Deepti R. Nagarkar

Thymic stromal lymphopoietin activity is increased in nasal polyps of patients with chronic rhinosinusitis

BACKGROUND Chronic rhinosinusitis with nasal polyps (CRSwNP) is associated with TH2-dominant inflammation. Thymic stromal lymphopoietin (TSLP) is a cytokine that triggers dendritic cell-mediated TH2 inflammatory responses and that enhances IL-1-dependent TH2 cytokine production in mast cells. Although increased TSLP mRNA levels have been found in nasal polyps (NPs), expression of TSLP protein and its function in patients with chronic rhinosinusitis (CRS) have not been fully explored. OBJECTIVES The objective of this study was to investigate the role of TSLP in patients with CRS. METHODS We investigated the presence and stability of TSLP protein in NPs using ELISA and Western blotting and investigated the function of TSLP in nasal tissue extracts with a bioassay based on activation of human mast cells. RESULTS Although TSLP mRNA levels were significantly increased in NP tissue from patients with CRSwNP compared with uncinate tissue from patients with CRS or control subjects, TSLP protein was significantly decreased in NP tissue, as detected by using the commercial ELISA kit. We found that recombinant TSLP was time-dependently degraded by NP extracts, and this degradation was completely inhibited by a protease inhibitor cocktail, suggesting that TSLP is sensitive to tissue proteases. Interestingly, NP extract-treated TSLP had higher activity in mast cells, although the amount of full-length TSLP was reduced up to 85%. NP extracts significantly enhanced IL-1β-dependent IL-5 production in mast cells compared with uncinate tissue homogenates, and responses were significantly inhibited by anti-TSLP, suggesting that NPs contain biologically relevant levels of TSLP activity. CONCLUSION TSLP and its metabolic products might play an important role in the inflammation seen in patients with CRSwNP.

Regulation and Function of the IL-1 Family Cytokine IL-1F9 in Human Bronchial Epithelial Cells

The IL-1 family of cytokines, which now includes 11 members, is well known to participate in inflammation. Although the most recently recognized IL-1 family cytokines (IL-1F5-11) have been shown to be expressed in airway epithelial cells, the regulation of their expression and function in the epithelium has not been extensively studied. We investigated the regulation of IL-1F5-11 in primary normal human bronchial epithelial cells. Messenger (m)RNAs for IL-1F6 and IL-1F9, but not IL-1F5, IL-1F8 or IL-1F10, were significantly up-regulated by TNF, IL-1β, IL-17 and the Toll-like receptor (TLR)3 ligand double-stranded (ds)RNA. mRNAs for IL-1F7 and IL-1F11 (IL-33) were weakly up-regulated by some of the cytokines tested. Notably, mRNAs for IL-1F6 and IL-1F9 were synergistically enhanced by the combination of TNF/IL-17 or dsRNA/IL-17. IL-1F9 protein was detected in the supernatant following stimulation with dsRNA or a combination of dsRNA and IL-17. IL-1F6 protein was detected in the cell lysate but was not detected in the supernatant. We screened for the receptor for IL-1F9 and found that lung fibroblasts expressed this receptor. We found that IL-1F9 activated mitogen-activated protein kinases and the transcription factor NF-κB in primary normal human lung fibroblasts. IL-1F9 also stimulated the expression of the neutrophil chemokines IL-8 and CXCL3 and the Th17 chemokine CCL20 in lung fibroblasts. These results suggest that epithelial activation by TLR3 (e.g., by respiratory viral infection) and exposure to cytokines from Th17 cells (IL-17) and inflammatory cells (TNF) may amplify neutrophilic inflammation in the airway via induction of IL-1F9 and activation of fibroblasts.

Airway epithelial cells activate TH2 cytokine production in mast cells through IL-1 and thymic stromal lymphopoietin

BACKGROUND Airway epithelial cells are important regulators of innate and adaptive immunity. Although mast cells are known to play a central role in manifestations of allergic inflammation and are found in the epithelium in patients with T(H)2-related diseases, their role is incompletely understood. OBJECTIVES The objective of this study was to investigate the role of airway epithelial cells in the production of T(H)2 cytokines in mast cells. METHODS Normal human bronchial epithelial (NHBE) cells were stimulated with TNF, IL-4, IFN-γ, IL-17A, and double-stranded RNA (dsRNA) alone or in combination. Human mast cells were stimulated with epithelial cell-derived supernatants or cocultured with NHBE cells. T(H)2 cytokine responses were blocked with neutralizing antibodies. RESULTS Supernatants from IL-4- and dsRNA-stimulated NHBE cells significantly enhanced T(H)2 cytokine production from mast cells. The combination of IL-4 and dsRNA itself or supernatants from NHBE cells stimulated with other cytokines did not activate mast cells, suggesting that mast cell responses were induced by epithelial cell factors that were only induced by IL-4 and dsRNA. Epithelial supernatant-dependent T(H)2 cytokine production in mast cells was suppressed by anti-IL-1 and anti- thymic stromal lymphopoietin (TSLP) and was enhanced by anti-IL-1 receptor antagonist. Similar results were observed in coculture experiments. Finally, we found dsRNA-dependent production of IL-1, TSLP, and IL-1 receptor antagonist in NHBE cells was regulated by T(H) cytokines, and their ratio in NHBE cells correlated with T(H)2 cytokine production in mast cells. CONCLUSIONS Pathogens producing dsRNA, such as respiratory viral infections, might amplify local T(H)2 inflammation in asthmatic patients through the production of TSLP and IL-1 by epithelial cells and subsequent activation of T(H)2 cytokine production by mast cells in the airways.