NKT-deficient Jα18 KO mice () were primed with ovalbumin and alum as in our standard protocol in Materials and methods

On day 11, the mice received 3.1 million purified NK/NKT cells containing 1 million WT NKT cells or DN TNFR25-tg NKT cells (DN NKT) or PBS by i.v. adoptive transfer, as indicated. The mice were exposed to ovalbumin aerosol on day 12 and analyzed on day 15. WT mice and Jα18 KO mice receiving WT NKT cells by adoptive transfer served as positive controls for induction of lung inflammation. Jα18 KO mice, immunized and ovalbumin aerosolized without adoptive cell transfer, served as negative controls. The data of three independent experiments and two mice in each group are shown. (A) Eosinophils in BALF. Error bars represent the mean ± the SEM. (B and C) Cytokine and TL1A mRNA expression in bronchial lymph nodes (LN; B) and lung parenchyma (C) determined by real time Taqman PCR. The fold increase or decrease of mRNA in Jα18 KO mice (Jα) adoptively transferred with WT NKT cells over mice adoptively transferred with DN TNFR25-tg NKT (DN NKT) cells is plotted.<p><b>Copyright information:</b></p><p>Taken from "Essential role of TNF receptor superfamily 25 (TNFRSF25) in the development of allergic lung inflammation"</p><p></p><p>The Journal of Experimental Medicine 2008;205(5):1037-1048.</p><p>Published online 12 May 2008</p><p>PMCID:PMC2373837.</p><p></p

Mice were immunized and subjected to ovalbumin aerosol according to our standard protocol

Administration of 50 μg blocking (L4G6) or nonblocking (L3A10) anti-Tl1A, or control hamster IgG i.p. was started at the time indicated relative to aerosol exposure and continued daily until analysis, which was at 76 h after aerosol. In the 72-h time points, anti-TL1A was administered 4 h before analysis. A, B, and C are separate experiments testing different schedules and controls. Note that nonblocking TL1A (L3A10) does not affect eosinophil exudation, similar to hamster IgG. (A and B) Data from three experiments with two mice. (C) Data from two experiments with five mice. (D and E) Histopathology of lung sections stained with HE and PAS. Five sections from each of three mice in each group were evaluated in a blinded fashion according to the scoring system described in Materials and methods. (F and G) Relative frequency of CD4 and CD8 cells in lung parenchyma after aerosol exposure and blockade of TL1A for different periods of time. Single-cell suspensions were analyzed by flow cytometry gating on the lymphocyte gate. (H–J) RNA was isolated from whole lungs and analyzed by real-time PCR as in . Error bars represent the mean ± the SEM.<p><b>Copyright information:</b></p><p>Taken from "Essential role of TNF receptor superfamily 25 (TNFRSF25) in the development of allergic lung inflammation"</p><p></p><p>The Journal of Experimental Medicine 2008;205(5):1037-1048.</p><p>Published online 12 May 2008</p><p>PMCID:PMC2373837.</p><p></p

(A) Diminished cellular exudation in BALF in anti-TL1A (L4G6)–treated mice

Mice were primed i.p. on day 0 and 5 with 66 μg ovalbumin absorbed to alum. On day 12, mice were aerosol challenged with 0.5% ovalbumin in PBS for 1 h using an ultrasonic nebulizer. Mice received 4 daily doses of 50 μg purified L4G6-IgG i.p. (anti-TL1A), beginning 1 d before aerosol. Controls received the same amount and schedule of purified hamster IgG. All mice were analyzed 3 d after aerosol antigen exposure ( = 4; representative of >10 experiments). *, P < 0.05; **, P < 0.01. (B) TL1A-blocking antibody L4G6 suppresses mucus production and lung inflammation. Lung histology after PAS staining after treatment of mice with control IgG (top) or L4G6-IgG (anti-TL1A; bottom). Notice the lack of mucus production and cell infiltration in L4G6-treated animals (arrows point to mucus in mice treated with control IgG). Experiments were repeated three times. (C) Diminished IL-5 and -13 production by ovalbumin restimulated bronchial lymph node cells after TL1A blockade with L4G6. Bronchial lymph node cells were harvested 3 d after aerosol and restimulated in vitro with 100 μg/ml ovalbumin for 4 d. IL-4 was not detectable (not depicted), even in the absence of anti-TL1A. = 4; **, P < 0.01; ***, P < 0.001. Experiments were repeated more than three times. (D) Cytokine expression in lung parenchyma after ovalbumin aerosol exposure. Lungs were harvested 1, 2, or 3 d after ovalbumin aerosol treatment. RNA was extracted, and after reverse transcription it was analyzed by Taqman PCR. Values are normalized to GAPDH cDNA and expressed as the fold increase of ovalbumin aerosol–treated over untreated mice. (E) Blocking anti-TL1A antibody L4G6 suppresses ovalbumin-induced cytokine expression in lung parenchyma. Mice were immunized twice with ovalbumin/alum, as described. 1 d before ovalbumin aerosol and for the next 3 d, mice received 50 μg blocking TL1A antibody L4G6 or control IgG i.p. Lungs were analyzed for expression of cytokine mRNA on day 1–3 after aerosol administration by Taqman PCR, as above. Data are presented as anti-TL1A–induced suppression of cytokine mRNA over control IgG.<p><b>Copyright information:</b></p><p>Taken from "Essential role of TNF receptor superfamily 25 (TNFRSF25) in the development of allergic lung inflammation"</p><p></p><p>The Journal of Experimental Medicine 2008;205(5):1037-1048.</p><p>Published online 12 May 2008</p><p>PMCID:PMC2373837.</p><p></p

(A) TL1A is not expressed on resting lymphocytes and up-regulated on activated T cells (top 6 graphs)

Resting splenocyte cell suspensions were gated using the respective labeled antibody as a population marker and the TL1A histogram displayed. Red curve, anti-TL1A; black curve, isotype control (bottom 3 graphs). Splenocytes were activated for 24 h with plate-bound anti-CD3 or with LPS and stained with anti-TL1A and with the population marker, as indicated. After gating on the population marker, TL1A expression on activated cells is shown as blue/shaded histogram. Red curve, resting cells; black curve, isotype control. Representative of more than three experiments. (B) TNFR25 and TL1A expression on cDNA transfected P815 and EL4. Transfected (right curve in each histogram) and untransfected cells were stained with the appropriate antibody and isotype controls and analyzed by flow cytometry. (C) TNFR25 activates NF-κB when triggered by agonistic antibody 4C12, by soluble TL1A or by membrane-bound TL1A. NF-κB activation was measured in EL4 cells transfected with TNFR25 in response to TNFR25 triggering. Cells were treated with the agonistic anti-TNFR25 antibody 4C12 (5 μg/ml) for 50 min; soluble TL1A was given for 25, 50, or 70 min, as indicated in the form of 25% supernatants from TL1A-transfected EL4 cells; membrane-bound TL1A (MTL1A) was given for 50 min by adding TL1A-transfected EL4-cells directly to TNFR25-transfected EL4. Controls received EL4 (untransfected) supernatants for 50 min. Nuclear extracts were prepared and analyzed by EMSA; the arrow indicates activated NF-κB. (D) Anti-TL1A antibody L4G6 blocks TL1A induced cell death of TNFR25-transfected cells. Soluble TL1A harvested from supernatants of P815-TL1A–transfected cells were mixed with Cr-labeled P815-TNFR25 target cells. Different anti-murine TL1A monoclonal antibodies were added into the assay, and Cr release was analyzed 5 h later. L4G6 antibody completely blocked the ability of TL1A to induce apoptosis in TNFR25-transfected P815 cells.<p><b>Copyright information:</b></p><p>Taken from "Essential role of TNF receptor superfamily 25 (TNFRSF25) in the development of allergic lung inflammation"</p><p></p><p>The Journal of Experimental Medicine 2008;205(5):1037-1048.</p><p>Published online 12 May 2008</p><p>PMCID:PMC2373837.</p><p></p

Bacterial Vaginosis Is Associated with Loss of Gamma Delta T Cells in the Female Reproductive Tract in Women in the Miami Women Interagency HIV Study (WIHS): A Cross Sectional Study

<div><p>Bacterial vaginosis (BV) is the most common female reproductive tract infection and is associated with an increased risk of acquiring and transmitting HIV by a mechanism that is not well understood. Gamma delta (GD) T cells are essential components of the adaptive and innate immune system, are present in the female reproductive tract, and play an important role in epithelial barrier protection. GD1 cells predominate in the mucosal tissue and are important in maintaining mucosal integrity. GD2 cells predominate in peripheral blood and play a role in humoral immunity and in the immune response to pathogens. HIV infection is associated with changes in GD T cells frequencies in the periphery and in the female reproductive tract. The objective of this study is to evaluate if changes in vaginal flora occurring with BV are associated with changes in endocervical GD T cell responses, which could account for increased susceptibility to HIV. Seventeen HIV-infected (HIV+) and 17 HIV-uninfected (HIV-) pre-menopausal women underwent collection of vaginal swabs and endocervical cytobrushes. Vaginal flora was assessed using the Nugent score. GD T cells were assessed in cytobrush samples by flow cytometry. Median Nugent score was 5.0 and 41% of women had abnormal vaginal flora. In HIV uninfected women there was a negative correlation between Nugent score and cervical GD1 T cells (b for interaction = - 0.176, p<0.01); cervical GD1 T cells were higher in women with normal vaginal flora than in those with abnormal flora (45.00% vs 9.95%, p = 0.005); and cervical GD2 T cells were higher in women with abnormal flora than in those with normal flora (1.70% vs 0.35%, p = 0.023). GD T cells in the genital tract are protective (GD1) and are targets for HIV entry (GD2). The decrease in cervical GD1 and increase in GD2 T cells among women with abnormal vaginal flora predisposes women with BV to HIV acquisition. We propose to use GD T cell as markers of female genital tract vulnerability to HIV.</p></div

Comparison of frequencies of endocervical GD2 T cells in HIV infected and uninfected women with normal and abnormal vaginal flora.

<p>VF: vaginal flora. Normal VF is defined as Nugent score less than 4. Abnormal VF is defined as Nugent score of 4 or greater than 4. In HIV- women, frequency of GD2 T cells was higher in women with abnormal vaginal flora than in those with normal vaginal flora (1.79 vs 0.55, p = 0.023). A 2 (HIV+, HIV-) x 2 (normal VF, abnormal VF) ANOVA indicated that there was a nonsignificant main effect of vaginal flora, <i>F</i>(1,22) = .03, <i>p</i> = .876, but a nonsignificant main effect of HIV status, <i>F</i>(1,22) = .43, <i>p</i> = .520. The interaction of vaginal flora and HIV status was statistically significant, <i>F</i>(1,22) = 5.67, <i>p</i> = .028. The interaction effect was further analyzed using pairwise comparisons with a Bonferroni correction, which showed that there was a statistically significant difference in GD2 T cells among HIV- women with abnormal vaginal flora (Nugent score more than 4) compared to those with normal vaginal flora (<i>p</i> = 0.023), but not among HIV+ women (<i>p</i> = 0.171).</p

pH, Nugent score, vaginal flora, bacterial vaginosis and frequency of GD T cells by HIV status (N = 34).

<p>pH, Nugent score, vaginal flora, bacterial vaginosis and frequency of GD T cells by HIV status (N = 34).</p

Comparison of percentages of endocervical GD1 T cells in HIV infected and uninfected women with normal and abnormal vaginal flora.

<p>VF: vaginal flora. Normal VF is defined as Nugent score less than 4. Abnormal VF is defined as Nugent score of 4 or greater than 4. In HIV- women, frequency of GD1 T cells was higher in women with normal vaginal flora than in those with abnormal vaginal flora (45.00 vs 9.95, p = 0.005). Results from a 2 (HIV+, HIV-) x 2 (normal VF, abnormal VF) ANOVA showed that there was a nonsignificant main effect of abnormal vaginal flora, <i>F</i>(1,33) = 2.12, <i>p</i> = 0.155, but a statistically significant main effect of HIV status, <i>F</i>(1,33) = 10.34, <i>p</i> = 0.003. The interaction of BV and HIV status was statistically significant, <i>F</i>(1,33) = 7.05, <i>p</i> = 0.013. Further analysis of the interaction effect of BV and HIV status using pairwise comparisons with a Bonferroni correction showed that there was a statistically significant difference in GD1 T cells among HIV- women with abnormal vaginal flora compared to those without abnormal vaginal flora (<i>p</i> = 0.005), but not among HIV+ women (<i>p</i> = 0.424).</p