OBJECTIVE: To investigate the changes in percentage of GATA3⁺ regulatory T (Treg) cells in patients with allergic rhinitis (AR) and mouse models. METHODS: The nasal mucosa specimens were obtained from 6 AR patients and 6 control patients for detection of nasal mucosal inflammation. Peripheral blood mononuclear cells (PBMC) were collected from 12 AP patients and 12 control patients to determine the percentages of Treg cells and GATA3⁺ Treg cells. In a C57BL/6 mouse model of AR, the AR symptom score, peripheral blood OVA-sIgE level, and nasal mucosal inflammation were assessed, and the spleen of mice was collected for detecting the percentages of Treg cells and GATA3⁺ Treg cells and the expressions of Th2 cytokines. RESULTS: Compared with the control patients, AR patients showed significantly increased eosinophil infiltration and goblet cell proliferation in the nasal mucosa (P < 0.01) and decreased percentages of Treg cells and GATA3⁺ Treg cells (P < 0.05). The mouse models of AR also had more obvious allergic symptoms, significantly increased OVA-sIgE level in peripheral blood, eosinophil infiltration and goblet cell hyperplasia (P < 0.01), markedly lowered percentages of Treg cells and GATA3⁺ Treg cells in the spleen (P < 0.01), and increased expressions of IL-4, IL-6 and IL-10 (P < 0.05). CONCLUSION The percentage of GATA3⁺ Treg cells is decreased in AR patients and mouse models. GATA3⁺ Treg cells possibly participate in Th2 cell immune response, both of which are involved in the occurrence and progression of AR, suggesting the potential of GATA3⁺ Treg cells as a new therapeutic target for AR.
Animals ; Mice ; Cytokines/metabolism* ; Disease Models, Animal ; GATA3 Transcription Factor ; Inflammation ; Leukocytes, Mononuclear/metabolism* ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Nasal Mucosa/metabolism* ; Ovalbumin ; Rhinitis, Allergic/therapy* ; T-Lymphocytes, Regulatory ; Th2 Cells/metabolism* ; Humans

Objective:To investigate the expression level and regulatory mechanism of 15-hydroxyprostaglandin dehydrogenase（HPGD） in chronic rhinosinusitis with nasal polyps（CRSwNP）. Methods:The expression pattern and level of HPGD in CRSwNP and control was observed using immunofluorescence, and western blot was used for analysis of HPGD expression in nasal polyp tissues. The effect of recombinant human high mobility group box-1（HMGB1） on HPGD expression in primary human nasal epithelial cells was observed, and the potential blocking effect of RAGE neutralizing antibody on HMGB1-induced HPGD expression was investigated. Results:The expression of HPGD was elevated in CRSwNP patients compared to the control, while the protein mainly localized at CD68-positive cells and epithelial cells. Recombinant human HMGB1 stimulated an increase in HPGD expression in primary human nasal mucosal epithelial cells at a time-dependent manner. Additionally, increased phosphorylation levels of MEK and elevated RAGE expression were also observed at 12 hours, but decreased at 24 hours after the incubation of HMGB1. The increase in the expression of HPGD induced by HMGB1 in primary human nasal epithelial cells was partly inhibited with RAGE neutralizing antibody. Conclusion:Elevated HPGD expression is observed in CRSwNP, predominantly in macrophages and epithelial cells. HMGB1 regulates HPGD expression through the RAGE-MEK signaling pathway, potentially providing a new target for future regulation of PGE2levels in CRSwNP.
Humans ; Antibodies, Neutralizing/metabolism* ; Chronic Disease ; HMGB1 Protein/metabolism* ; Mitogen-Activated Protein Kinase Kinases/metabolism* ; Nasal Mucosa/metabolism* ; Nasal Polyps/metabolism* ; Rhinitis

Objective: Transcriptome sequencing and bioinformatics analysis were performed on the gene expression of nasal epithelial cells in patients with seasonal allergic rhinitis (AR) and perennial AR, so as to obtain the differences in the gene expression of nasal epithelial cells between seasonal AR and perennial AR. Methods: The human nasal epithelial cell line(HNEpC) was cultured in vitro, treated with 100 μg/ml mugwort or house dust mite (HDM) extracts for 24 hours. Total cell RNA was extracted, and quantitative real-time polymerase chain reaction (qPCR) was used to detect the expression of cytokines, including IL-6, IL-8, IL-33 and thymic stromal lymphopoietin (TSLP). From November 2019 to November 2020, 3 seasonal AR patients, 3 perennial AR patients, and 3 healthy controls who attended the Department of Otolaryngology Head and Neck Surgery, China-Japan Union Hospital of Jilin University were analyzed. The patients' primary nasal epithelial cells were cultured in vitro, treated with corresponding allergens for 24 hours. Total RNA was extracted for transcriptome sequencing, and the sequencing results were analyzed by bioinformatics. Results: The qPCR results showed that the cytokines IL-6, IL-8, IL-33 and TSLP of HNEpC treated with mugworts extracts and HDM extracts had the same trend of change. After the nasal epithelial cells from patients with seasonal AR and perennial AR were treated with corresponding allergens, there were differences in biological processes and signal pathways between those and control. Gene ontology (GO) enrichment analysis showed that the differentially expressed genes (DEG) in AR patients allergic to mugwort were mainly enriched in the oxidation-reduction process, the negative regulation of apoptosis process, and the cell adhesion; the DEG in AR patients allergic to HDM were mainly enriched in cell adhesion, the negative regulation of cell proliferation and the response to drug. Enrichment analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway showed that the DEG of AR patients allergic to mugwort were significantly enriched in arachidonic acid metabolism, p53 signaling pathway and transforming growth factor β (TGF-β) signaling pathway, while the DEG of AR patients allergic to HDM were mainly enriched in cells cycle, Fanconi anemia pathway and DNA replication. Gene Set Enrichment Analysis (GSEA) showed that the inflammatory response, TNF-α/NF-κB signaling pathway and IL-2/STAT5 signaling pathway were significantly up-regulated in AR patients allergic to mugwort, indicating the promotion of inflammatory response; and AR patients allergic to HDM had significant down-regulation of G2M, E2F, and MYC, indicating the inhibition of cell proliferation. The protein-protein interaction network showed that TNF and CDK1 were the most interacting proteins in mugwort and HDM allergic AR patients, respectively. Conclusion: Seasonal AR and perennial AR may affect the different biological processes and signal pathways of nasal epithelial cells, leading to differences in the occurrence and development of AR.
Allergens ; Animals ; Computational Biology ; Cytokines/metabolism* ; Epithelial Cells/metabolism* ; Gene Expression ; Humans ; Interleukin-33/metabolism* ; Interleukin-6/metabolism* ; Interleukin-8 ; Nasal Mucosa/metabolism* ; Plant Extracts/metabolism* ; Pyroglyphidae ; RNA/metabolism* ; Rhinitis, Allergic/metabolism* ; Rhinitis, Allergic, Perennial ; Rhinitis, Allergic, Seasonal ; Seasons

Objective: To investigate the effects of dopamine on olfactory function and inflammatory injury of olfactory bulb in mice with allergic rhinitis (AR). Methods: AR mouse model was established by using ovalbumin (OVA), and the mice were divided into two groups: olfactory dysfunction (OD) group and without OD group through buried food pellet test (BFPT). The OD mice were randomly divided into 2 groups, and OVA combined with dopamine (3, 6, 9 and 12 days, respectively) or OVA combined with an equal amount of PBS (the same treatment time) was administered nasally. The olfactory function of mice was evaluated by BFPT. The number of eosinophils and goblet cells in the nasal mucosa were detected by HE and PAS staining. Western blotting, immunohistochemistry or immunofluorescence were used to detect the expression of olfactory marker protein (OMP) in olfactory epithelium, the important rate-limiting enzyme tyrosine hydroxylase (TH) of dopamine, and the marker proteins glial fibrillary acidic protein (GFAP) and CD11b of glial cell in the olfactory bulb. TUNEL staining was used to detect the damage of the olfactory bulb. SPSS 26.0 software was used for statistical analysis. Results: AR mice with OD had AR pathological characteristics. Compared with AR mice without OD, the expression of OMP in olfactory epithelium of AR mice with OD was reduced (F=26.09, P<0.05), the expression of GFAP and CD11b in the olfactory bulb was increased (F value was 38.95 and 71.71, respectively, both P<0.05), and the expression of TH in the olfactory bulb was decreased (F=77.00, P<0.05). Nasal administration of dopamine could shorten the time of food globule detection in mice to a certain extent, down-regulate the expression of GFAP and CD11b in the olfactory bulb (F value was 6.55 and 46.11, respectively, both P<0.05), and reduce the number of apoptotic cells in the olfactory bulb (F=25.64, P<0.05). But dopamine had no significant effect on the number of eosinophils and goblet cells in nasal mucosa (F value was 36.26 and 19.38, respectively, both P>0.05), and had no significant effect on the expression of OMP in the olfactory epithelium (F=55.27, P>0.05). Conclusion: Dopamine can improve olfactory function in mice with AR to a certain extent, possibly because of inhibiting the activation of glial cells in olfactory bulb and reducing the apoptotic injury of olfactory bulb cells.
Animals ; Disease Models, Animal ; Dopamine ; Humans ; Mice ; Mice, Inbred BALB C ; Nasal Mucosa/metabolism* ; Olfactory Bulb/pathology* ; Ovalbumin ; Rhinitis, Allergic/metabolism*

OBJECTIVETo detect the expression of epithelial-to-mesenchymal transition (EMT) biomarkers in nasal polyposis (NP) and to determine the effect of transforming growth factor β1 (TGF-β1) on EMT in cultured nasal epithelial cells.

METHODSThe specimens were obtained from sinus mucosa of 10 NP patients and inferior turbinate mucosa of 10 nasal septum deviation patients. The difference of mRNA expression of E-cadherin, β-catenin , zonula occludens 1 (ZO-1), vimentin and α-smooth muscle actin (α-SMA) in tissue and cultured nasal epithelial cells was detected by real-time PCR. The difference of protein exprssion of E-cadherin and vimentin in cultured nasal epithelial cells was detected by Western blot.SPSS 16.0 software was used to analyze the data.

RESULTSThe relative expression of E-cadherin and ZO-1 in NP tissues (0.012±0.007; 0.006±0.003) was higher than in normal nasal mucosa (0.041±0.024; 0.011±0.005), the difference was significant (t=3.675, P<0.01; t=2.956, P<0.05). However, there was no significant difference in the relative expression of β-catenin, vimentin and α-SMA between two groups (t value was 0.990, 0.429, 0.326, all P>0.05). In cultured nasal epithelial cells both from two groups, TGF-β1 induced the decreased E-cadherin, ZO-1 (tcontrol value was 3.639, 3.430, both P<0.05; tNP value was 3.279, 2.864, both P<0.05) and increased α-SMA, vimentin mRNA expression (tcontrol value was -6.393, -3.085, all P<0.05; tNP value was -2.981, -3.087, both P<0.05). Also, TGF-β1 induced the decreased E-cadherin and increased vimentin protein expression (tcontrol value was 3.583, -3.844, both P<0.05; tNP value was 5.113, -3.642, both P<0.05).

CONCLUSIONEMT is likely to contribute to nasal polyposis and TGF-β1 is involved in this process.

Actins ; metabolism ; Cadherins ; metabolism ; Cells, Cultured ; Epithelial-Mesenchymal Transition ; Humans ; Nasal Mucosa ; metabolism ; Nasal Polyps ; metabolism ; RNA, Messenger ; metabolism ; Real-Time Polymerase Chain Reaction ; Transforming Growth Factor beta1 ; pharmacology ; Vimentin ; metabolism ; Zonula Occludens-1 Protein ; metabolism ; beta Catenin ; metabolism

OBJECTIVEThe aim of the study was to investigate whether Orai1 antibody intraperitoneal injection could improve the condition of allergic rhinitis (AR) in mice.

METHODSTwenty-four BALB/C mice (SPF grade) were classified into 4 groups (AR group, Control group, Experimental group 1 and experimental group 2) according to a random number table. A mouse model of AR was established (Control group was established by phosphate buffered solution), and experimental group 1 and Experimental group 2 were established through intraperitoneal injection of 100 μg and 150 μg Orai1 antibody respectively. The number of sneezing and rubbing and eosinophilia in mice were assessed after different doses of Orai1 antibody intraperitoneal injection were applied. Then Orai1 protein and its mRNA in nasal mucosa, histomine, eosionphil cation protein (ECP), interlukin (IL)-1β, IL-4, IL-5 and IL-6 and their mRNA in nasal lavage fluid (NLF) and nasal mucosa were evaluated using enzyme linked immunosorbent assay (ELISA) and real-time reverse transcription-polymerase chain reaction (real-time RT-PCR). Furthermore, Orai1 protein and its mRNA in Th2 cells in peripheral blood, IL-4 and IL-5 in peripheral serum and their mRNAs in Th2 cells were also examined through ELISA and real-time RT-PCR. The data were analyzed by a statistical software of Graph Pad Prism 5.

RESULTSThere were significant differences in sneezing, nasal rubbing and local invading eosinophils in nasal mucosa after the treatment (t100 μg=7.88, t100 μg=9.92, t100 μg=4.30, respectively; t150 μg=16.43, t150 μg=16.31, t150 μg=9.35, respectively, all P-values<0.01). The Orai1 antibody intervention decreased contents of Orai1 in nasal mucosa, histomine, ECP, IL-1β, IL-4, IL-5 and IL-6. The contents of experimental group 1 were (0.186±0.015) μg/ml, (6.618±0.180) ng/ml, (2.555±0.031) ng/ml, (85.26±2.94) pg/ml, (55.12±1.21) pg/ml, (58.45±2.11) pg/ml and (77.12±2.13) pg/ml, respectively. The contents of experimental group 2 were (0.089±0.003) μg/ml, (4.501±0.310) ng/ml, (1.260±0.017) ng/ml, (48.49±2.12) pg/ml, (33.15±0.87) pg/ml, (38.24±0.95) pg/ml and (51.72±0.81) pg/ml, respectively. The differences were siginificant between group 1, group 2 and AR group(t value was 3.29, 10.44, 9.45, 17.53, 74.53, 87.06, 3.98; 8.54, 13.32, 23.00, 20.89, 80.73, 103.70, 13.34, all P<0.01). However, there were no significant differences in Orai1 protein and its mRNA in peripheral Th2 cells, IL-4 and IL-5 in peripheral serum and their mRNAs in Th2 cells (all P-values>0.05). In addition, the effect of 150 μg Orai1 antibody treatment was better than 100 μg one (all P-values<0.05).

CONCLUSIONOrai1 antibody intraperitoneal injection can improve the symptoms of AR mice, and alleviate the condition of allergic inflammation. Orai1 may become a novel aim in the AR study.

Animals ; Antibodies ; pharmacology ; Calcium Channels ; immunology ; metabolism ; Cytokines ; immunology ; Disease Models, Animal ; Enzyme-Linked Immunosorbent Assay ; Eosinophilia ; therapy ; Eosinophils ; immunology ; Immunotherapy ; Inflammation ; Injections, Intraperitoneal ; Mice ; Mice, Inbred BALB C ; Nasal Mucosa ; metabolism ; ORAI1 Protein ; RNA, Messenger ; Rhinitis, Allergic ; therapy ; Th2 Cells ; immunology

OBJECTIVE: To study the role of JNK (c-Jun N-terminal kinase) signal transduction pathway on the nasal mucosa remodeling in allergic rhinitis rats, to explore whether IL-1β participates the nasal mucosa remodeling in allergic rhinitis by JNK signal transduction pathway. METHOD: Totally 60 male Wistar rats (weighing about 200-250 g)were randomly divided into A (AR group) and B group (control group). The rats in A group were sensitized for inducing AR by intraperitoneal injection ovalbumin and Al(OH)₃. Ovalbumin was respectively dropped in each nasal cavity of every rat for 4,8,12 weeks(A4,A8,or A12 group) each had 10 rats. The rats in B group were sensitized by intraperitoneal injection saline. Saline was respectively dropped in each nasal cavity of every rat for 4,8, 12 weeks(B4, B8, or B12 group), and each had 10 rats. The concentration of IL-1β in serum and nasal lavage fluid were tested by ELASA. The protein expressions of P-JNK and P-c-Jun were detected by immunohistochemical technique. Linear correlation analysis showed the correlation between levels of IL-1β in serum and P-JNK protein, levels of IL-1β in nasal lavage fluid and P-JNK protein. RESULT: The concentrations of IL-1β in serum and nasal lavage fluid of A group were all significantly higher than those of the corresponding B group (all P < 0.01). Compared with A4 group and A8 group, concentrations of IL-1β in nasal lavage fluid of A12 group were significantly increased (all P < 0.01). However the levels of IL-1β in serum were not significantly different among them (all P > 0.05). Mean absorbance values of P-JNK and P-c-Jun in A group were significantly higher than those in corresponding B group (all P < 0.01) and compared with A4 group and A8 group, those of A12 group were significantly increased (all P < 0.01). Strong positive correlation were found between P-JNK and concentration of IL-1β in serum or nasal lavage fluid (r = 0.835 and r = 0.902, all P < 0.01). CONCLUSION JNK signal transduction pathway plays important role in the nasal mucosa remodeling in allergic rhinitis rats. IL-1β participates in AR nasal mucosa remodeling possibly partly through activating JNK signal transduction pathway.
Animals ; Disease Models, Animal ; Interleukin-1beta ; metabolism ; JNK Mitogen-Activated Protein Kinases ; metabolism ; MAP Kinase Signaling System ; Male ; Nasal Mucosa ; pathology ; Ovalbumin ; Paranasal Sinuses ; Rats ; Rats, Wistar ; Rhinitis, Allergic ; metabolism ; pathology ; Signal Transduction

OBJECTIVE: To explore the expression and significance of vasoactive intestinal peptide and Pituitary adenylate cyclase activiting polypeptide (VIP/PACAP) of nasal mucosa in rats with allergic rhinitis (AR), and the function of botulinum toxin-A(BTX-A) to inhibit the expression of VIP/PACAP in AR. METHOD: Thirty Sprague-Dawley rats were randomly divided into 3 groups, which were the AR group, the intervention group, and the control group. In the AR group, ovalbumin was used to sensitize healthy rats. In the intervention group, BTX-A was dripped into the nasal cavity of AR rats 7 times. In the control group, only physiological saline was used to drip into the nasal cavity of AR rats. Changes of the rats' behavior were observed. ELISA were used to detected the concentration variation of serum IFN-γ and IL-4. Histopathology and immunohistochemistry were employed to observe morphology in the rats' nasal mucosal and the expression of VIP/PACAP. Statistical analysis was also made. RESULT: (1)The typical symptoms marks of nasal scratching, sneezing, nasal blockage and rhinorrhea of AR group (7.5 ± 0.50) were higher than intervention group (1 ± 0.27) and control group (0.8 ± 0.31). (2) Comparing to intervention group and control group, the serm IFN-γ of the AR group obvious reduced (P < 0.05), the serm IL-4 of the AR group obvious rose (P < 0.01), and the serm Th1/Th2 (IFN-γ/IL-4) of the AR group obvious reduced (P < 0.01). (3) Comparing to intervention group and control group, the cilium loss, inflammatory cells infiltration, and inflammatory cells exudation of nasal mucosa in AR group were more obviously (P < 0.01), and the intervention group of the 3 indexes was obviously than control group. (4) The expression of VIP in the rats' nasal mucosa of the AR group (13.27 ± 2.74) were more intense than intervention group (5.21 ± 2.18) and control group (3.56 ± 5.30) (P < 0.01), and the expression of PACAP in the rats' nasal mucosa of the AR group (20.97 ± 2.14) were more intense than intervention group (6.33 ± 3.04) and control group (4.63 ± 1.25) (P < 0.01). (5) In all the 3 groups, there was positive correlation between expression of negative in VIP/PACAP and Thl/Th2 cell infiltration(r were respectively -0.340 and -0.223, P < 0.05). CONCLUSION The VIP/PACAP in the rats' nasal mucosa may play an important role in pathogenesis of AR, and BTX-A could improve the symptoms of AR through inhibition of the expression of VIP/ PACAP.
Animals ; Botulinum Toxins, Type A ; pharmacology ; Disease Models, Animal ; Interferon-gamma ; blood ; Interleukin-4 ; blood ; Nasal Mucosa ; drug effects ; metabolism ; Ovalbumin ; Paranasal Sinuses ; Pituitary Adenylate Cyclase-Activating Polypeptide ; antagonists & inhibitors ; metabolism ; Rats ; Rats, Sprague-Dawley ; Rhinitis, Allergic ; drug therapy ; Vasoactive Intestinal Peptide ; antagonists & inhibitors ; metabolism

PURPOSE: The nasal mucosa is the first site to encounter pathogens, and it forms continuous barriers to various stimuli. This barrier function is very important in the innate defense mechanism. Additionally, inflammation of the nasal sinus is known to be a hypoxic condition. Here, we studied the effect of hypoxia on barrier function in normal human nasal epithelial (NHNE) cells. MATERIALS AND METHODS: The expression levels of various junction complex proteins were assessed in hypoxia-stimulated NHNE cells and human nasal mucosal tissues. We performed real-time polymerase chain reaction analysis, western blotting, and immunofluorescence assays to examine differences in the mRNA and protein expression of ZO-1, a tight junction protein, and E-cadherin in NHNE cells. Moreover, we evaluated the trans-epithelial resistance (TER) of NHNE cells under hypoxic conditions to check for changes in permeability. The expression of ZO-1 and E-cadherin was measured in human nasal mucosa samples by western blotting. RESULTS: Hypoxia time-dependently decreased the expression of ZO-1 and E-cadherin at the gene and protein levels. In addition, hypoxia decreased the TER of NHNE cells, which indicates increased permeability. Human nasal mucosa samples, which are supposed to be hypoxic, showed significantly decreased levels of ZO-1 and E-cadherin expression compared with control. CONCLUSION: Our results demonstrate that hypoxia altered the expression of junction complex molecules and increased epithelial permeability in human nasal epithelia. This suggests that hypoxia causes barrier dysfunction. Furthermore, it may be associated with innate immune dysfunction after encountering pathogens.
Anoxia/etiology/*metabolism ; Blotting, Western ; Cadherins/*analysis/genetics ; Epithelium/chemistry/pathology ; Humans ; Membrane Proteins/*analysis ; Nasal Mucosa/*chemistry/pathology/*secretion ; Permeability/*radiation effects ; RNA, Messenger/genetics/metabolism ; Real-Time Polymerase Chain Reaction ; Tight Junctions/*metabolism ; Zonula Occludens-1 Protein

OBJECTIVE: To investigate the expression and role of Interleukin-33 (IL-33) and ST2 in the nasal polyps of human Eosinophilic and non-Eosinophilic chronic rhinosinusitis with nasal polyps (ECRS and non-ECRS). METHOD: IL-33 and ST2 protein expression in nasal polyps of ECRS and non-ECRS as well as in seemingly normal mucosa of the inferior turbinate tissue was investigated by immunohistochemical staining and messenger RNA (mRNA) expression of IL-33 and ST2 was assessed by realtime polymerase chain reaction (PCR) in 27 subjects with ECRS, 33 subjects with non-ECRS, and 11 control subjects. RESULT: (1) The ST2 was found both in nasal polyps of ECRS and non-ECRS,especially in ECRS, yet hardly found in the normal mucosa of the inferior turbinate tissue; (2) The expression of ST2 mRNA in nasal polyps of ECRS was higher than that in non-ECRS and normal inferior turbinate tissue, and the difference was both prominent in statistics (P<0.01); (3) The expression patterns of IL-33 at both mRNA and protein levels were not significantly different among the three groups (P>0.05). CONCLUSION The IL-33 and its receptor ST2 were both expressed in human nasal polyps including ECRS and non-ECRS, meanwhile the expression patterns of ST2 at both mRNA and protein levels were significantly higher in nasal polyps of ECRS. The current study suggests that IL-33 and its receptor ST2 may play important roles in the pathogenesis of chronic rhinosinusitis with nasal polyps, especially in ECRS through the increased expression of ST2 in Eosinophils as a hypothesis.
Chronic Disease ; Eosinophils ; immunology ; Humans ; Interleukin-1 Receptor-Like 1 Protein ; Interleukin-33 ; metabolism ; Nasal Mucosa ; metabolism ; Nasal Polyps ; immunology ; RNA, Messenger ; Real-Time Polymerase Chain Reaction ; Receptors, Cell Surface ; metabolism ; Rhinitis ; immunology ; Sinusitis ; immunology ; Turbinates ; metabolism