Alternative Method for Primary Nasal Epithelial Cell Culture Using Intranasal Brushing and Feasibility for the Study of Epithelial Functions in Allergic Rhinitis

Abstract

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.Purpose: Although differentiated normal human nasal epithelial (NHNE) cells can be used to study the role of human nasal epithelium, there is a need for effective culture models of nasal epithelium in sinonasal disease status, including allergic rhinitis (AR). We aimed to examine the feasibility of intranasal brushing for culture of nasal epithelial cells in AR patients and to verify the hypothesis that allergic nasal epithelial (ARNE) cells differ in histologic and physiologic characteristics. Methods: We established a system for isolating (via intranasal brushing) and culturing (with air-liquid interface, ALI) nasal epithelial cells from healthy volunteers (n=8) and AR patients (n=8). We used this system to compare the histologic findings and physiologic characteristics of NHNE and ARNE. Results: The histology results showed that fully differentiated ALI culture was obtained at least 14 days after confluence and that both ciliated and secretory cells were well differentiated in ALI culture using nasal brushing. The histology results of ARNE culture were significantly different from NHNE. The number of ciliated cells was lower, and secretory cells were more dominant in ARNE cell culture compared to NHNE cells. We also observed, by electron microscopy, loose tight junctions and short cilia in cultured ARNE cells. In addition, the mRNA level of TSLP which was one of the epithelial-derived allergic cytokines was significantly higher, and the expressions of genes involved in ciliogenesis were lower in cultured ARNE cells without allergen stimulation. Conclusions: Our findings suggest that ALI culture of ARNE cells using intranasal brushing may be an alternative method for epithelial cell culture in AR patients and that cultured ARNE cells will be useful for in vitro studies of the mechanisms at play during AR because they maintain unique allergic characteristics.

Nasal cytology: Methodology with application to clinical practice and research

“…The brush is immediately placed in a 5 mL polystyrene plastic tube containing 5 mL of PBS and cut‐off just above the bristles. The brush will be shaken vigorously in the solution and carefully brushed off against the wall of the tube . The tubes are centrifuged at 400 g for 10 minutes.…”

Type III interferons are critical host factors that determine susceptibility to Influenza A viral infection in allergic nasal mucosa

“…We found that cultured ARNE cells containing the histologic characteristics of allergic rhinitis nasal epithelium during the process of primary cell culture would be useful to investigate the susceptibility of allergic nasal mucosa to IAV infection. 21 The current findings showed that IAV-infected ARNE cells showed significant differences in viral loads, such as higher mRNA levels in cell lysate and viral titres in supernatant compared to IAV-infected NHNE cells. Furthermore, our clinical data showed that influenza-related symptoms such as fever, cough and throat pain in H1N1-positive individuals diagnosed with AR lasted longer and required hospitalization and intensive care.…”

“…Cells were cultured in a 1:1 mixture of basal epithelial growth medium and DMEM containing previously described supplements. 9,21 Cultures were grown while submerged for the first 9 days. The…”

Histamine H4 receptor gene polymorphisms: a potential predictor of oral H1 antihistamine efficacy for allergic rhinitis

“…From January 2013 to January 2015, a total of 142 AR patients were selected from Shanghai Ninth People's Hospital, Shanghai JiaoTong University School of Medicine as a case group. All the patients completed the following tests and questionnaires based on the World Health Organization (WHO) diagnostic criterion of ARIA 2007: anterior rhinoscopy, allergen skin test, history of allergic rhinitis, family history of allergy, and history of food and drug allergies . The inclusion criteria were as follows: (1) age from 15 to 60 years; (2) AR confirmed through skin prick or sensitinogen (fungi, mites and food‐allergens like fish, shrimp and crab) screening test; and (3) no antihistamine agent used in the 2 weeks prior to treatment.…”