Impact of the Junction Adhesion Molecule-A on Asthma

Abstract

Purpose: Junctional adhesion molecule (JAM)-A is an immunoglobulin-like molecule that colocalizes with tight junctions (TJs) in the endothelium and epithelium. It is also found in blood leukocytes and platelets. The biological significance of JAM-A in asthma, as well as its clinical potential as a therapeutic target, are not well understood. The aim of this study was to elucidate the role of JAM-A in a mouse model of asthma, and to determine blood levels of JAM-A in asthmatic patients.

Materials and methods: Mice sensitized and challenged with ovalbumin (OVA) or saline were used to investigate the role of JAM-A in the pathogenesis of bronchial asthma. In addition, JAM-A levels were measured in the plasma of asthmatic patients and healthy controls. The relationships between JAM-A and clinical variables in patients with asthma were also examined.

Results: Plasma JAM-A levels were higher in asthma patients (n=19) than in healthy controls (n=12). In asthma patients, the JAM-A levels correlated with forced expiratory volume in 1 second (FEV₁%), FEV₁/forced vital capacity (FVC), and the blood lymphocyte proportion. JAM-A, phospho-JNK, and phospho-ERK protein expressions in lung tissue were significantly higher in OVA/OVA mice than in control mice. In human bronchial epithelial cells treated with house dust mite extracts for 4 h, 8 h, and 24 h, the JAM-A, phospho-JNK, and phospho-ERK expressions were increased, as shown by Western blotting, while the transepithelial electrical resistance was reduced.

Conclusion: These results suggest that JAM-A is involved in the pathogenesis of asthma, and may be a marker for asthma.

Keywords: JAM-A; airway hyperresponsiveness; bronchial asthma; cell barrier; inflammation.

Fig. 1. JAM-A levels in healthy controls and patients with stable and exacerbated asthma. Kruskal-Wallis test with Dunn’s post hoc test for multiple comparisons; ^*p<0.05 vs. healthy control. JAM, junctional adhesion molecule.

Fig. 2. (A-E) Relationships of JAM-A with FVC%, FEV₁%, FEV₁/FVC, blood lymphocyte proportion, and blood eosinophil proportion in patients with stable and exacerbated asthma. Pearson or spearman correlation coefficients analysis. (F) JAM-A level between patients with skin test positive and negative in asthmatic patients. (G) JAM-A level between smoker and non-smoker in asthmatic patients. FVC, forced vital capacity; FEV₁, forced expiratory volume in 1 second; JAM, junctional adhesion molecule.

Fig. 3. Airway hyperresponsiveness (AHR) and inflammation in OVA/OVA-sensitized/challenged mice. (A) AHR was increased in OVA/OVA-sensitized/challenged mice compared to control mice. Values are mean±SEM (n=6 mice/group). Two-way repeated measures ANOVA with Tukey’s post hoc test for multiple comparisons; ^*p<0.05 vs. control. (B) Total and differential cell count in mouse bronchoalveolar lavage fluid (BALF). BALF was collected on day 25 post-challenge. Both the total and differential cell counts were higher in OVA/OVA-sensitized/challenged than control mice. One-tailed Student’s t-test; ^*p<0.05 vs. control. (C) Lung histology. Upper panel: more inflammatory cell infiltrations are seen in the lung tissue of OVA/OVA-sensitized/challenged than control mice, as shown by hematoxylin-and-eosin staining. Lower panel: PAS staining of airways reveals an increase in goblet cells in OVA/OVA-sensitized/challenged compared to control mice. (D) Comparison of histologic changes in the lung between OVA/OVA-sensitized/challenged and control mice, based on goblet cell numbers. One-tailed Student’s t-test; ^*p<0.05 vs. control. OVA, ovalbumin; SEM, standard error of the mean; PAS, Periodic Acid-Schiff.

Fig. 4. Levels of JAM-A protein in the lungs of OVA/OVA-sensitized/challenged mice, as determined by western blotting and immunohistochemistry (IHC). (A) JAM-A expression in lung tissue is significantly higher in OVA/OVA-sensitized/challenged than control mice. Three immunoblots were analyzed by densitometry and the densities of the bands were normalized to that of β-actin. (B) Bar graphs show the densitometric data (means±SE). One-tailed Student’s t-test; ^*p<0.05 vs. control. (C) phospho-JNK and phospho-ERK protein levels are higher in OVA/OVA-sensitized/challenged than control mice. Three immunoblots were analyzed by densitometry and the results were then normalized to those of unphosphorylated JNK and ERK. (D and E) Bar graphs show the densitometric data (means±SE). One-tailed Student’s t-test; ^*p<0.05 vs. control. (F) JAM-A levels in lung tissue are higher in OVA/OVA-sensitized/challenged than control mice, as shown by IHC analysis. (G) Bar graphs show the densitometric data. One-tailed Student’s t-test; ^*p<0.05 vs. control. JAM, junctional adhesion molecule; OVA, ovalbumin.

Fig. 5. Normal human bronchial epithelial (NHBE) cells treated with house dust mite (HDM) extract. (A) JAM-A protein levels are increased in HDM-treated NHBE cells compared to the control at 4 h. (B) Bar graphs show the densitometric data (mean±SE). One-tailed Student’s t-test; ^*p<0.05 vs. control. (C) phospho-JNK and phospho-ERK protein levels are higher in extracts from HDM-treated than control NHBE cells at 4 h. Three immunoblots were analyzed by densitometry and the results were normalized to the densities of unphosphorylated JNK and ERK. (D and E) Bar graphs show the densitometric data (mean±SE). One-tailed Student’s t-test; ^*p<0.05 vs. control. (F) Changes in transepithelial electrical resistance (TEER) in NHBE cells treated with HDM extracts, as determined in three immunoblots. Two-way repeated measures ANOVA with Tukey’s post hoc test for multiple comparisons; ^*p<0.05 vs. control. JAM, junctional adhesion molecule.