Fortilin, also known as histamine-releasing factor [HRF] and translationally controlled tumor protein, has been studied as a HRF and implicated as a mediator in late phase allergic reactions. It has a proinflammatory role in murine asthma and skin immediate hypersensitivity. However, the ability of allergen challenge to modulate the levels of fortilin in the nasal secretion of subjects with allergic rhinitis has not been reported. Nine adult subjects with allergic rhinitis and positive skin prick testing to ragweed pollen extract (RWPE) were recruited for this study. The mean age of the subjects was 41.6 years. Intranasal challenge with saline and provoking doses of RWPE solution were performed on two different days. During nasal challenge, symptom scores were recorded at baseline, 30 min, and hourly for a total of 6 hours. Nasal lavage was performed at baseline, 30 min and 5 hours post-challenge, and the collected nasal fluid was analyzed for fortilin levels by ELISA. Compared to saline challenge, RWPE challenge rapidly increased congestion, drainage, sneezing, and total symptom scores at 30 min post-challenge (p<0.05). These scores decreased 2 hours post-challenge. Compared to saline challenge, ragweed extract increased the level of fortilin 5 hours after challenge (p<0.05), but not at 30 min post challenge. Exposure to ragweed pollen induces a delayed secretion of fortilin in the nasal airway of ragweed-IgE skin test positive subjects with allergic rhinitis. Fortilin may play a role in the pathophysiology of allergic rhinitis.

RWPE contains NADPH oxidases that rapidly induce oxidative stress in the airways that is critical for allergic inflammation in sensitized mice. However response to oxidative stress is also dependent on host-specific factors. To identify these host factors, we performed saline or RWPE challenge in subjects with allergic rhinitis, and quantified 8-OHdG levels and 48 cytokines as markers of innate oxidative stress and host response respectively. Six RWPE allergic subjects were challenged intranasally with saline and RWPE, and symptom scores were recorded. Small strips of filter paper were applied to the nose 30 min post-challenge and stored. Cytokines were recovered from these filter strips, and 48 human cytokines were measured by Bio-Plex cytokine array. Nasal lavage fluid was also obtained 30 min post-challenge, and 8-OHdG in these fluids was measured by ELISA. RWE nasal challenge increased the patient symptom score at 30 min post-challenge (2.3 ± 0.8 saline, 6.2 ± 0.7 RWPE, p<0.05). In nasal lavage fluid, RWPE challenge increased 8-OHdG levels (0.2 ± 0.1 saline, 0.3 ± 0.1 RWPE, ng/ml, p<0.05). In nasal filter paper eluate, RWPE challenge increased levels of IL-10 (40 ± 40 saline, 72 ± 39 RWPE, pg/ml, p<0.05) and G-CSF (61 ± 39 saline, 154 ± 105 RWPE pg/ml, p<0.05). RWPE nasal challenge in allergic rhinitis patients increased symptoms, and levels of 8-OHdG, IL-10 and G-CSF. These observations suggest that RWPE rapidly induces ROS and DNA damage, but simultaneously increases anti-inflammatory cytokines IL-10 and G-CSF that may suppress host response to allergens.

RATIONALE: Intrinsic NADPH oxidases in RWE induce ROS in airway epithelium of mice and augment RWE-induced allergic airway inflammation. These effects are reversed by co-administering N-acetyl cysteine (NAC) antioxidant. To extend these observations to humans, we performed proteomic analysis of NLF following RWE or RWE + NAC challenge. METHODS: Eight RWE allergic subjects were challenged intranasally with saline and RWE, and symptom scores were recorded. We first examined whether proteins can be identified in NLF by 2-D gels. Subject 1 with only early phase symptoms was challenge with RWE, and NLF was obtained 6 h post-challenge. Protein spots in NLF were localized in 2-D gels, analyzed by Progenesis Samespots®, and identified by MALDI-TOF. Only one patient, Subject 2, developed early and late phase symptoms to RWE. The subject was challenged with saline, RWE, or RWE + NAC. NLF was obtained at baseline, 30 min, 5 h and 6 h post-challenge, and all 12 NLFs were subjected to 2-D gel analysis. RESULTS: We identified four proteins in NLF of Subject 1, lysozyme, lipocalin-1 precursor, cystatin-S and mammaglobin-B. In Subject 2, RWE challenge upregulated three proteins, A (12 kDa, pI 4.0), B (37 KDa, pI 8.5), and C (14.2 kDa, pI 5.6), downregulated two proteins, D (34.8 kDa, pI 6.4) and E (14.8 kDa, pI 9.1). Adding NAC with RWE challenge blocked modulation of these five proteins. CONCLUSIONS: We have demonstrated that nasal allergen challenge with associated oxidative stress dramatically alters secretion of five proteins in the nasal fluid. Identification of these proteins may provide valuable insight into pathogensis of allergic rhinitis.