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Expression of Lectin-Like Transcript 1, the Ligand for CD161, in Rheumatoid Arthritis.

Chalan P, Bijzet J, Huitema MG, Kroesen BJ, Brouwer E, Boots AM - PLoS ONE (2015)

Bottom Line: FACS analysis of digested ST confirmed LLT1 expression by CD68+ cells.Elevated systemic sLLT1 was found in all patient groups.Serum levels of sLLT1 were increased in all patient groups (patients with early- and late-stage RA, seropositive arthralgia and spondyloarthropathy) when compared to healthy subjects.

View Article: PubMed Central - PubMed

Affiliation: Department of Rheumatology and Clinical Immunology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands.

ABSTRACT

Objectives: Precursor Th17 lineage cells expressing CD161 are implicated in Rheumatoid Arthritis (RA) pathogenesis. CD4+CD161+ T-cells accumulate in RA joints and may acquire a non classical Th1 phenotype. The endogenous ligand for CD161 is lectin-like transcript 1 (LLT1). CD161/LLT1 ligation may co-stimulate T-cell IFN-γ production. We investigated the presence and identity of LLT1-expressing cells in RA synovial fluid (SF) and synovial tissue (ST). We also assessed levels of soluble LLT1 (sLLT1) in different phases of RA development.

Methods: Paired samples of peripheral blood mononuclear cells (MC) and SFMC (n = 14), digested ST cells (n = 4) and ST paraffin sections (n = 6) from late-stage RA were analyzed for LLT1 expression by flow cytometry and immunohistochemistry. sLLT1 was measured using a sandwich ELISA. Sera and SF from late-stage RA (n = 26), recently diagnosed RA patients (n = 39), seropositive arthralgia patients (SAP, n = 31), spondyloarthropathy patients (SpA, n = 26) and healthy controls (HC, n = 31) were assayed.

Results: In RA SF, LLT1 was expressed by a small proportion of monocytes. In RA ST, LLT1-expressing cells were detected in the lining, sublining layer and in areas with infiltrates. The LLT1 staining pattern overlapped with the CD68 staining pattern. FACS analysis of digested ST confirmed LLT1 expression by CD68+ cells. Elevated systemic sLLT1 was found in all patient groups.

Conclusions: In RA joints, LLT1 is expressed by cells of the monocyte/macrophage lineage. Serum levels of sLLT1 were increased in all patient groups (patients with early- and late-stage RA, seropositive arthralgia and spondyloarthropathy) when compared to healthy subjects.

No MeSH data available.


Related in: MedlinePlus

Surface-expressed LLT1 is found on SF monocytes.Monocytes from A) peripheral blood and B) synovial fluid were gated based on forward and side scatter characteristics. After excluding CD3+ and CD56+ lymphocytes, monocytes were gated based on CD14 and CD16 expression. The frequency of LLT1+ cells was assessed within the total monocyte population. C) The frequency of LLT1+ monocytes and D) LLT1 MFI from paired samples of PB and SF (n = 14). Mouse monoclonal anti-LLT1 antibody, clone 402659 (R&D Systems) was used.
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pone.0132436.g001: Surface-expressed LLT1 is found on SF monocytes.Monocytes from A) peripheral blood and B) synovial fluid were gated based on forward and side scatter characteristics. After excluding CD3+ and CD56+ lymphocytes, monocytes were gated based on CD14 and CD16 expression. The frequency of LLT1+ cells was assessed within the total monocyte population. C) The frequency of LLT1+ monocytes and D) LLT1 MFI from paired samples of PB and SF (n = 14). Mouse monoclonal anti-LLT1 antibody, clone 402659 (R&D Systems) was used.

Mentions: To investigate if LLT1 is upregulated in the pro-inflammatory environment of arthritic joints we first analyzed paired SFMC and PBMC from late-stage RA for LLT1 surface expression (Fig 1A–1D). Flow cytometric analysis detected LLT1 expression on a proportion of SF-derived monocytes whereas LLT1 was hardly detected on PB monocytes (median 5% [range 1.0–16.0%] vs 0.4%, [range 0,2–1,5%] of the CD14+ monocyte population) (Fig 1A, 1B and 1C). LLT1 mean fluorescence intensity (MFI) was also significantly increased within the SF-derived monocytes compared to PB monocytes (Fig 1D). In agreement with previous reports [11,13], we did not detect expression of LLT1 by circulating T-cells, B-cells, NK-cells, monocytes and granulocytes in PB samples of newly diagnosed RA or healthy controls (data not shown).


Expression of Lectin-Like Transcript 1, the Ligand for CD161, in Rheumatoid Arthritis.

Chalan P, Bijzet J, Huitema MG, Kroesen BJ, Brouwer E, Boots AM - PLoS ONE (2015)

Surface-expressed LLT1 is found on SF monocytes.Monocytes from A) peripheral blood and B) synovial fluid were gated based on forward and side scatter characteristics. After excluding CD3+ and CD56+ lymphocytes, monocytes were gated based on CD14 and CD16 expression. The frequency of LLT1+ cells was assessed within the total monocyte population. C) The frequency of LLT1+ monocytes and D) LLT1 MFI from paired samples of PB and SF (n = 14). Mouse monoclonal anti-LLT1 antibody, clone 402659 (R&D Systems) was used.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492745&req=5

pone.0132436.g001: Surface-expressed LLT1 is found on SF monocytes.Monocytes from A) peripheral blood and B) synovial fluid were gated based on forward and side scatter characteristics. After excluding CD3+ and CD56+ lymphocytes, monocytes were gated based on CD14 and CD16 expression. The frequency of LLT1+ cells was assessed within the total monocyte population. C) The frequency of LLT1+ monocytes and D) LLT1 MFI from paired samples of PB and SF (n = 14). Mouse monoclonal anti-LLT1 antibody, clone 402659 (R&D Systems) was used.
Mentions: To investigate if LLT1 is upregulated in the pro-inflammatory environment of arthritic joints we first analyzed paired SFMC and PBMC from late-stage RA for LLT1 surface expression (Fig 1A–1D). Flow cytometric analysis detected LLT1 expression on a proportion of SF-derived monocytes whereas LLT1 was hardly detected on PB monocytes (median 5% [range 1.0–16.0%] vs 0.4%, [range 0,2–1,5%] of the CD14+ monocyte population) (Fig 1A, 1B and 1C). LLT1 mean fluorescence intensity (MFI) was also significantly increased within the SF-derived monocytes compared to PB monocytes (Fig 1D). In agreement with previous reports [11,13], we did not detect expression of LLT1 by circulating T-cells, B-cells, NK-cells, monocytes and granulocytes in PB samples of newly diagnosed RA or healthy controls (data not shown).

Bottom Line: FACS analysis of digested ST confirmed LLT1 expression by CD68+ cells.Elevated systemic sLLT1 was found in all patient groups.Serum levels of sLLT1 were increased in all patient groups (patients with early- and late-stage RA, seropositive arthralgia and spondyloarthropathy) when compared to healthy subjects.

View Article: PubMed Central - PubMed

Affiliation: Department of Rheumatology and Clinical Immunology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands.

ABSTRACT

Objectives: Precursor Th17 lineage cells expressing CD161 are implicated in Rheumatoid Arthritis (RA) pathogenesis. CD4+CD161+ T-cells accumulate in RA joints and may acquire a non classical Th1 phenotype. The endogenous ligand for CD161 is lectin-like transcript 1 (LLT1). CD161/LLT1 ligation may co-stimulate T-cell IFN-γ production. We investigated the presence and identity of LLT1-expressing cells in RA synovial fluid (SF) and synovial tissue (ST). We also assessed levels of soluble LLT1 (sLLT1) in different phases of RA development.

Methods: Paired samples of peripheral blood mononuclear cells (MC) and SFMC (n = 14), digested ST cells (n = 4) and ST paraffin sections (n = 6) from late-stage RA were analyzed for LLT1 expression by flow cytometry and immunohistochemistry. sLLT1 was measured using a sandwich ELISA. Sera and SF from late-stage RA (n = 26), recently diagnosed RA patients (n = 39), seropositive arthralgia patients (SAP, n = 31), spondyloarthropathy patients (SpA, n = 26) and healthy controls (HC, n = 31) were assayed.

Results: In RA SF, LLT1 was expressed by a small proportion of monocytes. In RA ST, LLT1-expressing cells were detected in the lining, sublining layer and in areas with infiltrates. The LLT1 staining pattern overlapped with the CD68 staining pattern. FACS analysis of digested ST confirmed LLT1 expression by CD68+ cells. Elevated systemic sLLT1 was found in all patient groups.

Conclusions: In RA joints, LLT1 is expressed by cells of the monocyte/macrophage lineage. Serum levels of sLLT1 were increased in all patient groups (patients with early- and late-stage RA, seropositive arthralgia and spondyloarthropathy) when compared to healthy subjects.

No MeSH data available.


Related in: MedlinePlus