Neural crest-derived cells in nasal conchae of adult mice contribute to bone regeneration.

Neural crest-derived cells (NCDCs), a class of adult stem cells not restricted to embryonic tissues, are attractive tissue regenerative therapy candidates because of their ease of isolation, self-renewing properties, and multipotency. Although adult NCDCs can undergo osteogenic differentiation in vitro, whether they induce bone formation in vivo remains unclear. Previously, our group reported findings showing high amounts of NCDCs scattered throughout nasal concha tissues of adult mice. In the present study, NCDCs in nasal conchae labeled with enhanced green fluorescent protein (EGFP) were collected from adult P0-Cre/CAG-CAT-EGFP double transgenic mice, then cultured in serum-free medium to increase the number. Subsequently, NCDCs were harvested and suspended in type I atelocollagen gel, then an atelocollagen sponge was used as a scaffold for the cell suspension. Atelocollagen scaffolds with NCDCs were placed on bone defects created in a mouse calvarial bone defect model. Over the ensuing 12 weeks, micro-CT and histological analysis findings showed that mice with scaffolds containing NCDCs had slightly greater bone formation as compared to those with a scaffold alone. Furthermore, Raman spectroscopy revealed spectral properties of bone in mice that received scaffolds with NCDCs similar to those of native calvarial bone. Bone regeneration is important not only for gaining bone mass but also chemical properties. These results are the first to show the validity of biomolecule-free adult nasal concha-derived NCDCs for bone regeneration, including the chemical properties of regenerated bone tissue.

Strong correlation between air-liquid interface cultures and in vivo transcriptomics of nasal brush biopsy.

Air-liquid interface (ALI) cultures are ex vivo models that are used extensively to study the epithelium of patients with chronic respiratory diseases. However, the in vitro conditions impose a milieu different from that encountered in the patient in vivo, and the degree to which this alters gene expression remains unclear. In this study we employed RNA sequencing to compare the transcriptome of fresh brushings of nasal epithelial cells with that of ALI-cultured epithelial cells from the same patients. We observed a strong correlation between cells cultured at the ALI and cells obtained from the brushed nasal epithelia: 96% of expressed genes showed similar expression profiles, although there was greater similarity between the brushed samples. We observed that while the ALI model provides an excellent representation of the in vivo airway epithelial transcriptome for mechanistic studies, several pathways are affected by the change in milieu.

Expression of Estrogen Receptor-alpha in Nasal Polyps and the Effects of Dexamethasone on Estrogen Receptor-alpha Expression in RPMI 2650 Cells.

BACKGROUND: Studies have reported that epithelial cell proliferation may be involved in the pathogenesis of nasal polyps (NPs). Estrogen receptor (ER)-α, one type of ER, is related to anti-inflammatory action and cell survival in certain tissues. In this study, we examined the presence or absence of ER-α in NPs and healthy inferior turbinate mucosae. We also investigated the effect of dexamethasone on ER-α expression, cell viability, and apoptosis in RPMI 2650 cells. METHODS: Immunohistochemical staining and Western blot analysis were conducted to determine the expression of ER-α in 15 NPs and 15 healthy inferior turbinate mucosae. After treating RPMI 2650 cells with dexamethasone, ER-α expression was analyzed using Western blot analysis and cell viability was determined using the MTT assay. Western blot analysis and annexin V-phycoerythrin (PE) staining were used to examine apoptotic cell death. RESULTS: Western blot analysis showed that ER-α expression was upregulated in 13 of the 15 NP tissues. Immunohistochemical staining for ER-α confirmed the results of the Western blot analysis. When RPMI 2650 cells were treated with dexamethasone, both ER-α expression and cell viability were decreased. Furthermore, the treatment of RPMI 2650 cells with dexamethasone increased apoptotic cell death, as shown by increased levels of BAX and cleaved caspase-3, decreased levels of Bcl-2, and an increased percentage of positive annexin V-PE stained cells. CONCLUSION: ER-α expression was higher in NPs than in healthy inferior turbinate mucosae. When RPMI 2650 cells were treated with dexamethasone, ER-α expression was downregulated, cell viability decreased, and apoptosis increased. The decreased cell viability may be related, at least in part, to the decreased ER-α protein levels, which likely contributed to the induction of apoptotic cell death in RPMI 2650 cells.

15-Lipoxygenase 1 in nasal polyps promotes CCL26/eotaxin 3 expression through extracellular signal-regulated kinase activation.

BACKGROUND: 15-Lipoxygenase 1 (15LO1) is expressed in airway epithelial cells in patients with type 2-high asthma in association with eosinophilia. Chronic rhinosinusitis with nasal polyps (CRSwNP) is also associated with type 2 inflammation and eosinophilia. CCL26/eotaxin 3 has been reported to be regulated by 15LO1 in lower airway epithelial cells. However, its relation to 15LO1 in patients with CRSwNP or mechanisms for its activation are unclear. OBJECTIVE: We sought to evaluate 15LO1 and CCL26 expression in nasal epithelial cells (NECs) from patients with CRSwNP and healthy control subjects (HCs) and determine whether 15LO1 regulates CCL26 in NECs through extracellular signal-regulated kinase (ERK) activation. METHODS: 15LO1, CCL26, and phosphorylated ERK were evaluated in NECs from patients with CRSwNP and HCs. 15LO1/CCL26 and CCL26/cytokeratin 5 were colocalized by means of immunofluorescence. IL-13-stimulated NECs were cultured at an air-liquid interface with or without 15-lipoxygenase 1 gene (ALOX15) Dicer-substrate short interfering RNAs (DsiRNA) transfection, a specific 15LO1 enzymatic inhibitor, and 2 ERK inhibitors. Expression of 15LO1 and CCL26 mRNA and protein was analyzed by using quantitative RT-PCR, Western blotting, and ELISA. RESULTS: 15LO1 expression was increased in nasal polyp (NP) epithelial cells compared with middle turbinate epithelial cells from patients with CRSwNP and HCs. 15LO1 expression correlated with CCL26 expression and colocalized with CCL26 expression in basal cells of the middle turbinate and NPs from patients with CRSwNP. In primary NECs in vitro, IL-13 induced 15LO1 and CCL26 expression. 15LO1 knockdown and inhibition decreased IL-13-induced ERK phosphorylation and CCL26 expression. ERK inhibition (alone) similarly decreased IL-13-induced CCL26. Phosphorylated ERK expression was increased in NECs from CRSwNP subjects and positively correlated with both 15LO1 and CCL26 expression. CONCLUSIONS: 15LO1 expression is increased in NP epithelial cells and contributes to CCL26 expression through ERK activation. 15LO1 could be considered a novel therapeutic target for CRSwNP.

Decreased ciliary beat responsiveness to acetylcholine in the nasal polyp epithelium.

OBJECTIVE: We investigated the difference in ciliary beat responsiveness to acetylcholine in ex vivo and the difference in the expressions of associated molecules (M1/M3 muscarinic receptors, pannexin-1 and P2X7 purinergic receptor) between the nasal polyp and turbinate mucosa. STUDY DESIGN: Laboratorial study. PARTICIPANTS: Nasal polyp and inferior turbinate were collected from patients with hypertrophic rhinitis and/or nasal polyp during endoscopic sinonasal surgery. MAIN OUTCOME MEASURES: The mucosa was cut into thin strips, and ciliary movement was observed under a phase-contrast light microscope equipped with a high-speed digital video camera. The samples were also examined by scanning electron microscopy, fluorescence immunohistochemistry, and quantitative reverse transcription-polymerase chain reaction. RESULTS: Cilia were well preserved in both tissues at the ultrastructural level. The baseline ciliary beat frequency (CBF) was not different between the two tissues. The CBF of the turbinate was significantly increased by stimulation with acetylcholine (P < 0.001), but that of the polyp was not. The ratio of the acetylcholine-stimulated CBF to the baseline CBF was significantly lower in the polyp than in the turbinate (P < 0.001). Immunohistochemical study revealed that immunoreactivities for M3, pannexin-1 and P2X7 were weaker in the polyp than in the turbinate. The mRNA expressions of M1, M3 and P2X7 were significantly lower and that of pannexin-1 tended to be lower in the polyp than in the turbinate. CONCLUSIONS: These results indicate that ciliary beat responsiveness to acetylcholine is decreased in the nasal polyp. This may be explained by the decreased expressions of M3, P2X7 and probably pannexin-1 in this tissue.

Synergistic mucus secretion by histamine and IL-4 through TMEM16A in airway epithelium.

Histamine is an important mediator of allergic reactions, and mucus hypersecretion is a major allergic symptom. However, the direct effect of histamine on mucus secretion from airway mucosal epithelia has not been clearly demonstrated. TMEM16A is a Ca2+-activated chloride channel, and it is closely related to fluid secretion in airway mucosal epithelia. We investigated whether histamine directly induces fluid secretion from epithelial cells or submucosal glands (SMG) and mechanisms related, therewith, in allergic airway diseases. In pig airway tissues from the nose or trachea, histamine was a potent secretagogue that directly induced strong responses. However, gland secretion from human nasal tissue was not induced by histamine, even in allergic rhinitis patients. Histamine type 1 receptor (H1R) and histamine type 2 receptor (H2R) were not noted in SMG by in situ hybridization. Cultured primary human nasal epithelial (NHE) cells were used for the measurement of short-circuit current changes with the Ussing chamber. Histamine-induced slight responses of anion secretions under normal conditions. The response was enhanced by IL-4 stimulation through TMEM16A, which might be related to fluid hypersecretion in allergic rhinitis. Pretreatment with IL-4 augmented the histamine response that was suppressed by a TMEM16A inhibitor. TMEM16A expression was enhanced by 24-h treatment of IL-4 in human nasal epithelial cells. The expression of TMEM16A was significantly elevated in an allergic rhinitis group, compared with a control group. We elucidated histamine-induced fluid secretions in synergy with IL-4 through TMEM16A in the human airway epithelium. In addition, we observed species differences between pigs and humans in terms of gland secretion of histamine.

Activity of Multidrug Resistance-Associated Proteins 1-5 (MRP1-5) in the RPMI 2650 Cell Line and Explants of Human Nasal Turbinate.

The profound influence of ATP-binding cassette (ABC) transporters on the disposition of numerous drugs has led to increased interest in characterizing their expression profiles in various epithelial and endothelial barriers. The present work examined the presence and functional activity of five ABC efflux proteins, i.e., MRP 1-5, in freshly isolated human nasal epithelial cells and two in vitro models based on the human RPMI 2650 cell line. To evaluate the expression patterns of MRP1, MRP2, MRP3, MRP4, and MRP5 at the mRNA and protein levels in the ex vivo model and the differently cultured RPMI 2650 cells, reverse transcriptase polymerase chain reaction (RT-PCR), Western blot analysis, and indirect immunofluorescence staining were used. The functionality of the MRP transporters in the three models was assessed using efflux experiments and accumulation assays with the respective substrates and inhibitors. The mRNA and protein expression of all selected ABC transporters was detected in excised human nasal mucosa as well as in the corresponding cell culture models. Moreover, the functional expression of the MRP transport proteins was demonstrated in the three models for the first time. Therefore, the potential impact of multidrug resistance-associated proteins 1-5 on drug disposition after intranasal administration may be taken into consideration for future developments. The specimens of human nasal turbinate exhibited slightly lower efflux capacities of MRP1, MRP3, and MRP5 in relation to the submerged and ALI-cultured RPMI 2650 cells, but showed a promising comparability to both in vitro models concerning the activity of MRP2 and MRP4. In this regard, the different RPMI 2650 cell culture models will be able to provide useful experimental data in the preclinical phase to estimate the interaction of particular efflux transporters with drug candidates for nasal application.

1,8-Cineole potentiates IRF3-mediated antiviral response in human stem cells and in an ex vivo model of rhinosinusitis.

The common cold is one of the most frequent human inflammatory diseases caused by viruses and can facilitate bacterial superinfections, resulting in sinusitis or pneumonia. The active ingredient of the drug Soledum, 1,8-cineole, is commonly applied for treating inflammatory diseases of the respiratory tract. However, the potential for 1,8-cineole to treat primary viral infections of the respiratory tract remains unclear. In the present study, we demonstrate for the first time that 1,8-cineole potentiates poly(I:C)-induced activity of the antiviral transcription factor interferon regulatory factor 3 (IRF3), while simultaneously reducing proinflammatory nuclear factor (NF)-κB activity in human cell lines, inferior turbinate stem cells (ITSCs) and in ex vivo cultivated human nasal mucosa. Co-treatment of cell lines with poly(I:C) and 1,8-cineole resulted in significantly increased IRF3 reporter gene activity compared with poly(I:C) alone, whereas NF-κB activity was reduced. Accordingly, 1,8-cineole- and poly(I:C) treatment led to increased nuclear translocation of IRF3 in ITSCs and a human ex vivo model of rhinosinusitis compared with the poly(I:C) treatment approach. Nuclear translocation of IRF3 was significantly increased in ITSCs and slice cultures treated with lipopolysaccharide (LPS) and 1,8-cineole compared with the LPS-treated cells mimicking bacterial infection. Our findings strongly suggest that 1,8-cineole potentiates the antiviral activity of IRF3 in addition to its inhibitory effect on proinflammatory NF-κB signalling, and may thus broaden its field of application.

Electrical Impedance and Expression of Tight Junction Components of the Nasal Turbinate and Polyp.

PURPOSE: We investigated the electrical impedance and expression of tight junction components of the turbinate mucosa, nasal polyp, and normal skin. PROCEDURES: The inferior turbinate and nasal polyp of patients with chronic rhinosinusitis and the postauricular skin of patients with otitis media were examined. Electrical impedance was measured in vivo using a tissue conductance meter. Expressions of claudin-1 and tricellulin were examined by fluorescence immunohistochemistry and quantitative RT-PCR. RESULTS: Electrical impedance was higher in the skin than in the turbinate and polyp, but did not differ between the turbinate and polyp. Immunoreactivities for claudin-1 and tricellulin were seen in the epithelial/epidermal layer. Expression of claudin-1 was higher in the skin than in the turbinate and polyp. The polyp tended to show higher expression of claudin-1 but showed lower expression of tricellulin than the turbinate. The ratio of claudin-1 to tricellulin was highest in the skin and lowest in the turbinate. The correlation between expressions of the two tight junction components was strongly positive in the skin (r = 0.964) and negative (r = -0.527) in the turbinate and polyp. CONCLUSIONS: These results suggest that the roles of claudin-1 and tricellulin in barrier function may be complementary, and may thereby maintain a constant level of permeability of the mucosal tissues.

Comparison of nasal nitric oxide levels between the inferior turbinate surface and the middle meatus in patients with symptomatic allergic rhinitis.

BACKGROUND: Because of the anatomical complexity and the high output of the human nose, it has been unclear whether nasal nitric oxide (NO) serves as a reliable marker of allergic rhinitis (AR). We examined whether nasal NO levels in the inferior turbinate (IT) surface and the middle meatus (MM) differ in symptomatic AR patients. METHODS: We measured fractional exhaled NO (FeNO) and nasal NO in normal subjects (n = 50) and AR patients with mild symptoms (n = 16) or moderate or severe symptoms (n = 27). Nasal NO measurements were obtained using an electrochemical analyzer connected to a catheter and an air-suction pump (flow rate 50mL/sec). RESULTS: Compared to the normal subjects, the AR patients showed significantly higher nasal FeNO and nasal NO levels in the IT area. No significant difference in the MM area was observed among the three groups. The MM area showed higher NO levels than the IT area in all three groups. The ratio of nasal NO levels of the MM area to the IT area (MM/IT ratio) was significantly lower in the AR groups. The moderate/severe AR patients showed significantly higher nasal NO in the IT area (104.4 vs. 66.2ppb) and lower MM/IT ratios than those in the mild AR patients. The analysis of nasal brushing cells revealed significantly higher eosinophil cationic protein and nitrotyrosine levels in the AR groups. CONCLUSIONS: Nasal NO assessment in the IT area directly reflects persistent eosinophilic inflammation and may be a valid marker to estimate the severity of AR.

[1-11C]-Butanol Positron Emission Tomography reveals an impaired brain to nasal turbinates pathway in aging amyloid positive subjects.

BACKGROUND: Reduced clearance of cerebrospinal fluid (CSF) has been suggested as a pathological feature of Alzheimer's disease (AD). With extensive documentation in non-human mammals and contradictory human neuroimaging data it remains unknown whether the nasal mucosa is a CSF drainage site in humans. Here, we used dynamic PET with [1-11C]-Butanol, a highly permeable radiotracer with no appreciable brain binding, to test the hypothesis that tracer drainage from the nasal pathway reflects CSF drainage from brain. As a test of the hypothesis, we examined whether brain and nasal fluid drainage times were correlated and affected by brain amyloid. METHODS: 24 cognitively normal subjects (≥ 65 years) were dynamically PET imaged for 60 min. using [1-11C]-Butanol. Imaging with either [11C]-PiB or [18F]-FBB identified 8 amyloid PET positive (Aß+) and 16 Aß- subjects. MRI-determined regions of interest (ROI) included: the carotid artery, the lateral orbitofrontal (LOF) brain, the cribriform plate, and an All-turbinate region comprised of the superior, middle, and inferior turbinates. The bilateral temporalis muscle and jugular veins served as control regions. Regional time-activity were used to model tracer influx, egress, and AUC. RESULTS: LOF and All-turbinate 60 min AUC were positively associated, thus suggesting a connection between the brain and the nose. Further, the Aß+ subgroup demonstrated impaired tracer kinetics, marked by reduced tracer influx and slower egress. CONCLUSION: The data show that tracer kinetics for brain and nasal turbinates are related to each other and both reflect the amyloid status of the brain. As such, these data add to evidence that the nasal pathway is a potential CSF drainage site in humans. These data warrant further investigation of brain and nasal contributions to protein clearance in neurodegenerative disease.

Human Nasal Inferior Turbinate-Derived Neural Stem Cells Improve the Niche of Substantia Nigra Par Compacta in a Parkinson's Disease Model by Modulating Hippo Signaling.

BACKGROUND: Parkinson's disease (PD) is one of the most prevalent neurodegenerative diseases, following Alzheimer's disease. The onset of PD is characterized by the loss of dopaminergic neurons in the substantia nigra. Stem cell therapy has great potential for the treatment of neurodegenerative diseases, and human nasal turbinate-derived stem cells (hNTSCs) have been found to share some characteristics with mesenchymal stem cells. Although the Hippo signaling pathway was originally thought to regulate cell size in organs, recent studies have shown that it can also control inflammation in neural cells. METHODS: Dopaminergic neuron-like cells were differentiated from SH-SY5Y cells (DA-Like cells) and treated with 1-Methyl-4-phenylpyridinium iodide to stimulate Reactive oxidative species (ROS) production. A transwell assay was conducted to validate the effect of hNTSCs on the Hippo pathway. We generated an MPTP-induced PD mouse model and transplanted hNTSCs into the substantia nigra of PD mice via stereotaxic surgery. After five weeks of behavioral testing, the brain samples were validated by immunoblotting and immunostaining to confirm the niche control of hNTSCs. RESULTS: In-vitro experiments showed that hNTSCs significantly increased cell survival and exerted anti-inflammatory effects by controlling ROS-mediated ER stress and hippocampal signaling pathway factors. Similarly, the in-vivo experiments demonstrated an increase in anti-inflammatory effects and cell survival rate. After transplantation of hNTSCs, the PD mouse model showed improved mobility and relief from PD symptoms. CONCLUSION: hNTSCs improved the survival rate of dopaminergic neurons by manipulating the hippocampal pathway through Yes-associated protein (YAP)/transcriptional coactivator with a PDZ-binding motif (TAZ) by reducing inflammatory cytokines. In this study, we found that controlling the niche of hNTSCs had a therapeutic effect on PD lesions.

Antiviral activity of bovine type III interferon against bovine viral diarrhea virus is greatly reduced in bovine turbinate cells due to limited expression of IFN lambda receptor 1 (IL-28Rα).

Introduction: The antiviral activity of recombinant bovine interferon lambda 3 (bovIFN-λ3) against bovine viral diarrhea virus (BVDV) has been demonstrated in vitro in Madin-Darby bovine kidney cells (MDBK) and in vivo in cattle. However, anti-BVDV activity of bovIFN-λ3 has not been studied in bovine respiratory tract epithelial cells, supposedly a primary target of BVDV infection when entering the host by the oronasal route. Methods: Here we investigated the anti-BVDV activity of bovIFN-λ3 in bovine turbinate-derived primary epithelial cells (BTu) using BVDV infection and immunoperoxidase staining, TCID50, RT-qPCR, DNA and transcriptome sequencing, and transfection with plasmids containing the two subunits, IL-28Rα and IL-10Rß that constitute the bovIFN-λ3 receptor. Results: Our immunoperoxidase staining, RT-qPCR, and TCID50 results show that while BVDV was successfully cleared in MDBK cells treated with bovIFN-λ3 and bovIFN-α, only the latter, bovIFN-α, cleared BVDV in BTu cells. Preincubation of MDBK cells with bovIFN-λ3 before BVDV infection was needed to induce optimal antiviral state. Both cell types displayed intact type I and III IFN signaling pathways and expressed similar levels of IL-10Rß subunit of the type III IFN receptor. Sequencing of PCR amplicon of the IL-28Rα subunit revealed intact transmembrane domain and lack of single nucleotide polymorphisms (SNPs) in BTu cells. However, RT-qPCR and transcriptomic analyses showed a lower expression of IL-28Rα transcripts in BTu cells as compared to MDBK cells. Interestingly, transfection of BTu cells with a plasmid encoding IL-28Rα subunit, but not IL-10Rß subunit, established the bovIFN-λ3 sensitivity showing similar anti-BVDV activity to the response in MDBK cells. Conclusion: Our results demonstrate that the sensitivity of cells to bovIFN-λ3 depends not only on the quality but also of the quantity of the IL-28Rα subunit of the heterodimeric receptor. A reduction in IL-28Rα transcript expression was detected in BTu as compared to MDBK cells, despite the absence of spliced variants or SNPs. The establishment of bovIFN-λ3 induced anti-BVDV activity in BTu cells transfected with an IL-28Rα plasmid suggests that the level of expression of this receptor subunit is crucial for the specific antiviral activity of type III IFN in these cells.

Increased expression of dectin-1 in nasal polyps.

PURPOSE: A fungal etiology has been proposed to underlie severe nasal polyps (NP). Dectin-1 is an innate immune pattern recognition receptor which is involved in the recognition of some pathogenic fungi. We investigated the Dectin-1 levels in NP in order to evaluate the implication of such expression with respect to the development of NP. MATERIALS AND METHODS: Normal inferior turbinate tissues were obtained from forty patients undergoing surgery for augmentation rhinoplasty. Nasal polyp tissues were obtained from 53 patients who underwent endoscopic sinus surgery for chronic polypoid rhinosinusitis. Real-time polymerase chain reaction and Western blot analysis were performed to evaluate the mRNA and protein level of Dectin-1, respectively. ELISA was carried out to evaluate the cytokine production (IL-4, IL-5, IL-10, and TNF-α) in NP. RESULTS: Real-time polymerase chain reaction and Western-blot analysis showed that Dectin-1 expression in NP was increased compared with that in normal nasal inferior turbinate tissues. ELISA results suggest that the local expression of type-1 and type-2 inflammatory cytokine is skewed toward type-2 inflammatory cytokine in NP. CONCLUSIONS: These results suggest that Dectin-1 may play a role in the development of NP, and the production of Dectin-1, IL-4 and IL-5 (type-2 cytokines), may mainly participate in the inflammatory reaction in NP.

Concha bullosa surgery and the distribution of human olfactory neuroepithelium.

In bullous middle turbinate surgery, controversy exists over which side of the bullous middle turbinate should be removed, as the distribution of human olfactory neuroepithelium is unclear. This study evaluated whether the middle turbinate tissue of patients undergoing endoscopic concha bullosa surgery contains functional olfactory epithelium. This prospective clinical study was conducted in tertiary referable center. It detected 70 conchae bullosa in 48 patients with sinonasal symptoms, who underwent paranasal computed tomography (CT) that showed pneumatization of the middle concha. All samples were obtained under general anesthesia. Three samples were obtained from each bullous middle turbinate: one each from the anterior, medial, and lateral portions. The mucosa from each sample was stained with olfactory marker protein (OMP). In total, 210 middle turbinate samples were taken from 48 patients during endoscopic surgery for conchae bullosa. The patients were 22 females and 26 males. Of the 70 conchae bullosa, OMP-stained nerve tissue was found in the lateral, anterior and medial aspects of 57 (81.4 %), 42 (60.0 %) and 23 (32.8 %) of the bullous middle turbinates, respectively. OMP-stained nerve tissue was found in 122 (58.1 %) of the 210 bullous middle turbinate tissue samples. OMP-stained nerve tissue was found on the lateral surface of the bullous middle turbinate more often than the medial surface. Therefore, during the concha bullosa surgery, OMP-stained nerve tissue found at least in the medial part of concha, suggested that the opening of the medial part of middle concha.

Eosinophils and mast cells: a comparison of nasal mucosa histology and cytology to markers in nasal discharge in patients with chronic sino-nasal diseases.

Allergic rhinitis (AR), nasal polyps (NP) as well as chronic rhinosinusitis (CRS) are all known to be associated with eosinophilic infiltration and elevated numbers of mast cells (MC) within the mucosa. Both cell types and their markers eosinophilic cationic protein (ECP) and tryptase are utilized in the diagnosis and management of chronic sino-nasal diseases. Mucosal cytology samples were gathered by cytobrush, histological samples were obtained from the inferior turbinate. In both sample sets, the number of eosinophils and MC was determined. Their corresponding markers ECP and tryptase were quantified from nasal discharge. Patients were grouped with reference to their main diagnosis: AR (n = 34), NP (n = 25), CRS (n = 27) and controls (n = 34). Eosinophil counts from cytobrush and ECP levels were significantly elevated in NP compared to all other groups-31- and 13-fold over control, respectively. However, histologic review did not reveal any difference in eosinophil count among groups. Tryptase was significantly elevated threefold in AR versus CRS and controls. No correlation to cytological and histological MC counts could be found. ECP levels in nasal discharge as well as eosinophil counts can provide useful information with regard to the diagnosis. Likewise, tryptase concentrations can do. The presented data show that the measurement of markers in nasal discharge is superior in differentiating among diagnosis groups. Given that the collection of nasal secretions is more comfortable for patients than the more invasive techniques, we recommend first line ECP and tryptase testing performed on nasal secretions.

Immunohistochemical studies of wound healing after monopolar electrocautery and ultrasound submucosal inferior nasal turbinate reduction in sheep.

BACKGROUND: Extracellular matrix (ECM) proteins such as fibronectin and collagen III, enzymes such as matrix metalloproteinases and macrophages have been demonstrated to intervene in nasal and paranasal sinuses wound healing. AIM OF THE STUDY: To compare concentration of ECM proteins, enzymes and the recruitment of macrophages during wound repair after monopolar electrocautery in contrast with ultrasound submucosal surgical tissue reduction of inferior nasal turbinate (INT) tested in sheep. MATERIALS AND METHODS: Prospective controlled study in sheep. Immunostaining for collagen III, fibronectin, CD68 and matrix metalloproteinase-9 (MMP9) was applied in tissue specimens of INT mucosa after monopolar electrocoagulation (MEC) and ultrasound tissue reduction (UTR). Twelve INTs were studied 1, 3 and 8 weeks post-operatively in each interventional group (MEC and UTR) and 5 INTs were studied in animals of the control group (without surgery). The immunoreactivity was quantitatively graded between 0% to 100% immunoreactivity by a blinded senior pathologist. RESULTS: At the end of the study period collagen III, fibronectin and MMP9 were increased in both groups compared to the levels of the control group. When compared to control group, CD68 immunoreactivity was found higher in MEC group but not in UTR group. Fibronectin subepithelial immunoreactivity exhibited a substantial negative correlation with mucosal epithelial cell necrosis, a substantial positive correlation with fibrosis in MEC-treated specimens and a significant positive correlation with sinusoid engorgement in UTR-treated specimens. Collagen III tissue immunoreactivity showed a particularly significant negative correlation with sinusoid engorgement in MEC-treated specimens. CONCLUSION: Correlation of fibronectin and collagen III immunoreactivity to histopathologic findings suggests different ECM repair processes between MEC and UTR turbinate tissue reduction. The use of CD68 and MMP9 provides additional clues to the mode of actions of these techniques and to the molecular and cellular events of the nasal mucosa wound healing process.

Localization and up-regulation of cysteinyl leukotriene-2 receptor in human allergic nasal mucosa.

BACKGROUND: Cysteinyl leukotrienes (CysLTs) are lipid mediators that have been implicated in the pathogenesis of allergic rhinitis. Pharmacological studies using CysLTs indicate that 2 classes of receptors exist, namely, CysLT1 and CysLT2 receptors. The former class of receptors is sensitive to the CysLT1 antagonists currently used to treat asthma and allergic rhinitis, and its localization has been previously examined by our group using immunohistochemistry and in situ hybridization techniques. We investigated the expression and localization of the CysLT2 receptor in human nasal mucosa by western blot and immunohistochemical analyses. METHODS: Human turbinates were obtained after turbinectomy from 16 patients with nasal obstruction refractory to medical therapy. To identify the cells expressing the CysLT2 receptor, double immunostaining was performed by using anti-CysLT2 receptor antibody and anti-CD31 (endothelial cell) antibody or anti-smooth muscle actin antibody. RESULTS: A 39 kDa band was detected on the western blots of human turbinates samples by using the anti-CysLT2 receptor antibody. The expression level of the CysLT2 receptor in patients with nasal allergy was higher than that in patients with non-allergic rhinitis. The immunohistochemical study also showed an intense immunoreactivity for CysLT2 receptor in both vascular endothelial cells and vascular smooth muscles. CONCLUSIONS: The results indicated that the CysLT2 receptor plays a primary role in the vascular responses in the upper respiratory tract.

Expression and localization of purinergic P2Y(12) receptor in human nasal mucosa.

BACKGROUND: Extracellular nucleotides such as ATP and UTP are released from essentially all cells, and they interact with cell surface P2 receptors to produce a broad range of physiological responses. P2Y12 receptor is the major platelet receptor that mediates ADP-induced aggregation, P2Y12 receptor inhibitors such as clopidogrel and prasugrel inhibit platelet aggregation, and thus, they are used in the treatment and prevention of coronary artery disease. Recently, studies have focused on the P2Y12 receptor as a receptor for leukotriene E4 (LTE4), because this receptor is required for LTE4-mediated pulmonary inflammation. To establish the presence of P2Y12 receptor in human nasal mucosa, we investigated the expression and the localization of the P2Y12 receptor in human nasal mucosa. METHODS: Human turbinates were obtained by turbinectomy from 12 patients with nasal obstruction refractory to medical therapy. The expression of P2Y12 receptor was evaluated by RT-PCR, western blotting, and immunohistochemical analysis. RESULTS: RT-PCR analysis of total RNA extracted from human nasal turbinate, primary cultured human nasal epithelial cells and nasal vascular endothelial cells demonstrated the expression of P2Y12 receptor mRNA. A band of approximately 55 kDa was detected in human turbinates by western blot analysis using anti-P2Y12 receptor antibody. We could not find any differences between P2Y12 receptor levels in allergic and non-allergic nasal mucosa. An immunohistochemical study revealed that epithelial cells, submucosal glands and vascular endothelial cells showed intense immunoreactivity for the P2Y12 receptor. CONCLUSIONS: The results may have important clinical implications for understanding the role of P2Y12 receptor in upper airway diseases such as allergic rhinitis and non-allergic rhinitis.

Characteristics of Human Nasal Turbinate Stem Cells under Hypoxic Conditions.

This study investigated the influence of hypoxic culture conditions on human nasal inferior turbinate-derived stem cells (hNTSCs), a subtype of mesenchymal stem cells (MSCs). It aimed to discern how hypoxia affected hNTSC characteristics, proliferation, and differentiation potential compared to hNTSCs cultured under normal oxygen levels. After obtaining hNTSCs from five patients, the samples were divided into hypoxic and normoxic groups. The investigation utilized fluorescence-activated cell sorting (FACS) for surface marker analysis, cell counting kit-8 assays for proliferation assessment, and multiplex immunoassays for cytokine secretion study. Differentiation potential-osteogenic, chondrogenic, and adipogenic-was evaluated via histological examination and gene expression analysis. Results indicated that hNTSCs under hypoxic conditions preserved their characteristic MSC phenotype, as confirmed by FACS analysis demonstrating the absence of hematopoietic markers and presence of MSC markers. Proliferation of hNTSCs remained unaffected by hypoxia. Cytokine expression showed similarity between hypoxic and normoxic groups throughout cultivation. Nevertheless, hypoxic conditions reduced the osteogenic and promoted adipogenic differentiation potential, while chondrogenic differentiation was relatively unchanged. These insights contribute to understanding hNTSC behavior in hypoxic environments, advancing the development of protocols for stem cell therapies and tissue engineering.