The significant expression of TRPV3 in nasal polyps of eosinophilic chronic rhinosinusitis

BackgroundThe number of patients with eosinophilic chronic rhinosinusitis (ECRS) has been increasing in recent years in Japan. In ECRS, nasal polyps recur immediately after endoscopic sinus surgery. The molecular biological mechanism underlying the refractoriness of ECRS is unclear.MethodsWhole-transcriptome analysis with next-generation sequencing (RNA-seq) was conducted to investigate the molecular biological mechanism of ECRS. Real-time PCR, immunohistochemical staining, and immunofluorescence staining were performed to validate the results of RNA-seq.ResultsRNA-seq analysis revealed that in the nasal polyps of ECRS, the levels of 3 transcripts were elevated significantly and those of 7 transcripts were diminished significantly. Among the genes encoding these transcripts, TRPV3 (transient receptor potential cation channel, subfamily V, member 3) was identified as the only gene that is highly expressed in ECRS nasal polyps but this gene\u27s expression was not previously detected using DNA microarray analysis in peripheral blood eosinophils. TRPV3 is newly identified here as a gene transcribed in ECRS. Our analysis also revealed that TRPV3 was highly expressed in the infiltrating eosinophils and mucosal epithelium of the nasal polyps of ECRS, and further that the more severe the refractoriness was after surgery, the higher the TRPV3 expression was in nasal polyps.ConclusionsTRPV3 might play a role in the refractoriness of ECRS. Additional studies are required to evaluate the function of TRPV3 in ECRS

Periostin as a novel biomarker for postoperative recurrence of chronic rhinosinitis with nasal polyps

We previously reported that chronic rhinosinusitis with nasal polyps (CRSwNP) was subdivided into four chronic rhinosinusitis (CRS) subtypes using the JESREC scoring system. We sought to identify the gene expression profile and biomarkers related with CRSwNP by RNA-sequence. RNA-sequencing was performed to identify differentially expressed genes between nasal polyps (NPs) and inferior turbinate mucosa from 6 patients with CRSwNP, and subsequently, quantitative real-time PCR was performed to verify the results. ELISA was performed to identify possible biomarkers for postoperative recurrence. In the RNA-sequencing results, periostin (POSTN) expression was the highest in NP. We focused on POSTN and investigated the protein level of POSTN by immunohistochemistry and ELISA. POSTN was diffusely expressed in moderate and severe eosinophilic CRS using immunohistochemistry, and its staining pattern was associated with the severity of the phenotype of the CRSwNP (P < 0.05). There was a significant difference between the POSTN high/low groups for postoperative recurrence when the cutoff point was set at 115.5 ng/ml (P = 0.0072). Our data suggests that the protein expression level of POSTN was associated with the severity of CRSwNP, and serum POSTN can be a novel biomarker for postoperative recurrence of CRSwNP

Serum IgG4 as a biomarker reflecting pathophysiology and post-operative recurrence in chronic rhinosinusitis

Background: Type 2 chronic rhinosinusitis (CRS), especially eosinophilic CRS (ECRS), is an intractable upper airway inflammatory disease. Establishment of serum biomarkers reflecting the pathophysiology of CRS is desirable in a clinical setting. As IgG4 production is regulated by type 2 cytokines, we sought to determine whether serum IgG4 levels can be used as a biomarker for CRS. Methods: Association between the serum IgG4 levels and clinicopathological factors was analyzed in 336 CRS patients. Receiver operating characteristics (ROC) analysis was performed to determine the cut-off value of serum IgG4 levels that can be used to predict the post-operative recurrence. Results: Serum IgG4 levels were significantly higher in patients with moderate to severe ECRS versus those with non to mild ECRS. The levels were also significantly higher in asthmatic patients and patients exhibiting recurrence after surgery compared to controls. ROC analysis determined that the best cut-off value for the serum IgG4 level to predict the post-operative recurrence was 95 mg/dL. The corresponding sensitivity and specificity were 39.7% and 80.5%, respectively. When we combined the two cut-off values for the serum IgG4 and periostin, patients with high serum levels of either IgG4 or periostin exhibited a high post-operative recurrence (OR: 3.95) as compared to patients having low serum levels of both IgG4 and periostin. Conclusions: The present results demonstrate that the serum IgG4 level is associated with disease severity and post-operative course in CRS. In particular, the combination of serum IgG4 and periostin could be a novel biomarker that predicts post-operative recurrence

Variants of C-C Motif Chemokine 22 (CCL22) Are Associated with Susceptibility to Atopic Dermatitis: Case-Control Studies

Atopic dermatitis (AD) is a common inflammatory skin disease caused by multiple genetic and environmental factors. AD is characterized by the local infiltration of T helper type 2 (Th2) cells. Recent clinical studies have shown important roles of the Th2 chemokines, CCL22 and CCL17 in the pathogenesis of AD. To investigate whether polymorphisms of the CCL22 gene affect the susceptibility to AD, we conducted association studies and functional studies of the related variants. We first resequenced the CCL22 gene and found a total of 39 SNPs. We selected seven tag SNPs in the CCL22 gene, and conducted association studies using two independent Japanese populations (1st population, 916 cases and 1,032 controls; 2nd population 1,034 cases and 1,004 controls). After the association results were combined by inverse variance method, we observed a significant association at rs4359426 (meta-analysis, combined P = 9.6×10−6; OR, 0.74; 95% CI, 0.65–0.85). Functional analysis revealed that the risk allele of rs4359426 contributed to higher expression levels of CCL22 mRNA. We further examined the allelic differences in the binding of nuclear proteins by electrophoretic mobility shift assay. The signal intensity of the DNA-protein complex derived from the G allele of rs223821, which was in absolute LD with rs4359426, was higher than that from the A allele. Although further functional analyses are needed, it is likely that related variants play a role in susceptibility to AD in a gain-of-function manner. Our findings provide a new insight into the etiology and pathogenesis of AD

ORAI1 Genetic Polymorphisms Associated with the Susceptibility of Atopic Dermatitis in Japanese and Taiwanese Populations

Atopic dermatitis is a chronic inflammatory skin disease. Multiple genetic and environmental factors are thought to be responsible for susceptibility to AD. In this study, we collected 2,478 DNA samples including 209 AD patients and 729 control subjects from Taiwanese population and 513 AD patients and 1027 control subject from Japanese population for sequencing and genotyping ORAI1. A total of 14 genetic variants including 3 novel single-nucleotide polymorphisms (SNPs) in the ORAI1 gene were identified. Our results indicated that a non-synonymous SNP (rs3741596, Ser218Gly) associated with the susceptibility of AD in the Japanese population but not in the Taiwanese population. However, there is another SNP of ORAI1 (rs3741595) associated with the risk of AD in the Taiwanese population but not in the Japanese population. Taken together, our results indicated that genetic polymorphisms of ORAI1 are very likely to be involved in the susceptibility of AD

Trimellitic anhydride induces low-grade mast cell degranulation without specific IgE

Objectives: Low-molecular-weight (LMW) substances are known to be causative agents of occupational asthma (OA) and occupational rhinitis (OR). Although most LMW substances are irritants or allergens, some can cause immediate type immunoglobulin E (IgE)-mediated allergic reactions. Trimellitic anhydride (TMA) is one such LMW substance, which is known as an immunological sensitizer. However, the exact molecular biological details of the effects of TMA remain unclear. Methods: We measured the β-hexosaminidase release from mast cells after directly exposing the cells to various LMW substances. The tyrosine phosphorylation of whole cellular molecules and the phosphorylation of extracellular signal-regulated kinase (ERK) were assessed by immunoblot assay. Results: Among the LMW substances tested, only TMA induced β-hexosaminidase release. However, the mast cell degranulation induced by TMA was lower than that induced by an antigen or a calcium ionophore. Moreover, the pattern of tyrosine phosphorylation of whole cellular molecules was quite different between IgE-mediated antigen stimulation and TMA exposure. The TMA effect on mast cells was independent of not only IgE but also Ca2+ influx. ERK phosphorylation was not detected in mast cells exposed to TMA. Conclusions: TMA induced mild degranulation of mast cells without IgE, even though the phosphorylation of ERK was not detected. This reaction suggests that TMA affects humans even upon first exposure. Therefore, it is imperative to avoid human exposure to high concentrations of TMA. In order to stop the development of severe asthma in individuals with OR, we need to be able to identify cases of OR caused by TMA as soon as possible. Keywords: Occupational rhinitis, Respiratory hypersensitivity, Trimellitic anhydride, Mast cel

Integrative RNA-Seq and H3 Trimethylation ChIP-Seq Analysis of Human Lung Cancer Cells Isolated by Laser-Microdissection

Our previous integrative study in gastric cancer discovered cryptic promoter activation events that drive the expression of important developmental genes. However, it was unclear if such cancer-associated epigenetic changes occurred in cancer cells or other cell types in bulk tissue samples. An integrative analysis consisting of RNA-Seq and H3K4me3 ChIP-Seq was used. This workflow was applied to a set of matched normal lung tissues and non-small cell lung cancer (NSCLC) tissues, for which the stroma and tumor cell parts could be isolated by laser-microdissection microscopy (LMD). RNA-Seq analysis showed subtype-specific differential expressed genes and enriched pathways in NSCLC. ChIP-Seq analysis results suggested that the proximal altered H3K4me3 regions were located at differentially expressed genes involved in cancer-related pathways, while altered distal H3K4me3 regions were annotated with enhancer activity of cancer regulatory genes. Interestingly, integration with ENCODE data revealed that proximal tumor-gained promoters were associated with EZH2 and SUZ12 occupancies, which are the core components of polycomb repressive complex 2 (PRC2). This study used LMD on clinical samples for an integrative analysis to overcome the tissue heterogeneity problem in cancer research. The results also contribute to the overall understanding of genetic and epigenetic dysregulation of lung malignancy

Reduced nasal nitric oxide levels in patients with eosinophilic chronic rhinosinusitis

Background: In Eosinophilic chronic rhinosinusitis (ECRS), it is difficult to estimate the refractoriness and recurrence risk for each patient. Fraction of exhaled nitric oxide (FeNO) is known as a biomarker of eosinophilic inflammation in lower airway. It has been reported that nasal NO has some crucial functions in the upper and lower airways. However, in upper airway, paranasal sinuses, the usefulness of NO measurement remains controversial. The purpose of this study is to identify the usefulness of nasal NO measurement in ECRS and the involvement of nasal NO in the pathogenesis of ECRS. Methods: We compared the nasal NO levels of ECRS, non-ECRS, and normal control groups. Correlation between nasal NO levels and clinical findings were observed. Then, we compared nasal NO levels before and after endoscopic sinus surgery (ESS). We also examine whether nasal NO levels might discriminate ECRS by the receiver operating characteristic (ROC) curve analysis. Results: Nasal NO levels were significantly decreased in ECRS compared to the other two groups. Moreover, nasal NO levels in ECRS significantly and negatively correlated with eosinophil levels and CT score. However, they did not correlate with the nasal polyp score. Nasal NO levels were not upregulated soon after opening the sinus ostium by ESS. The ROC curves for nasal NO levels were used to discriminate all CRS patients and ECRS patients from normal controls. Conclusions: Nasal NO may be useful as a marker of ECRS severity and low nasal NO levels in ECRS may contribute to its pathogenesis. Keywords: Asthma, Chronic rhinosinusitis, Eosinophilic chronic rhinosinusitis, Eosinophilic inflammation, Nasal nitric oxide leve