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Fibroblasts from different sites may promote or inhibit recruitment of flowing lymphocytes by endothelial cells.

McGettrick HM, Smith E, Filer A, Kissane S, Salmon M, Buckley CD, Rainger GE, Nash GB - Eur. J. Immunol. (2009)

Bottom Line: Interestingly, co-culture with dermal fibroblasts caused a marked reduction in cytokine-induced adhesion, while synovial fibroblasts had variable effects depending on their source.Exogenous IL-6 was also found to inhibit response to TNF-alpha+IFN-gamma.Normal stromal fibroblasts appear to regulate the cytokine-sensitivity of vascular endothelium, while fibroblasts associated with chronic inflammation bypass this and develop a directly inflammatory phenotype.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cardiovascular Sciences, The Medical School, The University of Birmingham, Birmingham, UK.

ABSTRACT
We examined the hypothesis that stromal fibroblasts modulate the ability of endothelial cells (EC) to recruit lymphocytes in a site-specific manner. PBL were perfused over HUVEC that had been cultured with fibroblasts isolated from the inflamed synovium or the skin of patients with rheumatoid arthritis or osteoarthritis, or from normal synovium, with or without exposure to the inflammatory cytokines TNF-alpha+IFN-gamma. Fibroblasts from inflamed synovium, but no others, caused unstimulated HUVEC to bind flowing lymphocytes. This adhesion was supported by alpha(4)beta(1)-VCAM-1 interaction and stabilised by activation of PBL through CXCR4-CXCL12. Antibody neutralisation of IL-6 during co-culture effectively abolished the ability of EC to bind lymphocytes. Cytokine-stimulated EC supported high levels of lymphocyte adhesion, through the presentation of VCAM-1, E-selectin and chemokine(s) acting through CXCR3. Interestingly, co-culture with dermal fibroblasts caused a marked reduction in cytokine-induced adhesion, while synovial fibroblasts had variable effects depending on their source. In the dermal co-cultures, neutralisation of IL-6 or TGF-beta caused partial recovery of cytokine-induced lymphocyte adhesion; this was complete when both were neutralised. Exogenous IL-6 was also found to inhibit response to TNF-alpha+IFN-gamma. Normal stromal fibroblasts appear to regulate the cytokine-sensitivity of vascular endothelium, while fibroblasts associated with chronic inflammation bypass this and develop a directly inflammatory phenotype. Actions of IL-6 might be pro-inflammatory or anti-inflammatory, depending on the local milieu.

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Effect of neutralising IL-6 or TGF-β, or of adding IL-6 on lymphocyte adhesion. (A) Neutralisation in unstimulated endothelial-RA synovial fibroblasts co-cultures; (B) neutralisation in cytokine-stimulated endothelial-RA dermal fibroblasts co-cultures; (C) addition of IL-6 to unstimulated or cytokine-stimulated EC mono-cultures. Data are mean±SEM from at least three experiments, using at least three different donors for each cell type. In all cases, ANOVA showed a significant effect of treatment on lymphocyte adhesion (p<0.01). *p<0.05 and **p<0.01 by Bonferroni test compared with co-cultures without neutralisation in (A) and (B), or to cytokine-treated mono-culture without IL-6 in (C).
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fig05: Effect of neutralising IL-6 or TGF-β, or of adding IL-6 on lymphocyte adhesion. (A) Neutralisation in unstimulated endothelial-RA synovial fibroblasts co-cultures; (B) neutralisation in cytokine-stimulated endothelial-RA dermal fibroblasts co-cultures; (C) addition of IL-6 to unstimulated or cytokine-stimulated EC mono-cultures. Data are mean±SEM from at least three experiments, using at least three different donors for each cell type. In all cases, ANOVA showed a significant effect of treatment on lymphocyte adhesion (p<0.01). *p<0.05 and **p<0.01 by Bonferroni test compared with co-cultures without neutralisation in (A) and (B), or to cytokine-treated mono-culture without IL-6 in (C).

Mentions: We previously identified IL-6 as a soluble promoter of the adhesion of neutrophils in a similar unstimulated co-culture model 9. Here, significantly more IL-6 was produced in endothelial co-cultures with synovial fibroblasts than in co-cultures with dermal fibroblasts from the same RA donors in the absence of exogenous cytokines (2.92±0.52 ng/mL compared with 0.64±0.14 ng/mL, respectively; mean±SEM n=9, p<0.01 by paired t-test). Neutralising antibodies against IL-6 or TGF-β were added to the co-cultures for 24 h before the assay. Neutralisation of IL-6 caused a significant reduction in lymphocyte adhesion to EC cultured with synovial fibroblasts, while neutralisation of TGF-β had no significant effect (Fig. 5A).


Fibroblasts from different sites may promote or inhibit recruitment of flowing lymphocytes by endothelial cells.

McGettrick HM, Smith E, Filer A, Kissane S, Salmon M, Buckley CD, Rainger GE, Nash GB - Eur. J. Immunol. (2009)

Effect of neutralising IL-6 or TGF-β, or of adding IL-6 on lymphocyte adhesion. (A) Neutralisation in unstimulated endothelial-RA synovial fibroblasts co-cultures; (B) neutralisation in cytokine-stimulated endothelial-RA dermal fibroblasts co-cultures; (C) addition of IL-6 to unstimulated or cytokine-stimulated EC mono-cultures. Data are mean±SEM from at least three experiments, using at least three different donors for each cell type. In all cases, ANOVA showed a significant effect of treatment on lymphocyte adhesion (p<0.01). *p<0.05 and **p<0.01 by Bonferroni test compared with co-cultures without neutralisation in (A) and (B), or to cytokine-treated mono-culture without IL-6 in (C).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2821685&req=5

fig05: Effect of neutralising IL-6 or TGF-β, or of adding IL-6 on lymphocyte adhesion. (A) Neutralisation in unstimulated endothelial-RA synovial fibroblasts co-cultures; (B) neutralisation in cytokine-stimulated endothelial-RA dermal fibroblasts co-cultures; (C) addition of IL-6 to unstimulated or cytokine-stimulated EC mono-cultures. Data are mean±SEM from at least three experiments, using at least three different donors for each cell type. In all cases, ANOVA showed a significant effect of treatment on lymphocyte adhesion (p<0.01). *p<0.05 and **p<0.01 by Bonferroni test compared with co-cultures without neutralisation in (A) and (B), or to cytokine-treated mono-culture without IL-6 in (C).
Mentions: We previously identified IL-6 as a soluble promoter of the adhesion of neutrophils in a similar unstimulated co-culture model 9. Here, significantly more IL-6 was produced in endothelial co-cultures with synovial fibroblasts than in co-cultures with dermal fibroblasts from the same RA donors in the absence of exogenous cytokines (2.92±0.52 ng/mL compared with 0.64±0.14 ng/mL, respectively; mean±SEM n=9, p<0.01 by paired t-test). Neutralising antibodies against IL-6 or TGF-β were added to the co-cultures for 24 h before the assay. Neutralisation of IL-6 caused a significant reduction in lymphocyte adhesion to EC cultured with synovial fibroblasts, while neutralisation of TGF-β had no significant effect (Fig. 5A).

Bottom Line: Interestingly, co-culture with dermal fibroblasts caused a marked reduction in cytokine-induced adhesion, while synovial fibroblasts had variable effects depending on their source.Exogenous IL-6 was also found to inhibit response to TNF-alpha+IFN-gamma.Normal stromal fibroblasts appear to regulate the cytokine-sensitivity of vascular endothelium, while fibroblasts associated with chronic inflammation bypass this and develop a directly inflammatory phenotype.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cardiovascular Sciences, The Medical School, The University of Birmingham, Birmingham, UK.

ABSTRACT
We examined the hypothesis that stromal fibroblasts modulate the ability of endothelial cells (EC) to recruit lymphocytes in a site-specific manner. PBL were perfused over HUVEC that had been cultured with fibroblasts isolated from the inflamed synovium or the skin of patients with rheumatoid arthritis or osteoarthritis, or from normal synovium, with or without exposure to the inflammatory cytokines TNF-alpha+IFN-gamma. Fibroblasts from inflamed synovium, but no others, caused unstimulated HUVEC to bind flowing lymphocytes. This adhesion was supported by alpha(4)beta(1)-VCAM-1 interaction and stabilised by activation of PBL through CXCR4-CXCL12. Antibody neutralisation of IL-6 during co-culture effectively abolished the ability of EC to bind lymphocytes. Cytokine-stimulated EC supported high levels of lymphocyte adhesion, through the presentation of VCAM-1, E-selectin and chemokine(s) acting through CXCR3. Interestingly, co-culture with dermal fibroblasts caused a marked reduction in cytokine-induced adhesion, while synovial fibroblasts had variable effects depending on their source. In the dermal co-cultures, neutralisation of IL-6 or TGF-beta caused partial recovery of cytokine-induced lymphocyte adhesion; this was complete when both were neutralised. Exogenous IL-6 was also found to inhibit response to TNF-alpha+IFN-gamma. Normal stromal fibroblasts appear to regulate the cytokine-sensitivity of vascular endothelium, while fibroblasts associated with chronic inflammation bypass this and develop a directly inflammatory phenotype. Actions of IL-6 might be pro-inflammatory or anti-inflammatory, depending on the local milieu.

Show MeSH
Related in: MedlinePlus