Lieve Coorevits

Reciprocal changes of Foxp3 expression in blood and intestinal mucosa in IBD patients responding to infliximab.

Background: Immune therapies may act in inflammatory bowel diseases (IBD) by modulating regulatory T cells (Tregs). Therefore, we investigated the effect of infliximab (IFX) therapy on Forkhead box protein3 (Foxp3) T cells in blood and intestinal mucosa from Crohns disease (CD) and ulcerative colitis (UC). Methods: Forty patients with active IBD (23 CD / 17 UC) were treated with IFX 5 mg/kg intravenously at weeks 0, 2, 6, and each 8 weeks thereafter. Blood samples were obtained before every infusion and T‐lymphocyte subsets were characterized by flow cytometry. Foxp3 expression in intestinal biopsies from 43 patients with active IBD (19 CD / 24 UC) before and after IFX infusion and from 6 controls were assessed by quantitative reverse‐transcription polymerase chain reaction and immunohistochemistry. Plasma C‐reactive protein (CRP), clinical response, and endoscopic healing data were collected in parallel. Results: IFX therapy resulted in a significant and sustained relative increase of CD4+CD25+Foxp3+ Treg and of CD4+CD25−Foxp3+ Treg cells in peripheral blood (both P < 0.0001 compared to baseline), particularly in responders (both P < 0.05 compared to nonresponders). The change in CRP over time inversely correlated with the increase of CD25+Foxp3+ cells (P < 0.001, r = −0.39) and durable clinical response was associated with a sustained increase of circulating Foxp3+ cells. Surprisingly, IFX therapy downregulated mucosal mRNA and protein expression of Foxp3 in UC and CD responders (both P < 0.001) but not in nonresponders. Conclusions: IFX therapy has opposite effects in Foxp3+ Treg cells in blood and gut mucosa, which suggests a redistribution of this important T‐cell subset. Inflamm Bowel Dis 2010

Production of granulocyte-macrophage colony-stimulating factor by T cells is regulated by B7 and IL-1 beta.

Granulocyte–macrophage colony‐stimulating factor (GM‐CSF) has proliferation‐ and differentiation‐inducing effects on immature myeloid cells in the bone marrow, and it can modulate the function of several types of mature myeloid cells. We have stimulated purified human T cells with immobilized anti‐CD3 or mitogenic anti‐CD2 (a combination of monoclonal antibodies 9‐1 and 9.6) which could induce GM‐CSF production. The cytokines interleukin‐1β (IL‐1β) and IL‐2 strongly enhanced GM‐CSF production, while IL‐4, IL‐6, GM‐CSF, interferon‐γ (IFN‐γ) and tumour necrosis factor (TNF) had no effect. Activation of protein kinase C by phorbol myristate acetate or triggering of CD28 on T cells by monoclonal antibody 9.3 provided accessory signals for enhanced GM‐CSF production in activated T cells. Most important, the addition of mouse cells transfected with human B7‐1 (CD80), a natural ligand for CD28, provided a potent accessory signal for GM‐CSF production by activated T cells, which could not be blocked by cyclosporin A. The effect of IL‐1β was in fact indirect, and resulted from enhanced IL‐2 production, while the effect of B7 resulted from both IL‐2‐dependent and IL‐2‐independent pathways. We conclude that antigen‐presenting cells (APC) can up‐regulate GM‐CSF production through IL‐1β and through CD28 triggering by B7 molecules. As GM‐CSF itself up‐regulates B7 expression and IL‐1β production by APC, a bidirectional regulatory feedback pathway between APC and T cells seems to modulate GM‐CSF production.

Identification of an enriched CD4+ CD8α++ Cd8β+ T-cell subset among tumor-infiltrating lymphocytes in human renal cell carcinoma

Three‐color immunofluorescence flow‐cytometric analysis of freshly isolated tumor‐infiltrating lymphocytes (TIL) from patients with primary renal cell carcinoma (RCC) revealed a unique, not previously described TIL subset with a CD3+ CD4+ CD8α++ CD8β+ phenotype. This subset represented at least 5% of CD3+ TIL in 15 of 21 patients with clear cell RCC, whereas it was not or only marginally represented in patients with papillary RCC or sarcomatoid RCC. In one‐third of the patients with clear cell RCC, more than 20% of CD3+ TIL and in one patient more than half of the CD3+ TIL displayed this phenotype. The occurrence of this subset was not associated with pathological stage, tumor diameter, nuclear grade, cytogenetic abnormalities or vascular invasion in this patient cohort. When present, the CD3+ CD8α++ CD8β+ subset was detected in similar proportions in tumor tissue and tumor capsula, and it was also detected in adjacent non‐tumoral renal tissue, albeit in much lower proportions. Despite strong cell surface expression of various activation markers (CD69, CD54 and HLA‐DR), CD3+ CD4+ CD8α++ anti‐CD3‐redirected cytotoxicity assay. In contrast with CD3+ CD4+ CD8− cells from the same tumor sample, they were markedly deficient in IL‐2Rα up‐regulation following anti‐CD3 triggering. The possibility that these cells represent either anergic cells or a highly specialized effector population with a discrete, as yet undescribed function is discussed. Int. J. Cancer 71:178–182, 1997.

Phenotype, Cytokine Production and Cytolytic Capacity of Fresh (Uncultured) Tumour-Infiltrating T Lymphocytes in Human Renal Cell Carcinoma

We investigated the phenotype and functional capacities of tumour-infiltrating lymphocytes (TIL), freshly isolated from primary renal cell carcinoma (RCC) specimens (n = 20). Three-colour flow cytometry immunophenotyping revealed that RCC TIL consist mainly of CD3+ T cells, with a clear predominance of CD4- CD8+ over CD4+ CD8- T cells, and a marked population of CD4+ CD8+ T cells. Natural killer (NK) cells were also strongly represented (> 25% in 15 of 20 tumour samples), while B cells constituted a minor TIL subset (< 5% in 18 of 20 tumour samples). More importantly, the T and NK cells within the tumour displayed a significantly higher expression of the early activation marker CD69 than their counterparts in adjacent normal renal tissue and in peripheral blood. Expression of CD54 and of HLA-DR was also elevated on CD3+ TIL, and HLA-DR expression was further vigorously up-regulated following ex vivo stimulation with anti-CD3, all suggesting enhanced immune activity within the tumour microenvironment. CD3+ CD4+ TIL displayed a normal capacity to up-regulate CD25 expression and to secrete both Th1-type (IL-2, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma)) and Th2-type (IL-4, IL-5 and IL-10) cytokines upon triggering with anti-CD3. Furthermore, cytokine production was susceptible to modulation by CD28 costimulation. CD3+ CD8+ TIL, on the other hand, consistently demonstrated a poor up-regulation of CD25 upon triggering with anti-CD3, and displayed poor ex vivo cytolytic activity in an anti-CD3-redirected 4-h cytotoxicity assay against murine P815 cells. Collectively, our findings indicate that the CD3+ CD4+ TIL in RCC have normal functional capacities, whereas the proportionally major CD3+ CD8+ TIL are functionally impaired. The relevance of these findings to the in vivo local immune response in RCC is discussed.