Nasal Sensitization with Ragweed Pollen Induces Local-Allergic-Rhinitis-Like Symptoms in Mice

<div><p>Recently, the concept of local allergic rhinitis (LAR) was established, namely rhinitis symptoms with local IgE production and negative serum antigen-specific IgE. However, the natural course of LAR development and the disease pathogenesis is poorly understood. This study investigated the pathophysiology of mice with allergic rhinitis that initially sensitized with ragweed pollen through the nasal route. Mice were nasally administrated ragweed pollen over consecutive days without prior systemic immunization of the allergen. Serial nasal sensitization of ragweed pollen induced an allergen-specific increase in sneezing, eosinophilic infiltration, and the production of local IgE by day 7, but serum antigen-specific IgE was not detected. Th2 cells accumulated in nose and cervical lymph nodes as early as day 3. These symptoms are characteristic of human LAR. Continual nasal exposure of ragweed pollen for 3 weeks resulted in the onset of classical AR with systemic atopy and adversely affected lung inflammation when the allergen was instilled into the lung. <i>Fcer1a</i>^−/− mice were defective in sneezing but developed normal eosinophilic infiltration. Contrary, <i>Rag2</i>^−/− mice were defective in both sneezing and eosinophilic infiltration, suggesting that T cells play a central role in the pathogenesis of the disease. These observations demonstrate nasal allergen sensitization to non-atopic individuals can induce LAR. Because local Th2 cell accumulation is the first sign and Th2 cells have a central role in the disease, a T-cell-based approach may aid the diagnosis and treatment of LAR.</p></div

Repeated nasal ragweed sensitization induces systemic atopy.

<p>Nasal, ragweed (RW)-sensitized mice were analyzed 1 day after final sensitization. (A) Experimental schema. (B) Serum immunoglobulin levels. (C) Production of cytokines in cLN cells. (D–F) Histological examinations. (D) Hematoxylin and eosin (HE) staining of coronal section of nasal cavity. (E) HE staining of nasal lateral mucosa; black square in (D). (F) Periodic acid-Schiff staining of nasal septum; red square in (D). (G) Presence of eosinophils in nasal mucosa. Data representative of three independent experiments (means, SEMs, n = 5). *P<0.05, ***P<0.001. N.D. not detected.</p

Nasal sensitization of ragweed adversely affects allergic inflammation in lungs.

<p>Nasal, ragweed (RW)-sensitized mice were intratracheally challenged with ragweed 3 days after final sensitization. (A) Experimental schema. (B) BALF CD45⁺ cells and eosinophils. Hematoxylin and eosin (C) and Periodic acid-Schiff (D) staining of lungs. Data representative of three independent experiments (means and SEMs, n = 4). *P<0.05, **P<0.01.</p

Local IgE production by nasal ragweed sensitization.

<p>Mice were nasally administered ragweed (RW) pollen on indicated consecutive days. GLTs and PSTs expression in cLN (A) and nasal (B) B cells. (C) qPCR analysis of <i>Aicda</i> expression in cLN and nasal B cells. Data representative of three independent experiments (n = 2). Nasal B cells were pooled from five mice.</p

Acquired immunity is central for pathogenesis of nasally sensitized allergic rhinitis.

<p>WT and <i>Rag2</i>^−/− mice were nasally administered ragweed (RW) pollen or PBS as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103540#pone-0103540-g003" target="_blank">Figure 3A</a>. (A, C) or for 7 consecutive days (B). (A) Number of sneezes. (B, C) Presence of eosinophils in nasal mucosa. Data representative of three independent experiments (means, SEMs n = 4). **P<0.01, ***P<0.001 (in (A), comparing WT to <i>Rag2</i>^−/− mice). ^†P<0.05, ^††P<0.01, ^†††P<0.001 (in (A), comparing to day 0 in <i>Rag2</i>^−/− mice).</p

(A) Complex of indicated poly I:C and protein (MDA5 or RIG-I) was observed by AFM

Height is on a scale from 0 to 4 nm, with a low area depicted in dark brown and a higher area depicted in brighter color. Scale area, 500 nm. Bars, 100 nm. Those are representative images from several pictures. (B and C) Statistical height analyses of molecules corresponding to pictures in A.<p><b>Copyright information:</b></p><p>Taken from "Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid–inducible gene-I and melanoma differentiation–associated gene 5"</p><p></p><p>The Journal of Experimental Medicine 2008;205(7):1601-1610.</p><p>Published online 7 Jul 2008</p><p>PMCID:PMC2442638.</p><p></p

(A and B) Indicated genotype of MEFs were treated with 1 μg/ml of the indicated RNAs (10 μg/ml in the case of high dose) for 24 h

s, sense ssRNA; as, antisense ssRNA; s+as, dsRNA generated by annealing s with as. s and as are chemically synthesized 70-bp ssRNAs having a 5′ hydroxyl end in A. s and as are in vitro transcribed capped-ssRNAs in (B). The production of IFN-β in the supernatant was measured by ELISA. (C) WT, , , and MEFs were treated with 1 μg/ml of in vitro–transcribed capped dsRNAs for 16 h. The production of IFN-β in the supernatant was measured by ELISA. Error bars show SDs between triplicates.<p><b>Copyright information:</b></p><p>Taken from "Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid–inducible gene-I and melanoma differentiation–associated gene 5"</p><p></p><p>The Journal of Experimental Medicine 2008;205(7):1601-1610.</p><p>Published online 7 Jul 2008</p><p>PMCID:PMC2442638.</p><p></p

(A) The indicated RNAs are shown on the ethidium bromide–stained agarose gel

M, DNA marker. (B) WT, , 5, and 5 MEFs were treated with 1 μg/ml of untreated or RNase III–treated poly I:C for 16 h. The production of IFN-β in the supernatant was measured by ELISA. mock, no RNA. (C) Relative induction of IFN-β after stimulation with poly I:C treated with RNase III for the indicated periods. (D) The production level of IFN-β in the stimulation with RNase A–treated, Bal31-treated, or alkaline-hydrolyzed poly I:C. Error bars show the SDs between triplicates.<p><b>Copyright information:</b></p><p>Taken from "Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid–inducible gene-I and melanoma differentiation–associated gene 5"</p><p></p><p>The Journal of Experimental Medicine 2008;205(7):1601-1610.</p><p>Published online 7 Jul 2008</p><p>PMCID:PMC2442638.</p><p></p

(A) RNA samples harvested from uninfected (mock), EMCV-, VSV-, or influenza virus (flu)–infected cells were transfected into WT, , and MEFs

The production of IFN-β in the culture supernatant 10 h after transfection was measured by ELISA. (B) RNA harvested from noninfected (mock) or EMCV-, VSV-, or influenza virus–infected cells with CIAP-, RNase III-, both CIAP-, and RNase III-treatments or nontreatment (enzyme-) was transfected into WT MEFs. The production of IFN-β in the supernatant 10 h after transfection was measured by ELISA. (C) dsRNA in uninfected (mock), EMCV-, VSV-, or influenza virus-infected cells was measured by ELISA. (D) Immunostaining for dsRNA in MEFs infected with EMCV, VSV, and influenza virus for 8 h. Red, dsRNA; blue, nucleus. Error bars show SDs between triplicates. (E) RNA harvested from noninfected (mock) or VSV-infected cells (indicated periods) was electrophoresed in 1.5% agarose gel, transferred to a nylon membrane, and blotted by anti-dsRNA antibody. Reovirus genome RNAs were indicated as the size control. The arrow shows VSV dsRNA. (F) dsRNA blotting of RNA harvested from EMCV- or VSV-infected cells. RNAs were electrophoresed in nondenaturing 10% polyacrylamide gel. Reovirus genome RNAs were indicated (left). Arrows (right) show EMCV and VSV dsRNA.<p><b>Copyright information:</b></p><p>Taken from "Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid–inducible gene-I and melanoma differentiation–associated gene 5"</p><p></p><p>The Journal of Experimental Medicine 2008;205(7):1601-1610.</p><p>Published online 7 Jul 2008</p><p>PMCID:PMC2442638.</p><p></p

(A) WT, , and GM-CSF-DCs were infected with the indicated multiplicity of infection of reovirus

The production of IFN-β in the supernatant was measured by ELISA. (B and C) The indicated genotypes of MEFs were treated with 1 μg/ml of reovirus genome RNA (B) or 0.1 μg/ml of dsRNA segments (C) for 16 h. The production of IFN-β in the supernatant was measured by ELISA. The reoviral genome is shown on the ethidium bromide-stained gel (C, right), and the S (1.2–1.4 kbp), M (2.2–2.3 kbp), and L (3.9 kbp) segments are indicated. Error bars show SDs between triplicates.<p><b>Copyright information:</b></p><p>Taken from "Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid–inducible gene-I and melanoma differentiation–associated gene 5"</p><p></p><p>The Journal of Experimental Medicine 2008;205(7):1601-1610.</p><p>Published online 7 Jul 2008</p><p>PMCID:PMC2442638.</p><p></p