1,25(OH)2D3 and dexamethasone additively suppress synovial fibroblast activation by CCR6+ T helper memory cells and enhance the effect of tumor necrosis factor alpha blockade

Background

Despite recent improvements in the treatment of rheumatoid arthritis (RA), an insufficient treatment response and the development of treatment resistance in many patients illustrates the need for new therapeutic strategies. Chronic synovial inflammation could be suppressed by targeting RA synovial fibroblast (RASF) activation by, for example, interleukin (IL)-17A-producing CCR6+ T helper memory (memTh) cells. Here, we modulated this interaction by combining the active vitamin D metabolite 1,25(OH)2D3 with dexamethasone (DEX) and explored the potential therapeutic applications.

Methods

CCR6+ memTh cells from peripheral blood mononuclear cells (PBMCs) of healthy donors or treatment-naive early RA patients were cultured alone or with RASF from established RA patients for 3 days and treated with or without 1,25(OH)2D3, DEX, or etanercept. Treatment effects were assessed using enzyme-linked immunosorbent assay (ELISA) and flow cytometry.

Results

1,25(OH)2D3, and to lesser extent DEX, reduced production of the pro-inflammatory cytokines IL-17A, IL-22, and interferon (IFN)ő≥ in CCR6+ memTh cells. Tumor necrosis factor (TNF)őĪ was only inhibited by the combination of 1,25(OH)2D3 and DEX. In contrast, DEX was the strongest inhibitor of IL-6, IL-8, and tissue-destructive enzymes in RASF. As a result, 1,25(OH)2D3 and DEX additively inhibited inflammatory mediators in CCR6+ memTh-RASF cocultures. Interestingly, low doses of mainly DEX, but also 1,25(OH)2D3, combined with etanercept better suppressed synovial inflammation in this coculture model compared with etanercept alone.

Conclusion

This study suggests that 1,25(OH)2D3 and DEX additively inhibit synovial inflammation through targeting predominantly CCR6+ memTh cells and RASF, respectively. Furthermore, low doses of DEX and 1,25(OH)2D3 enhance the effect of TNFőĪ blockade in inhibiting RASF activation, thus providing a basis to improve RA treatment.

Authors

Wendy Dankers, Claudia González-Leal, Nadine Davelaar, Patrick S. Asmawidjaja, Adriana M. C. Mus, Johanna M. W. Hazes, Edgar M. Colin & Erik Lubberts

Link

https://doi.org/10.1186/s13075-018-1706-9

Human Memory Th17 Cell Populations Change Into Anti-inflammatory Cells With Regulatory Capacity Upon Exposure to Active Vitamin D

Background

Chronic synovial inflammation is an important hallmark of inflammatory arthritis, but the cells and mechanisms involved are incompletely understood. Previously, we have shown that CCR6+ memory T-helper (memTh) cells and synovial fibroblasts (SF) activate each other in a pro-inflammatory feedforward loop, which potentially drives persistent synovial inflammation in inflammatory arthritis. However, the CCR6+ memTh cells are a heterogeneous population, containing Th17/Th22 and Th17.1 cells. Currently, it is unclear which of these subpopulations drive SF activation and how they should be targeted. In this study, we examined the individual contribution of these CCR6+ memTh subpopulations to SF activation and examined ways to regulate their function.

Methods

Th17/Th22 (CXCR3‚ąíCCR4+), Th17.1 (CXCR3+CCR4‚ąí), DP (CXCR3+CCR4+), and DN (CXCR3‚ąíCCR4‚ąí) CCR6+ memTh, cells sorted from PBMC of healthy donors or treatment-na√Įve early rheumatoid arthritis (RA) patients, were cocultured with SF from RA patients with or without anti-IL17A, anti-IFNő≥, or 1,25(OH)2D3. Cultures were analyzed by RT-PCR, ELISA, or flow cytometry.

Results

Th17/Th22, Th17.1, DP, and DN cells equally express RORC but differ in production of TBX21 and cytokines like IL-17A and IFNő≥. Despite these differences, all the individual CCR6+ memTh subpopulations, both from healthy individuals and RA patients, were more potent in activating SF than the classical Th1 cells. SF activation was partially inhibited by blocking IL-17A, but not by inhibiting IFNő≥ or TBX21. However, active vitamin D inhibited the pathogenicity of all subpopulations leading to suppression of SF activation.

Conclusions

Human CCR6+ memTh cells contain several subpopulations that equally express RORC but differ in TBX21, IFNő≥, and IL-17A expression. All individual Th17 subpopulations are more potent in activating SF than classical Th1 cells in an IFNő≥-independent manner. Furthermore, our data suggest that IL-17A is not dominant in this T cell-SF activation loop but that a multiple T cell cytokine inhibitor, such as 1,25(OH)2D3, is able to suppress CCR6+ memTh subpopulation-driven SF activation.

Authors

Wendy Dankers, Nadine Davelaar, Jan Piet van Hamburg, Jeroen van de Peppel, Edgar M Colin, Erik Lubberts

Link

https://doi.org/10.3389/fimmu.2019.01504

The heterogeneous human memory CCR6+ T helper-17 populations differ in T-bet and cytokine expression but all activate synovial fibroblasts in an IFNő≥-independent manner

Background

Chronic synovial inflammation is an important hallmark of inflammatory arthritis, but the cells and mechanisms involved are incompletely understood. Previously, we have shown that CCR6+ memory T-helper (memTh) cells and synovial fibroblasts (SF) activate each other in a pro-inflammatory feedforward loop, which potentially drives persistent synovial inflammation in inflammatory arthritis. However, the CCR6+ memTh cells are a heterogeneous population, containing Th17/Th22 and Th17.1 cells. Currently, it is unclear which of these subpopulations drive SF activation and how they should be targeted. In this study, we examined the individual contribution of these CCR6+ memTh subpopulations to SF activation and examined ways to regulate their function.

Methods

Th17/Th22 (CXCR3‚ąíCCR4+), Th17.1 (CXCR3+CCR4‚ąí), DP (CXCR3+CCR4+), and DN (CXCR3‚ąíCCR4‚ąí) CCR6+ memTh, cells sorted from PBMC of healthy donors or treatment-na√Įve early rheumatoid arthritis (RA) patients, were cocultured with SF from RA patients with or without anti-IL17A, anti-IFNő≥, or 1,25(OH)2D3. Cultures were analyzed by RT-PCR, ELISA, or flow cytometry.

Results

Th17/Th22, Th17.1, DP, and DN cells equally express RORC but differ in production of TBX21 and cytokines like IL-17A and IFNő≥. Despite these differences, all the individual CCR6+ memTh subpopulations, both from healthy individuals and RA patients, were more potent in activating SF than the classical Th1 cells. SF activation was partially inhibited by blocking IL-17A, but not by inhibiting IFNő≥ or TBX21. However, active vitamin D inhibited the pathogenicity of all subpopulations leading to suppression of SF activation.

Conclusions

Human CCR6+ memTh cells contain several subpopulations that equally express¬†RORC¬†but differ in¬†TBX21, IFNő≥, and IL-17A expression. All individual Th17 subpopulations are more potent in activating SF than classical Th1 cells in an IFNő≥-independent manner. Furthermore, our data suggest that IL-17A is not dominant in this T cell-SF activation loop but that a multiple T cell cytokine inhibitor, such as 1,25(OH)2D3, is able to suppress CCR6+ memTh subpopulation-driven SF activation.

Authors

Wendy Dankers, Hannah den Braanker, Sandra M. J. Paulissen, Jan Piet van Hamburg, Nadine Davelaar, Edgar M. Colin & Erik Lubberts

Link

https://doi.org/10.1186/s13075-021-02532-9

Translational Research Studies Unraveling the Origins of Psoriatic Arthritis: Moving Beyond Skin and Joints

Patients with psoriatic arthritis (PsA) are suffering from a decreased quality of life despite currently available treatments. In the latest years, novel therapies targeting the IL-17/IL-23 and TNF pathways improved clinical outcome. Despite this, remission of disease is not achieved in a considerable group of patients, continuous treatment is very often required to reach clinical remission, and prevention of PsA in patients with psoriasis (PsO) is currently impossible. A better understanding of PsA pathogenesis is required to develop novel treatment strategies that target inflammation and destruction more effectively and at an early stage of the disease, or even before clinically manifest disease. The skin is considered as one of the sites of onset of immune activation, triggering the inflammatory cascade in PsA. PsO develops into PsA in 30% of the PsO patients. Influenced by environmental and genetic factors, the inflammatory process in the skin, entheses, and/or gut may evolve into synovial tissue inflammation, characterized by influx of immune cells. The exact role of the innate and adaptive immune cells in disease pathogenesis is not completely known. The involvement of activated IL-17A+ T cells could implicate early immunomodulatory events generated in lymphoid organs thereby shaping the pathogenic inflammatory response leading to disease. In this perspective article, we provide the reader with an overview of the current literature regarding the immunological changes observed during the earliest stages of PsA. Moreover, we will postulate future areas of translational research aimed at increasing our knowledge on the molecular mechanisms driving disease development, which will aid the identification of novel potential therapeutic targets to limit the progression of PsA.

Keywords: animal models; early psoriatic arthritis; immunopathogenesis; psoriasis; translational.

Copyright © 2021 Bolt, van Ansenwoude, Hammoura, van de Sande and van Baarsen.

Authors

Janne W Bolt, Chaja M J van Ansenwoude, Ihsan Hammoura, Marleen G van de Sande, Lisa G M van Baarsen

Link

https://doi.org/10.3389/fmed.2021.711823

Lymph node stromal cells: subsets and functions in health and disease

Lymph nodes (LNs) aid the interaction between lymphocytes and antigen-presenting cells, resulting in adequate and prolonged adaptive immune responses. LN stromal cells (LNSCs) are crucially involved in steering adaptive immune responses at different levels. Most knowledge on LNSCs has been obtained from mouse studies, and few studies indicate similarities with their human counterparts. Recent advances in single-cell technologies have revealed significant LNSC heterogeneity among different subsets with potential selective functions in immunity. This review provides an overview of current knowledge of LNSCs based on human and murine studies describing the role of these cells in health and disease.

Authors

C Grasso, C Pierie, R E Mebius, L G M van Baarsen

Link

https://doi.org/10.1016/j.it.2021.08.009

Bridging Insights From Lymph Node and Synovium Studies in Early Rheumatoid Arthritis

Objective: To identify molecular changes in synovium before arthritis development in individuals at risk of developing rheumatoid arthritis (RA).

Materials and methods: We included 67 IgM rheumatoid factor and/or anti-citrullinated protein antibody positive individuals with arthralgia but without arthritis. Synovial biopsies were collected after which individuals were prospectively followed for at least 2 years during which 17 developed arthritis. An exploratory genome-wide transcriptional profiling study was performed in 13 preselected individuals to identify transcripts associated with arthritis development (n = 6). Findings were validated using quantitative real-time PCR and immunohistochemistry in the total cohort.

Results: Microarray-based survival analyses identified 5588 transcripts whose expression levels in synovium were significantly associated with arthritis development. Pathway analysis revealed that synovial tissue of at risk individuals who later developed arthritis display higher expression of genes involved in adaptive immune response-related pathways compared to at risk individuals who did not develop arthritis. Lower expression was observed for genes involved in extracellular matrix receptor interaction, Wnt-mediated signal transduction and lipid metabolism. Two-way hierarchical clustering analyses of a 27-gene signature separated the total at risk cohort into two groups, where pre-RA individuals preferred to cluster together. Immunohistochemistry studies revealed more podoplanin positive cells and lower lipid droplet staining in synovial tissue from pre-RA individuals.

Conclusion: Synovial alterations in adaptive immune response and lipid metabolism are associated with future development of arthritis. Since this data show synovial changes without overt cellular infiltration, these may be attributed to preclinical changes in resident synovial tissue cells such as fibroblasts, macrophages and tissue resident T cells.

Authors

Aoife M O’Byrne,¬†Tineke A de Jong,¬†Lisa G M van Baarsen

Link

https://doi.org/10.3389/fmed.2021.820232

Ultrasound-guided lymph node biopsy sampling to study the immunopathogenesis of rheumatoid arthritis: a well-tolerated valuable research tool

Objective: To identify molecular changes in synovium before arthritis development in individuals at risk of developing rheumatoid arthritis (RA).

Materials and methods: We included 67 IgM rheumatoid factor and/or anti-citrullinated protein antibody positive individuals with arthralgia but without arthritis. Synovial biopsies were collected after which individuals were prospectively followed for at least 2 years during which 17 developed arthritis. An exploratory genome-wide transcriptional profiling study was performed in 13 preselected individuals to identify transcripts associated with arthritis development (n = 6). Findings were validated using quantitative real-time PCR and immunohistochemistry in the total cohort.

Results: Microarray-based survival analyses identified 5588 transcripts whose expression levels in synovium were significantly associated with arthritis development. Pathway analysis revealed that synovial tissue of at risk individuals who later developed arthritis display higher expression of genes involved in adaptive immune response-related pathways compared to at risk individuals who did not develop arthritis. Lower expression was observed for genes involved in extracellular matrix receptor interaction, Wnt-mediated signal transduction and lipid metabolism. Two-way hierarchical clustering analyses of a 27-gene signature separated the total at risk cohort into two groups, where pre-RA individuals preferred to cluster together. Immunohistochemistry studies revealed more podoplanin positive cells and lower lipid droplet staining in synovial tissue from pre-RA individuals.

Conclusion: Synovial alterations in adaptive immune response and lipid metabolism are associated with future development of arthritis. Since this data show synovial changes without overt cellular infiltration, these may be attributed to preclinical changes in resident synovial tissue cells such as fibroblasts, macrophages and tissue resident T cells.

Authors

Renée H. Fiechter, Janne W. Bolt, Marleen G. H. van de Sande, Caroline J. Aalbers, Robert B. M. Landewé, Mario Maas, Sander W. Tas & Lisa G. M. van Baarsen

Link

https://doi.org/10.1186/s13075-022-02728-7

Synovial gene signatures associated with the development of rheumatoid arthritis in at risk individuals: A prospective study

Objective: To identify molecular changes in synovium before arthritis development in individuals at risk of developing rheumatoid arthritis (RA).

Materials and methods: We included 67 IgM rheumatoid factor and/or anti-citrullinated protein antibody positive individuals with arthralgia but without arthritis. Synovial biopsies were collected after which individuals were prospectively followed for at least 2 years during which 17 developed arthritis. An exploratory genome-wide transcriptional profiling study was performed in 13 preselected individuals to identify transcripts associated with arthritis development (n = 6). Findings were validated using quantitative real-time PCR and immunohistochemistry in the total cohort.

Results: Microarray-based survival analyses identified 5588 transcripts whose expression levels in synovium were significantly associated with arthritis development. Pathway analysis revealed that synovial tissue of at risk individuals who later developed arthritis display higher expression of genes involved in adaptive immune response-related pathways compared to at risk individuals who did not develop arthritis. Lower expression was observed for genes involved in extracellular matrix receptor interaction, Wnt-mediated signal transduction and lipid metabolism. Two-way hierarchical clustering analyses of a 27-gene signature separated the total at risk cohort into two groups, where pre-RA individuals preferred to cluster together. Immunohistochemistry studies revealed more podoplanin positive cells and lower lipid droplet staining in synovial tissue from pre-RA individuals.

Conclusion: Synovial alterations in adaptive immune response and lipid metabolism are associated with future development of arthritis. Since this data show synovial changes without overt cellular infiltration, these may be attributed to preclinical changes in resident synovial tissue cells such as fibroblasts, macrophages and tissue resident T cells.

Authors

Tineke A de Jong, Maria J H de Hair, Marleen G H van de Sande, Johanna F Semmelink, Ivy Y Choi, Danielle M Gerlag, Paul P Tak, Lisa G M van Baarsen

Link

https://doi.org/10.1016/j.jaut.2022.102923

Altered lipid metabolism in synovial fibroblasts of individuals at risk of developing rheumatoid arthritis

Early in vivo embryonic retinal development is a well-documented and evolutionary conserved process. The specification towards eye development is temporally controlled by consecutive activation or inhibition of multiple key signaling pathways, such as the Wnt and hedgehog signaling pathways. Recently, with the use of retinal organoids, researchers aim to manipulate these pathways to achieve better human representative models for retinal development and disease. To achieve this, a plethora of different small molecules and signaling factors have been used at various time points and concentrations in retinal organoid differentiations, with varying success. Additions differ from protocol to protocol, but their usefulness or efficiency has not yet been systematically reviewed. Interestingly, many of these small molecules affect the same and/or multiple pathways, leading to reduced reproducibility and high variability between studies. In this review, we make an inventory of the key signaling pathways involved in early retinogenesis and their effect on the development of the early retina in vitro. Further, we provide a comprehensive overview of the small molecules and signaling factors that are added to retinal organoid differentiation protocols, documenting the molecular and functional effects of these additions. Lastly, we comparatively evaluate several of these factors using our established retinal organoid methodology.

Authors

T A de Jong, J F Semmelink, S W Denis, M G H van de Sande, R H L Houtkooper, L G M van Baarsen

Link

https://doi.org/10.1016/j.jaut.2022.102974