Limits...
Resting CD4+ effector memory T cells are precursors of bystander-activated effectors: a surrogate model of rheumatoid arthritis synovial T-cell function.

Brennan FM, Smith NM, Owen S, Li C, Amjadi P, Green P, Andersson A, Palfreeman AC, Hillyer P, Foey A, Beech JT, Feldmann M - Arthritis Res. Ther. (2008)

Bottom Line: Here, using magnetic beads and fluorescence-activated cell sorting, we extensively phenotype the Tck effector cells and conclude that effector function resides within the CD4+CD45RO+, CCR7-, CD49dhigh population, and that these cells are derived from the effector memory CD4+ T cells in resting blood.Moreover, blockade of these ligands also resulted in inhibition of spontaneous tumour necrosis factor-alpha production in RA synovial mononuclear cell cultures.Taken together, these data strengthen our understanding of T-cell effector function, highlight the multiple involvement of different cell surface ligands in cell-cell contact and, provide novel insights into the pathogenesis of inflammatory RA disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Kennedy Institute of Rheumatology Division, Imperial College Faculty of Medicine, Aspenlea Road, London, W6 8LH, UK. f.brennan@imperial.ac.uk

ABSTRACT

Background: Previously we described a system whereby human peripheral blood T cells stimulated for 8 days in a cytokine cocktail acquired effector function for contact-dependent induction of proinflammatory cytokines from monocytes. We termed these cells cytokine-activated (Tck) cells and found that the signalling pathways elicited in the responding monocytes were identical whether they were placed in contact with Tck cells or with T cells isolated from rheumatoid arthritis (RA) synovial tissue.

Methods: Here, using magnetic beads and fluorescence-activated cell sorting, we extensively phenotype the Tck effector cells and conclude that effector function resides within the CD4+CD45RO+, CCR7-, CD49dhigh population, and that these cells are derived from the effector memory CD4+ T cells in resting blood.

Results: After stimulation in culture, these cells produce a wide range of T-cell cytokines, undergo proliferation and differentiate to acquire an extensively activated phenotype resembling RA synovial T cells. Blocking antibodies against CD69, CD18, or CD49d resulted in a reduction of tumour necrosis factor-alpha production from monocytes stimulated with CD4+CD45RO+ Tck cells in the co-culture assay. Moreover, blockade of these ligands also resulted in inhibition of spontaneous tumour necrosis factor-alpha production in RA synovial mononuclear cell cultures.

Conclusion: Taken together, these data strengthen our understanding of T-cell effector function, highlight the multiple involvement of different cell surface ligands in cell-cell contact and, provide novel insights into the pathogenesis of inflammatory RA disease.

Show MeSH

Related in: MedlinePlus

Proliferation pattern of cytokine-stimulated lymphocytes. (a) Resting peripheral blood lymphocytes (PBLs) undergo cell divisions when they are stimulated with cytokines over 8 days. Freshly elutriated lymphocytes were labeled with 5 μmol/l carboxyfluoroscein succinimidyl ester (CFSE) and stimulated with IL-2, IL-6 and tumour necrosis factor-α for 8 days. The number of cell divisions was analysed on day 8 using CellQuest software. Markers (M1 to M7) indicate sequential cell divisions. (b-e) Proliferation of subpopulations of PBLs stimulated with cytokines. CFSE-labelled PBLs were counterstained with antibodies to CD3 (panel b), CD4 (panel c), CD8 (panel d) and CD56 (panel e). On day 8 the number of cell divisions for each subpopulation of PBLs was acquired and analyzed using CellQuest software. Results for all figures are shown from one donor representative of seven different experiments.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2452984&req=5

Figure 1: Proliferation pattern of cytokine-stimulated lymphocytes. (a) Resting peripheral blood lymphocytes (PBLs) undergo cell divisions when they are stimulated with cytokines over 8 days. Freshly elutriated lymphocytes were labeled with 5 μmol/l carboxyfluoroscein succinimidyl ester (CFSE) and stimulated with IL-2, IL-6 and tumour necrosis factor-α for 8 days. The number of cell divisions was analysed on day 8 using CellQuest software. Markers (M1 to M7) indicate sequential cell divisions. (b-e) Proliferation of subpopulations of PBLs stimulated with cytokines. CFSE-labelled PBLs were counterstained with antibodies to CD3 (panel b), CD4 (panel c), CD8 (panel d) and CD56 (panel e). On day 8 the number of cell divisions for each subpopulation of PBLs was acquired and analyzed using CellQuest software. Results for all figures are shown from one donor representative of seven different experiments.

Mentions: Figure 1a illustrates the CFSE dilution of Tck cells at day 8 counterstained for CD3 (Figure 1b), CD4 (Figure 1c), CD8 (Figure 1d) and CD56 (Figure 1e). At day 8 approximately 40% of cells were undivided (peak M1), whereas 60% of cells (peaks M2 to M7) underwent up to six rounds of cell division (Figure 1a). The CD3+CD4+ and CD3+CD8+ cells were mostly found within the undivided population (M1; Figure 1b,c,d), whereas most CD56+ natural killer (NK) cells (Figure 1e) underwent several rounds of cell division. Pooling data from seven different donors indicated that overall there was a small increase in cell number from 1 million/ml at day 0 to 1.43 million/ml at day 8, accounted for predominantly by increases in CD3-CD56+ NK cells from 0.07 to 0.25 million/ml (about 3.5-fold) and CD3+CD8+ T cells from 0.29 to 0.64 million/ml (2-fold). CD3+CD4+ T cells numbers were increased less, from 0.51 to 0.67 million/ml. CD19+ B cells did not survive the 8-day culture (data not shown).


Resting CD4+ effector memory T cells are precursors of bystander-activated effectors: a surrogate model of rheumatoid arthritis synovial T-cell function.

Brennan FM, Smith NM, Owen S, Li C, Amjadi P, Green P, Andersson A, Palfreeman AC, Hillyer P, Foey A, Beech JT, Feldmann M - Arthritis Res. Ther. (2008)

Proliferation pattern of cytokine-stimulated lymphocytes. (a) Resting peripheral blood lymphocytes (PBLs) undergo cell divisions when they are stimulated with cytokines over 8 days. Freshly elutriated lymphocytes were labeled with 5 μmol/l carboxyfluoroscein succinimidyl ester (CFSE) and stimulated with IL-2, IL-6 and tumour necrosis factor-α for 8 days. The number of cell divisions was analysed on day 8 using CellQuest software. Markers (M1 to M7) indicate sequential cell divisions. (b-e) Proliferation of subpopulations of PBLs stimulated with cytokines. CFSE-labelled PBLs were counterstained with antibodies to CD3 (panel b), CD4 (panel c), CD8 (panel d) and CD56 (panel e). On day 8 the number of cell divisions for each subpopulation of PBLs was acquired and analyzed using CellQuest software. Results for all figures are shown from one donor representative of seven different experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2452984&req=5

Figure 1: Proliferation pattern of cytokine-stimulated lymphocytes. (a) Resting peripheral blood lymphocytes (PBLs) undergo cell divisions when they are stimulated with cytokines over 8 days. Freshly elutriated lymphocytes were labeled with 5 μmol/l carboxyfluoroscein succinimidyl ester (CFSE) and stimulated with IL-2, IL-6 and tumour necrosis factor-α for 8 days. The number of cell divisions was analysed on day 8 using CellQuest software. Markers (M1 to M7) indicate sequential cell divisions. (b-e) Proliferation of subpopulations of PBLs stimulated with cytokines. CFSE-labelled PBLs were counterstained with antibodies to CD3 (panel b), CD4 (panel c), CD8 (panel d) and CD56 (panel e). On day 8 the number of cell divisions for each subpopulation of PBLs was acquired and analyzed using CellQuest software. Results for all figures are shown from one donor representative of seven different experiments.
Mentions: Figure 1a illustrates the CFSE dilution of Tck cells at day 8 counterstained for CD3 (Figure 1b), CD4 (Figure 1c), CD8 (Figure 1d) and CD56 (Figure 1e). At day 8 approximately 40% of cells were undivided (peak M1), whereas 60% of cells (peaks M2 to M7) underwent up to six rounds of cell division (Figure 1a). The CD3+CD4+ and CD3+CD8+ cells were mostly found within the undivided population (M1; Figure 1b,c,d), whereas most CD56+ natural killer (NK) cells (Figure 1e) underwent several rounds of cell division. Pooling data from seven different donors indicated that overall there was a small increase in cell number from 1 million/ml at day 0 to 1.43 million/ml at day 8, accounted for predominantly by increases in CD3-CD56+ NK cells from 0.07 to 0.25 million/ml (about 3.5-fold) and CD3+CD8+ T cells from 0.29 to 0.64 million/ml (2-fold). CD3+CD4+ T cells numbers were increased less, from 0.51 to 0.67 million/ml. CD19+ B cells did not survive the 8-day culture (data not shown).

Bottom Line: Here, using magnetic beads and fluorescence-activated cell sorting, we extensively phenotype the Tck effector cells and conclude that effector function resides within the CD4+CD45RO+, CCR7-, CD49dhigh population, and that these cells are derived from the effector memory CD4+ T cells in resting blood.Moreover, blockade of these ligands also resulted in inhibition of spontaneous tumour necrosis factor-alpha production in RA synovial mononuclear cell cultures.Taken together, these data strengthen our understanding of T-cell effector function, highlight the multiple involvement of different cell surface ligands in cell-cell contact and, provide novel insights into the pathogenesis of inflammatory RA disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Kennedy Institute of Rheumatology Division, Imperial College Faculty of Medicine, Aspenlea Road, London, W6 8LH, UK. f.brennan@imperial.ac.uk

ABSTRACT

Background: Previously we described a system whereby human peripheral blood T cells stimulated for 8 days in a cytokine cocktail acquired effector function for contact-dependent induction of proinflammatory cytokines from monocytes. We termed these cells cytokine-activated (Tck) cells and found that the signalling pathways elicited in the responding monocytes were identical whether they were placed in contact with Tck cells or with T cells isolated from rheumatoid arthritis (RA) synovial tissue.

Methods: Here, using magnetic beads and fluorescence-activated cell sorting, we extensively phenotype the Tck effector cells and conclude that effector function resides within the CD4+CD45RO+, CCR7-, CD49dhigh population, and that these cells are derived from the effector memory CD4+ T cells in resting blood.

Results: After stimulation in culture, these cells produce a wide range of T-cell cytokines, undergo proliferation and differentiate to acquire an extensively activated phenotype resembling RA synovial T cells. Blocking antibodies against CD69, CD18, or CD49d resulted in a reduction of tumour necrosis factor-alpha production from monocytes stimulated with CD4+CD45RO+ Tck cells in the co-culture assay. Moreover, blockade of these ligands also resulted in inhibition of spontaneous tumour necrosis factor-alpha production in RA synovial mononuclear cell cultures.

Conclusion: Taken together, these data strengthen our understanding of T-cell effector function, highlight the multiple involvement of different cell surface ligands in cell-cell contact and, provide novel insights into the pathogenesis of inflammatory RA disease.

Show MeSH
Related in: MedlinePlus