthermore, chemokine receptor expression revealed that these IL-6 producing T cells could not be categorised as conventional T helper 1 (Th1), Th2, Th17 or Tfh cells. These data indicate that IL-6-secreting T cells are a distinct population of T cells. Finally, we have also studied whether these IL-6 producing T cells are also present in other tissues. Indeed, we have found these cells also in sc adipose tissues and synovium of OA patients, but only at low frequencies in blood. Conclusion In conclusion, we have found a novel population of CD4+ T cells which secrete IL-6 directly ex vivo and are in an activated state, indicating that these CD4+ T cells might recognise adipose tissue antigens and could be involved in the inflammatory processes present in human adipose tissue. Moreover, they are a source of IL-6 in the OA joint, thereby potentially contributing to joint inflammation.

Functional and phenotypical analysis of IL-6-secreting CD4+ T cells in human adipose tissue

Emerging evidence indicates that a dynamic interplay between the immune system and adipocytes contributes to the disturbed homeostasis in adipose tissue of obese subjects. Recently, we observed IL‐6‐secretion by CD4+ T cells from the stromal vascular fraction (SVF) of the infrapatellar fat pad (IFP) of knee osteoarthritis patients directly ex vivo. Here we show that human IL‐6+CD4+ T cells from SVF display a more activated phenotype than the IL‐6− T cells, as evidenced by the expression of the activation marker CD69. Analysis of cytokines secretion, as well as expression of chemokine receptors and transcription factors associated with different Th subsets (Treg, Th1, Th2, Th17 and Tfh) revealed that IL‐6‐secreting CD4+ T cells cannot be assigned to a conventional Th subset. TCRβ gene analysis revealed that IL‐6+ and IL‐6−CD4+ T cells appear clonally unrelated to each other, suggesting a different specificity of these cells. In line with these observations, adipocytes are capable of enhancing IL‐6 production by CD4+ T cells. Thus, IL‐6+CD4+ T cells are TCRαβ T cells expressing an activated phenotype potentially resulting from an interplay with adipocytes that could be involved in the inflammatory processes in the OA joint.

02.40 Lack of obesity-related features in adipocytes and inflammatory cells in the infrapatellar fat pad (ifp) of oa patients

Background Obesity is associated with the development and progression of osteoarthritis (OA). The infrapatellar fat pad (IFP) could contribute to this association due to its localization in the knee joint and secretion of inflammatory mediators. However, little is known about the effects of obesity on the IFP. Therefore, the aim of this study was to investigate the presence of obesity-related features in adipocytes and infiltrating immune cells in the IFP of OA patients. Materials and methods IFP volume was determined by MRI in 79 knee OA patients. IFP and subcutaneous adipose tissue (SCAT) were obtained from 106 knee OA patients (total n=155: 68% women, mean age 65 years, mean (SD) body mass index (BMI) 29.9 kg/m2 (5.7)) undergoing joint replacement surgery. Crown-like structures (CLS) were determined using immunohistochemistry. Adipocyte size was determined by light microscopy and histology. Stromal vascular fraction (SVF) cells were characterised by flow cytometry. Results IFP volume (mean(SD) 23.6 (5.4) mm3) associated with gender and height, but not with BMI. Likewise, volume and size of IFP adipocytes (mean 271 pl, mean 1933 μm) was not correlated with BMI. Few CLS were observed in IFP and the number did not correlate with BMI. Moreover, high BMI was not associated with higher SVF immune cell numbers in IFP, nor with changes in their phenotype. No molecular differences were observed with BMI, besides an increase in TNFα expression. Extensive characterisation of IFP macrophages revealed that CD206 and CD163, usually associated with an anti-inflammatory phenotype were the most abundantly expressed surface markers on macrophages (81% and 41% respectively), while macrophages produced predominantly IL-6 and TNFα, but little IL-10. Interestingly, surface marker and cytokine expression revealed that CD163+ macrophages had an activated and pro-inflammatory phenotype. Conclusions Obesity-related differences usually observed in SCAT and visceral adipose tissue could not be detected in IFP of OA patients, a fat depot implicated in OA pathogenesis.

The infrapatellar fat pad of osteoarthritic patients has an inflammatory phenotype

Background and objectives Obesity is a risk factor for the development of osteoarthritis (OA) in hands and knees. It has become apparent during the last years that adipose tissue can secrete different adipokines with powerful immunomodulatory effects. Because the infrapatellar fat pad (IFP) is an intra-articular organ in the vicinity of synovium and cartilage, the authors hypothesised that IFP-derived soluble factors could contribute to pathological processes in the knee joint. Therefore the authors have extensively compared the release of inflammatory mediators and characterised the adipocytes and immune cell infiltrate in IFP and Sc adipose tissue (ScAT). Materials and methods Paired IFP and ScAT samples were obtained from 27 primary OA patients. The stromal vascular cell fraction (SVF) was isolated and characterised by FACS. Cytokine and adipokine release in fat- and adipocyte-conditioned media was measured by luminex. Results IFP secreted higher levels of inflammatory mediators, like IL-6, adipsin, adiponectin and visfatin than ScAT. This could be due to differences in the phenotype of adipocytes or/and in composition and phenotype of the SVF cells. Indeed, a similar trend was seen for IL-6 and adipsin when adipocyte-conditioned media from IFP and ScAT were compared. Moreover, the SVF fraction of IFP contained more cells per gram tissue, a lower percentage of T cells and a higher percentage of mast cells than ScAT. In addition, IFP-derived T cells displayed a predominantly proinflammatory phenotype, while macrophages in IFP presented a mixed pro- and anti-inflammatory phenotype. Finally, tumour necrosis factor-α release by IFP was correlated with BMI, indicating BMI-related inflammatory changes in IFP. Conclusions The authors show profound differences in secreted inflammatory factors and immune cell composition between IFP and ScAT. These data indicate that IFP is qualitatively different from ScAT and IFP-derived soluble mediators could contribute to pathophysiological processes in the OA knee joint.

Adiponectin is a predictor for radiographic progression in early RA patients, independently of anti-CCP antibodies

Background and objectives Adipose tissue can secrete soluble mediators (adipokines) with potent immune regulatory functions. Some adipokines have been previously associated with radiographic damage in rheumatoid arthritis (RA) in cross-sectional studies. In the present study we investigated the capacity of adipokines to predict radiographic progression over a period of 4 years and studied their contribution relative to other known risk factors, such as anticyclic citrullinated peptide (anti-CCP) antibodies. Materials and methods Serum concentrations of leptin, visfatin, resistin, adiponectin, adipsin, tumour necrosis factor (TNF) α and interleukin (IL) -6 were determined in baseline sera of 253 RA patients from the Early Arthritis Cohort. The association between levels of these adipokines, and radiographic progression was determined using multivariable regression analysis correcting for age, gender, body mass index (BMI) and the presence of antiCCP antibodies. Results IL-6, TNFα, visfatin and adiponectin levels associated positively with radiographic progression over four years. This association was independent of BMI. However, only adiponectin and IL-6 levels remained independent predictors for progression when corrected for anti-CCP antibodies. The association of both TNFα and visfatin with radiographic damage seemed dependent on anti-CCP antibodies, which is in line with the fact that the levels of both cytokines correlated significantly with anti-CCP levels in these patients. Stratification for the presence of anti-CCP antibodies revealed that adiponectin associated with progression only in anti-CCP+ disease, whereas a trend for a positive association was apparent for IL-6 in both anti-CCP+ and anti-CCP− disease. Conclusions Our results indicate that adipokines are predictors for radiographic progression in RA, possibly through distinct underlying biological mechanisms. Within the adipokines studied, adiponectin was found to predict radiographic progression independently of anti-CCP antibodies and only in anti-CCP+ disease, which indicates it as a potential therapeutic target in RA.