Intranasal α-GalCer administration activates lung iNKT cells to secrete cytokines.

<p>Mice were intranasally administered with vehicle or α-GalCer. Two hours later, BALF was collected. (A) The absolute numbers of iNKT (CD3⁺PBS57 loaded CD1d tetramer⁺) cells were measured. (B) IFN-γ and IL-4 expression from iNKT cells was assayed by intracellular staining with anti-mouse IFN-γ and IL-4 Abs. The shaded area is an isotype control; the open area reflects cytokine expression. The percentage of IFN-γ and IL-4-producing iNKT cells were shown. (C) IL-4 and IFN-γ levels in the BALF were measured by ELISA. a-GC, α-GalCer. n = 5 mice for each group. *, p<0.05 using Mann-Whitney U test.</p

IL-4 enhances <i>Mmp12</i> expression in macrophages.

<p>Bone marrow derived macrophages were stimulated with different concentrations of IL-4 or IFN-γ and the expression of <i>Mmp12</i> was measured by quantitative RT-PCR. Summarized results of 4 independent experiments are shown. Each symbol represents an individual experiment and the horizontal lines represent the means. *, p<0.05 using Kruskal-Wallis one-way ANOVA followed Dunn's post test.</p

Intranasal α-GalCer administration induced acute inflammation in the lung.

<p>Mice were intranasally administered with vehicle or α-GalCer. BALF was collected at days 0, 3 and 7. (A) BALF total cells and the absolute numbers of macrophages, lymphocytes, neutrophils, and eosinophils were measured at the indicated time points. (B) The relative expressions of cytokines and chemokines compared to positive control (set to a value of 1) of the assay kit in BALF at 3 days after α-GalCer administration of mice were determined by a dot blot immunoassay. Samples were pooled from 5–8 mice in two independent experiments. (C) Cytokines IFN-γ, IL-4, TNF-α, IL-17, and IL-13 in the BALF were assayed by ELISA. n = 5–8 mice for each group. *, p<0.05; **, p<0.01; ***, p<0.001 using Kruskal-Wallis one-way ANOVA followed Dunn's post test.</p

Increased airway inflammation in mice repeatedly administered with α-GalCer.

<p>Mice received intranasal α-GalCer administration once a week for 6 weeks and airway inflammation was measured 2 weeks after the last administration. (A) Analysis of lung function. Changes in airway resistance, dynamic compliance, and airway elastance were measured on mechanically ventilated mice, in response to increasing doses of methacholine. (B) Representative results of H-E examination of lung sections (x100 magnification). Blue arrowheads indicate cell infiltration in the lung. Black arrowheads indicate airspace enlargement in the lung. (C) The total numbers of cells present in BALF. (D) The absolute numbers of macrophages, lymphocytes, neutrophils, and eosinophils, present in BALF. Mac, macrophage; Lym, lymphocyte; Neu, neutrophil; Eos, eosinophil. (E) The absolute numbers of CD4⁺ and CD8⁺ T cells were examined. (F) CD8/CD4 ratios of T cells were measured. (G) Lung tissues were isolated and the expression of TNF-α and IL-6 was detected by quantitative RT-PCR. n = 9–14 mice for each group. (H) Macrophages from BALF were isolated and their expression of TNF-α and IL-6 was detected by quantitative RT-PCR. Individual symbols represent a sample pooled from 3–5 mice and the horizontal lines represent the mean values. a-GC, α-GalCer. *, p<0.05; **, p<0.01; ***, p<0.001 using Mann-Whitney U test.</p

Anti-IL-4 Abs reduced emphysema and mucus production in mice repeatedly administered with α-GalCer.

<p>Mice were injected with neutralizing Abs to IL-4 or isotype controls 1 hour prior to each α-GalCer administration. Histopathological examination was measured 2 weeks after the last α-GalCer administration. (A) The expression of <i>Mmp-12</i> in lung tissues was analyzed by quantitative RT-PCR. (B) The expression of <i>Mmp-12</i> in lung macrophages was analyzed by quantitative RT-PCR. (C) The mean linear intercept of the alveolar septa was measured. (D) The absolute numbers of macrophages, neutrophils, eosinophils, and lymphocytes were measured. Mac, macrophage; Lym, lymphocyte; Neu, neutrophil; Eos, eosinophil. (E) The expression of <i>Muc5ac</i> in lung tissues was detected by quantitative RT-PCR. n = 9–16 mice for each group. *, p<0.05; **, p<0.01; ***, p<0.001 using Mann-Whitney U test.</p

The non-canonical Notch signaling is essential for the control of fertility in <i>Aedes aegypti</i>

<div><p>The Notch signaling pathway is a highly evolutionarily-conserved cell-cell signaling pathway that regulates many events during development. It plays a pivotal role in the regulation of fundamental cellular processes, such as cell proliferation, stem cell maintenance, and differentiation during embryonic and adult development. However, functions of Notch signaling in <i>Aedes aegypti</i>, the major mosquito vector for dengue, are largely unknown. In this study, we identified a unique feature of <i>A</i>. <i>aegypti</i> Notch (AaNotch) in the control of the sterile-like phenotype in female mosquitoes. Silencing AaNotch with a reverse genetic approach significantly reduced the fecundity and fertility of the mosquito. Silencing AaNotch also resulted in the prevention of micropyle formation, which led to impaired fertilization. In addition, JNK phosphorylation (a signaling molecule in the non-canonical Notch signaling pathway) was inhibited in the absence of AaNotch. Furthermore, treatment with a JNK inhibitor in the mosquito resulted in impaired fecundity and fertility. Taken together, our results demonstrate that non-canonical Notch signaling is essential for controlling fertility in the <i>A</i>. <i>aegypti</i> mosquito.</p></div

Chemical inhibition of JNK reduces egg tanning and hatching.

<p>(A) Total oviposited eggs from control, DMSO, or JNK inhibitor injected female mosquitoes (Scale bar = 0.5 mm.) (B) Melanized and non-melanized eggs (number in the parentheses denotes the total number of eggs counted). Y-axis is the percentage of melanized eggs of total eggs counted. (C) Eggs from control, DMSO, or JNK inhibitor (JNKi)-treated mosquitoes subjected to deoxygenation-induced hatching for melanized (MZ) and non-melanized (non-MZ) eggs. Number in the parentheses denotes total number of eggs examined. (D) Eggs from control, DMSO-, and JNK inhibitor-treated mosquitoes separated into melanized (MZ) and non-melanized (non-MZ) eggs. Scale bar = 10 μm. (E) Percentage of eggs with complete micropylar pore formation (Y-axis). Number in the parentheses denotes the number of eggs with complete micropylar formation divided by the total number counted.</p

Bilirubin concentrations and hydrogen peroxide levels increased similarly during the first few days.

<p>The correlation between these two variables was 0.454, <i>P</i><0.001. The left y-axis is the bilirubin concentration, whereas the right y-axis is the H₂O₂ (luminol) signal. The y-axis scale is logarithmic because of the high luminal signal values.</p

Enrollment of the study population.

<p>Enrollment of the study population.</p

A scatter plot figure with bilirubin (on the X-axis) and the corresponding hydrogen peroxide levels with Logarithmic transformation (on the Y-axis) was presented.

<p>There is a linear relationship between these 2 values. The regression equation is H₂O₂ signal = 3.371+0.108*Bilirubin. The P value is <0.0001.</p