Figure 1.
Airways hyperresponsiveness dose response to house dust mite exposure.
The indicated strains of mice (B6 = C57BL/6; Bc = BALB/c; B6 Itk−/− = C57BL/6/Itk−/−; Bc Itk−/− = BALB/c/Itk−/−) were exposed to A) 5; B) 25; and C) 100 µg of HDM per day for 10 consecutive days. Twenty four hours after the final exposure, animals were analyzed for hyperresponsiveness to increasing doses of methacholine as described in the materials and methods section (*p<0.05, Bc vs. B6, Bc vs. Bc Itk−/− , B6 vs. B6 Itk−/−, Bc Itk−/− vs. B6 Itk−/−, 1-way ANOVA). D) The Area Under the Curve (AUC) was determined for the methacholine response at each dose of HDM.
Figure 2.
Histopathological analysis of the dose response to house dust mite exposure.
The indicated strains of mice were exposed to HDM as in figure 1. Twenty four hours after the final exposure, animals were sacrificed and lungs collected for A) H&E staining or B) PAS staining. All images are at 10×.
Figure 3.
Airway cytokine dose response to house dust mite exposure.
A) The indicated strains of mice were exposed to HDM as in figure 1, then sacrificed 24 hours after the final exposure and lungs collected for mRNA analysis. Q-RT-PCR analysis was performed for A) IL-4, B) IL-13 and C) IFN-γ. Data were normalized to PBS exposed animals and expressed as fold increased over the PBS controls (which was set at 1). (Note log scales on both y- and x-axes, *p<0.05, Bc vs. B6, †p<0.05, Bc vs. Bc Itk−/−, #B6 vs. B6 Itk−/−, @p<0.05, Bc Itk−/− vs. B6 Itk−/−, students t-test).
Figure 4.
Airway chemokine dose response to house dust mite exposure.
A) The indicated strains of mice were exposed to HDM as in figure 1, then sacrificed 24 hours after the final exposure and lungs collected for mRNA analysis. Q-RT-PCR analysis was performed for A) CCL-7, B) CCL-11 and Data were normalized to PBS exposed animals and expressed as fold increased over the PBS controls as in figure 1. (Note log scales on both y- and x-axes, *p<0.05, Bc vs. B6, †p<0.05, Bc vs. Bc Itk−/−, #B6 vs. B6 Itk−/−, @p<0.05, Bc Itk−/− vs. B6 Itk−/−, students t-test).
Figure 5.
Itk regulates the development of susceptibility or resistance to allergic airway inflammation dependent on background.
A) Model of data analysis. Strain 1 and 2 differ with regards to their dose response to allergen. This difference can be captured by the slope of their dose response (Left panel), the significance of which can inform on whether there is a response (p<0.05) or not (p>0.05) (right panel). These p values can be used as described in the text to determine resistance (R), i.e. no response (with p>0.05,) or the degrees of susceptibility (with p<0.05), which is related to the slope of their response, and thus the p values of those slopes. B) Analysis of the slopes of the IL-4, IL-13 and IFN-γ (left panel) or CCL-7 and CCL-11 (middle panel) dose responses for the indicated mice. Analysis of the inverse of the slopes of the data shown in (A) (right panel, note log scales on the y-axis). C) Analysis of Susceptible (Saai) or Resistance (Raai) as described in thematerials and methods . The dotted line indicates the cutoff of 0.77 between resistance and susceptibility. D) Hierarchical clustering of the values in (C) to generate a cluster tree (B6 = C57BL/6; Bc = BALB/c; B6/Itk−/− = C57BL/6/Itk−/−; Bc/Itk−/− = BALB/c/Itk−/−).