(A and B) Flow cytometry analysis of cells isolated from the organs of 8–12-wk-old mice

Plots are gated on CD3 cells (A) or GFP CD3 cells (B). Numbers indicate mean percent cells in quadrants ± SD obtained with at least three mice. LPLs, lamina propria lymphocytes isolated from small intestine; mLN, mesenteric LNs; BM, bone marrow. (C) Immunofluorescence histology of RORγt cells in the small intestine of mice. Most RORγt cells in villi are T cells, whereas RORγt cells in cryptopatches located between crypts are CD3 LTi cells. Bar, 50 μm. (D) Expression of CD4 and CD8α by spleen GFPTCR-β and lung or GFPTCR-δ cells.<p><b>Copyright information:</b></p><p>Taken from "In vivo equilibrium of proinflammatory IL-17 and regulatory IL-10 Foxp3 RORγt T cells"</p><p></p><p>The Journal of Experimental Medicine 2008;205(6):1381-1393.</p><p>Published online 9 Jun 2008</p><p>PMCID:PMC2413035.</p><p></p

MACS-sorted naive (CD62L) CD4 T cells from the spleens of mice were stimulated in duplicates with anti-CD3 and anti-CD28 in the presence of blocking anti–IFN-γ and anti–IL-4 antibodies and the indicated cytokines or RA

After different periods of time, cells were restimulated with PMA/ionomycin for 5 h and analyzed by flow cytometry for the expression of GFP, Foxp3, IL-17, and IL-10. All plots are gated on TCR-β cells, except plots for IL-10 that are gated on GFPTCR-β cells. Numbers indicate percent cells in quadrants. Data are representative of three independent experiments.<p><b>Copyright information:</b></p><p>Taken from "In vivo equilibrium of proinflammatory IL-17 and regulatory IL-10 Foxp3 RORγt T cells"</p><p></p><p>The Journal of Experimental Medicine 2008;205(6):1381-1393.</p><p>Published online 9 Jun 2008</p><p>PMCID:PMC2413035.</p><p></p

(A) Cells isolated from the spleen and mesenteric LNs of mice were sorted into eight distinct populations based on their expression of GFP, CD3, TCR-β, TCR-δ, CD4, and CD25 (Fig

S1), and gene expression was assessed using real-time PCR. Ct values were normalized to the mean Ct of five housekeeping genes. Data are the mean of two or three independent experiments. (B) Foxp3 RORγt T cells express IL-10. Cells isolated from LNs of mice were restimulated in vitro with PMA/ionomycin for 5 h and subjected to intracellular staining for GFP, IL-17, Foxp3, and IL-10 or an isotype control. Numbers indicate percent cells in quadrants. Results are representative of at least three individual experiments.<p><b>Copyright information:</b></p><p>Taken from "In vivo equilibrium of proinflammatory IL-17 and regulatory IL-10 Foxp3 RORγt T cells"</p><p></p><p>The Journal of Experimental Medicine 2008;205(6):1381-1393.</p><p>Published online 9 Jun 2008</p><p>PMCID:PMC2413035.</p><p></p

(A) mice were treated with DSS in the drinking water for 6 d, followed by water for 10 d

This protocol was repeated for a total of three cycles. After the last cycle, cells isolated from the colon were restimulated in vitro with PMA/ionomycin for 5 h and subjected to intracellular staining for GFP, IL-17, and Foxp3. Histograms (from left to right) report percent GFPTCR-β cell subsets in total T cells (see ), total numbers of RORγt Tαβ cells present in the organ, and the ratio of IL-17–producing to Foxp3 cells within RORγt Tαβ cells (see ). Right panels show immunofluorescence histology of a colon from a healthy or a treated mouse. Bar, 100 μm. (B) mice were infected intranasally with 100 PFUs of influenza A virus for 7 d. Cells were then isolated from the lung and processed as in A. Right panels show immunofluorescence histology of a lung from healthy or an infected mouse. Bar, 50 μm. (C) Cells were isolated from the mesenteric LNs of a 4-mo-old × mouse and processed as in A. Right panels show immunofluorescence histology of a mesenteric LN from a normal or a tumor-bearing mouse. Bar, 100 μm. Data shown are representative of at least three independent experiments. Three to four mice were analyzed per group. *, P < 0.05 as compared with control (mock-treated or WT mice).<p><b>Copyright information:</b></p><p>Taken from "In vivo equilibrium of proinflammatory IL-17 and regulatory IL-10 Foxp3 RORγt T cells"</p><p></p><p>The Journal of Experimental Medicine 2008;205(6):1381-1393.</p><p>Published online 9 Jun 2008</p><p>PMCID:PMC2413035.</p><p></p

In patients with acute bacterial meningitis, (A) CCL20 cerebrospinal fluid (CSF) levels were significantly elevated compared with controls.

<p>CCL20 CSF levels of patients with pneumococcal meningitis correlated to (B) CSF white blood cell (WBC) counts, (C) CSF blood glucose ratio, and (D) CSF protein levels (see text for details).</p

CCR6-deficient mice suffer from increased brain edema after antibiotic therapy.

<p>(A) Infected mice with CCR6-deficiency developed more pronounced brain edema than infected wild type mice 48 h after infection, reflected in an increase in the estimated brain volume. (B) At the same time point, an increase of brain albumin content was noted in infected <i>Ccr6</i>^−/− mice, indicating blood-brain barrier disruption. These differences were only seen after but not before initiation of antibiotic therapy (24 h after infection). (C–G) Intracranial bleeding was similar in infected <i>Ccr6</i>^−/− and wild type mice. (*) p<0.05 compared with WT control animals. Number of animals: 24 h: <i>Ccr6</i>^−/− n = 12, WT n = 12; 48 h: <i>Ccr6</i>^−/− n = 12, WT n = 13.</p

CCL20 is expressed mainly during acute pneumococcal meningitis.

<p>(A) Increased CCL20 levels were found in mice brain homogenates during acute bacterial meningitis using ELISA. After initiation of antibiotic therapy (starting 24 h after infection), they decreased quickly to normal ((*) p<0.01 compared with uninfected controls). In uninfected control mice, (B, C) only a very subtle CCL20-positive staining was observed (white arrow). In contrast, in animals with pneumococcal meningitis, CCL20-positive staining was found in (D) epithel cells of the choroid plexus (black arrow) and (E) the subarachnoid inflammatory infiltrate (#). Number of animals: 6 h: n = 6, 24 h: n = 9, 30 h: n = 6, 48 h: n = 6, 72 h: n = 6, and 120 h: n = 7. Uninfected animals were used as controls (n = 11).</p

CCR6-deficient mice are more susceptible to pneumococcal meningitis.

<p>CCR6-deficient mice were more strongly affected from a clinical perspective by infection with <i>Streptococcus pneumoniae</i> than wild type controls. This was reflected (A) in increased clinical scores and (B) mortality. (C) CSF pleocytosis was lower in infected CCR6-deficient animals 24 h and 48 h after infection. (D) This was associated with higher brain bacterial titers at the time of initiation of antibiotic therapy (24 h after infection). (*) p<0.05 compared with infected wild type animals. Number of animals: 24 h: <i>Ccr6</i>^−/− n = 12, WT n = 12; 48 h: <i>Ccr6</i>^−/− n = 12, WT n = 13.</p

The pro-inflammatory effect of the CCL20/CCR6 axis seems independent of IL-17 production.

<p>IL17 was up-regulated in the CSF of (A) humans and (B) mice with pneumococcal meningitis. (C) However, antibody blockage of IL-17 in experimental pneumococcal meningitis did not lead to a reduction of inflammation in the CSF. P<0.05 as compared with uninfected controls. Number of animals: n = 5 mice per group.</p