Mucosa In Health Research Articles

Characterizing the T Cell Repertoire in the Proximal Airway in Health and Disease.

Recent translational scientific efforts in subglottic stenosis (SGS) support a disease model where epithelial alterations facilitate microbiome displacement, dysregulated immune activation, and localized fibrosis. Given the observed immune cell infiltrate in SGS, we sought to test the hypothesis that SGS cases possessed a low diversity (highly clonal) adaptive immune response when compared with healthy controls. Single cell RNA sequencing (scRNA-seq) of subglottic mucosal scar in iSGS (n = 24), iLTS (n = 8), and healthy controls (n = 7) was performed. T cell receptor (TCR) sequences were extracted, analyzed, and used to construct repertoire structure, compare diversity, interrogate overlap, and define antigenic targets using the Immunarch bioinformatics pipeline. The proximal airway mucosa in health and disease are equally diverse via Hill framework quantitation (iSGS vs. iLTS vs. Control, p > 0.05). Repertoires do not significantly overlap between individuals (Morisita <0.02). Among iSGS patients, clonality of the TCR repertoire is driven by CD8+ T cells, and iSGS patients possess numerous TCRs targeting viral and intercellular pathogens. High frequency clonotypes do not map to known targets in public datasets. SGS cases do not possess a lower diversity adaptive immune infiltrate when compared with healthy controls. Interestingly, the TCR repertoire in both health and disease contains a restricted number of high frequency clonotypes that do not significantly overlap between individuals. The target of the high frequency clonotypes in health and disease remain unresolved. NA Laryngoscope, 134:1757-1764, 2024.

Spatially resolved transcriptomics reveals pro-inflammatory fibroblast involved in lymphocyte recruitment through CXCL8 and CXCL10.

The interplay among different cells in a tissue is essential for maintaining homeostasis. Although disease states have been traditionally attributed to individual cell types, increasing evidence and new therapeutic options have demonstrated the primary role of multicellular functions to understand health and disease, opening new avenues to understand pathogenesis and develop new treatment strategies. We recently described the cellular composition and dynamics of the human oral mucosa; however, the spatial arrangement of cells is needed to better understand a morphologically complex tissue. Here, we link single-cell RNA sequencing, spatial transcriptomics, and high-resolution multiplex fluorescence in situ hybridisation to characterise human oral mucosa in health and oral chronic inflammatory disease. We deconvolved expression for resolution enhancement of spatial transcriptomic data and defined highly specialised epithelial and stromal compartments describing location-specific immune programs. Furthermore, we spatially mapped a rare pathogenic fibroblast population localised in a highly immunogenic region, responsible for lymphocyte recruitment through CXCL8 and CXCL10 and with a possible role in pathological angiogenesis through ALOX5AP. Collectively, our study provides a comprehensive reference for the study of oral chronic disease pathogenesis.

Impairment of Tissue-Resident Mesenchymal Stem Cells in Chronic Ulcerative Colitis and Crohn's Disease.

Little is known about the presence and function of tissue-resident mesenchymal stem cells [MtSCs] within the gastrointestinal mucosa in health and inflammatory bowel disease [IBD]. The contribution of MtSCs to the generation of inflammatory fibroblasts during IBD is also poorly understood. We hypothesized that IBD-MtSCs are impaired and contribute to the generation of the pathological myofibroblasts in IBD. In a cohort of clinically and endoscopically active IBD patients and normal controls, we used quantitative RT-PCR and stem cell differentiation assays, as well as confocal microscopy, to characterize MtSCs. Expression of two stem cell markers, Oct4 and ALDH1A, was increased in the inflamed IBD colonic mucosa and correlated with an increase of the mesenchymal lineage marker Grem1 in ulcerative colitis [UC], but not Crohn's disease [CD]. Increased proliferation and aberrant differentiation of Oct4+Grem1+ MtSC-like cells was observed in UC, but not in CD colonic mucosa. In contrast to normal and UC-derived MtSCs, CD-MtSCs lose their clonogenic and most of their differentiation capacities. Our data also suggest that severe damage to these cells in CD may account for the pathological PD-L1low phenotype of CD myofibroblasts. In contrast, aberrant differentiation of MtSCs appears to be involved in the appearance of pathological partially differentiated PD-L1high myofibroblasts within the inflammed colonic mucosa in UC. Our data show, for the first time, that the progenitor functions of MtSCs are differentially impaired in CD vs UC, providing a scientific rationale for the use of allogeneic MSC therapy in IBD, and particularly in CD.

Oral seeding and niche-adaptation of middle ear biofilms in health

The entrenched dogma of a sterile middle ear mucosa in health is incongruent with its periodic aeration and seeding with saliva aerosols. To test this, we sequenced 16S rRNA-V4 amplicons from otic secretions collected at the nasopharyngeal orifice of the tympanic tube and, as controls, oropharyngeal and buccal samples. The otic samples harbored a rich diversity of oral keystone genera and similar functional traits but were enriched in anaerobic genera in the Bacteroidetes (Prevotella and Alloprevotella), Fusobacteria (Fusobacterium and Leptotrichia) and Firmicutes (Veillonella) phyla. Facultative anaerobes in the Streptococcus genus were also abundant in the otic and oral samples but corresponded to distinct, and sometimes novel, cultivars, consistent with the ecological diversification of the oral migrants once in the middle ear microenvironment. Neutral community models also predicted a large contribution of oral dispersal to the otic communities and the positive selection of taxa better adapted to growth and reproduction under limited aeration. These results challenge the traditional view of a sterile middle ear in health and highlight hitherto unknown roles for oral dispersal and episodic ventilation in seeding and diversifying otic biofilms.

Defining human mesenchymal and epithelial heterogeneity in response to oral inflammatory disease.

Human oral soft tissues provide the first barrier of defence against chronic inflammatory disease and hold a remarkable scarless wounding phenotype. Tissue homeostasis requires coordinated actions of epithelial, mesenchymal, and immune cells. However, the extent of heterogeneity within the human oral mucosa and how tissue cell types are affected during the course of disease progression is unknown. Using single-cell transcriptome profiling we reveal a striking remodelling of the epithelial and mesenchymal niches with a decrease in functional populations that are linked to the aetiology of the disease. Analysis of ligand-receptor interaction pairs identify potential intercellular hubs driving the inflammatory component of the disease. Our work establishes a reference map of the human oral mucosa in health and disease, and a framework for the development of new therapeutic strategies.

Tu1622 - Acute Stress Impacts Clock Genes and Barrier Integrity in the Intestinal Mucosa in Health

Tu1622 - Acute Stress Impacts Clock Genes and Barrier Integrity in the Intestinal Mucosa in Health

Barrier function of the nasal mucosa in health and type-2 biased airway diseases.

The mucosal lining of the upper airways represents the outer surface of the body to the ambient air and its contents and is prepared for it as the first line of defense. Apart from the well-described physical barrier and the mucociliary clearance, a variety of systems, including the airway microbiome, antimicrobial proteins, damage-associated molecular patterns, innate lymphoid cells, epithelial-derived cytokines and chemokines, and finally the adaptive immune system, as well as eosinophils as newly appreciated defense cells form different levels of protection against and response to any possible intruder. Of interest especially for allergic airway disease, mucosal germs might not just elicit a classical Th1/Th17-biased inflammatory response, but may directly induce a type-2 mucosal inflammation. Innovative therapeutic interventions may be possible at different levels also; however, whether modulations of the innate or adaptive immune responses will finally be more successful, and how the correction of the adaptive immune response might impact on the innate side, will be determined in the near future.

Mucin profile of Upper Gastrointestinal Tract Lesions

Background: Mucin secretions reflect the composition changes in the functional state of the mucosa in health and disease of the upper gastrointestinal tract. Objectives: To study the different types of upper gastrointestinal tract lesions and to determine the role of mucin histochemistry in the interpretation of lesions of upper gastrointestinal tract. Methods: A prospective study from September 2006 to August 2007 was carried out. 60 endoscopic biopsy specimens obtained from upper gastrointestinal tract lesions from the department of Gastroenterology at R.L. Jalappa Hospital & Research Centre was sent for histopathological examination. All the sections were stained by routine Haematoxylin and Eosin stain followed by special stains for mucins i.e Alcian blue pH 2.5 / PAS and Hale\'s Colloidal Iron. Results: Age of presentation of various upper gastrointestinal lesions was between 26-90 years. Females dominated the study group i.e. 39 (65%) when compared to males i.e. 21 (35%). Alcian Blue/Periodic Acid Schiff (AB/PAS) staining in oesophageal lesions could detect mixture of acidic mucins & neutral mucins and sulphomucins in Barrett\'s oesophagus. In gastritis, predominantly mixture of acidic and neutral mucins and in one case, sulphomucins were detected. Adenocarcinoma of stomach showed combination of acidic and neutral mucins. Duodenitis showed acidic, neutral and sialomucins. Adenocarcinoma of duodenum showed combination of acidic & neutral mucins. Hale\'s Colloidal Iron (HCI) stain in inflammatory & neoplastic lesions of stomach & duodenum, could detect only acidic mucins. Conclusion: In neoplastic condition of stomach this study suggests that those tumours which secrete neutral and mixed mucins probably arise from gastric epithelium and neck cells of the gastric glands. In those cases where the tumour produces acidic mucins it is possible the tumour arose from areas of intestinal metaplasia. Key words: Mucins, Histochemistry, Alcian Blue / Periodic Acid Schiff, Hale\'s Colloidal Iron

Genetics of Complex Airway Disease

The past 3 years have seen highly significant genetic effects identified for a wide variety of common complex diseases, including the airway disorders of asthma and chronic obstructive pulmonary disease. It appears that only a portion of the genetically mediated susceptibility to complex diseases has been identified, and there is much left to be discovered. This review briefly describes the results of the genome-wide association studies of asthma and gives an overview of the parallel and increasingly large-scale studies that are taking place with chronic obstructive pulmonary disease. The future impact is discussed of technological advances that allow increasingly large-scale gene expression studies, next-generation sequencing, and genome-wide testing for epigenetic effects. The use of genetic technology to examine the airway microbiota that interact with the mucosa in health and disease is described.

Looking after local nasal and sinus mucosa in health, disease and after surgery

The paper emphasises the need for looking after of nasal and sinus mucosa in health, disease and after nasal surgery. It is a systematic arrangement of steps required to restore nasal mucosa to healthy state. These steps have been arranged to co relate them to diseases and symptoms for symptomatic and curative treatment. It can rationalise the need for surgery in cases non responsive to maximum medical treatment. It can improve postoperative surgical results after functional endoscopic sinus surgery (FESS) and other endoscopic assisted procedures.

Intramucosal Distribution of WNT Signaling Components in Human Esophagus

The molecular mechanisms governing the biology and pathobiology of esophageal squamous mucosa in health and disease are not completely understood. Earlier genome-wide expression study of normal-looking esophageal squamous mucosa has shown differential expression of the Wingless-type MMTV integration site family (Wnt) modulators Dickkopf (Dkk) homologs among healthy individuals and patients with reflux esophagitis and Barrett metaplasia suggesting that the Wnt pathway may be involved in esophageal mucosal biology. Seven full-thickness human donor esophagi were cryosectioned for immunohistochemical analysis, and lamina propria (LP), basal (BC), intermediate (IC), and superficial (SC) cells were also dissected by laser-capture microdissection for real-time polymerase chain reaction. Wnt1, 2b, and 3a were expressed primarily in BC, Wnt3, and 5b in LP, and Wnt5a in IC. Frizzled 1, low-density lipoprotein receptor-related protein 6, secreted frizzled-related protein 1, T-cell-specific transcription factor 3, and dishevelled 3 were expressed highest in LP decreasing precipitously medially toward SC. Dkk1 predominantly expressed in SC was more than 100-folds greater than other layers (P<0.001). Dkk4 was expressed primarily in SC but Dkk3 was opposite with greatest expression in LP. Immunohistochemical analysis showed Wnt1 and 3a in BC, Wnt5a in IC and SC, Dkk1 predominantly in SC, Dkk4 in SC and IC, and Dkk3 and SFRP1 in LP and BC

The immunohistology of CD40 in human oral epithelium in health and disease

CD40 has a role in the regulation of immune responses, cell proliferation and migration, and apoptosis. Little is known of its distribution in oral mucosal pathology. Oral keratinocyte lines were tested for CD40 protein by Western blotting. Immunohistochemistry was used to stain paraffin sections of oral mucosa in health and in inflammatory, reactive, dysplastic and malignant disease. Western blotting confirmed the presence of CD40 in oral keratinocytes. CD40 was generally expressed by keratinocytes in the basal layer, with variable parabasal expression. Langerhans cells also stained positively. Expression was lost in nine of 33 (27%) epithelial dysplasias, seven of which were severe. Eighty-one percent of well, 69% of moderately and 50% of poorly differentiated oral squamous cell carcinomas (OSCC) expressed CD40. Overall, 45 of 65 (69%) OSCC were positive. The pattern of expression was unrelated to tumour differentiation. CD40 expression by basal and parabasal oral keratinocytes is physiological. Expression is lost in approximately one-third of oral epithelial dysplasias and OSCC. The significance of such loss remains unknown, but may be related to immunological or other abnormalities of keratinocyte homeostasis.

Nasal nitric oxide measurements before and after repeated humming maneuvers.

It has been recently shown that humming greatly increases nasal nitric oxide (NO). This is most likely owing to a rapid washout of sinus NO caused by the oscillating sound waves. During repeated humming manoeuvres nasal NO gradually decreases, likely because NO accumulated in the sinuses is washed out. We studied whether humming before measurements would affect nasally exhaled NO. NO output was measured by the chemiluminescence technique in orally and nasally exhaled air in 38 subjects: 18 healthy subjects (HS), 15 subjects with allergic rhinitis (AR) and five subjects with allergic nasal polyposis (AP). Each subject performed a NO measurement during quiet nasal exhalation either preceded by a period of silence/free speaking or immediately after five consecutive humming manoeuvres (posthumming). Mean nasal NO output (95% CI) after a period of silence/free speaking was 231 nL min-1 (178-284) in HS, 434 nL min-1 (347-522) in AR (P < 0.001) and 262 nL min-1 (163-361) in AP. Post-humming nasal NO output was 16% (5 to 50%) lower in HS and 14% (1 to 49%) lower in AR, while it remained unchanged in AP subjects. Intra-subject coefficient of variation of quiet nasal exhalation was 12% in HS, 13% in AR and 5% in AP. Post humming intraindividual coefficient of variation significantly decreased in both HS and AR, but it did not change in AP. Nasal NO levels measured immediately after repeated humming manoeuvres are consistently lower and more reproducible than nasal NO levels measured after a period of silence or free speaking. Repeated humming effectively empties the sinuses, thereby probably minimizing the normal contribution from the sinuses to nasal NO. This may be useful to better estimate NO output from the nasal cavity mucosa in health and disease.

Prebiotics: actual and potential effects in inflammatory and malignant colonic diseases.

This paper will summarize the most recent clinical and experimental data on the effects of prebiotics in inflammatory and cancerous diseases of the large intestine. Animal studies, as well as data obtained in in-vitro cell culture systems, have underlined the potential of certain prebiotics to protect against inflammatory and cancerous processes in the large intestine. Clinical trials are now in progress to assess the relevance of these promising results. The biochemical mechanisms are still incompletely deciphered, but both the promotion of lactic acid-producing bacteria and the production of short-chain fatty acids, particularly butyrate, during the fermentation of prebiotics could be key factors. Enteric resident bacteria are involved in inflammatory bowel diseases and may contribute to colonic carcinogenesis. Dietary manipulation of the flora may thus represent a useful aid to prevent or to treat these diseases, and this could be a place for prebiotics. Inulin-like prebiotics have shown encouraging results in animal models, but clinical and epidemiological trials are necessary to define their efficacy in humans. In the next few years, important advances are expected in understanding the interactions between prebiotics, intestinal flora and the colonic mucosa in health and diseases, enabling the improvement of therapy as well as better nutritional handling of susceptible individuals.

Mature dendritic cells infiltrate the T cell-rich region of oral mucosa in chronic periodontitis: in situ, in vivo, and in vitro studies.

Previous studies have analyzed the lymphoid and myeloid foci within the gingival mucosa in health and chronic periodontitis (CP); however, the principal APCs responsible for the formation and organizational structure of these foci in CP have not been defined. We show that in human CP tissues, CD1a(+) immature Langerhans cells predominantly infiltrate the gingival epithelium, whereas CD83(+) mature dendritic cells (DCs) specifically infiltrate the CD4(+) lymphoid-rich lamina propria. In vivo evidence shows that exacerbation of CP results in increased levels of proinflammatory cytokines that mediate DC activation/maturation, but also of counterregulatory cytokines that may prevent a Th-polarized response. Consistently, in vitro-generated monocyte-derived DCs pulsed with Porphyromonas gingivalis strain 381 or its LPS undergo maturation, up-regulate accessory molecules, and release proinflammatory (IL-1beta, PGE(2)) and Th (IL-10, IL-12) cytokines. Interestingly, the IL-10:IL-12 ratio elicited from P. gingivalis-pulsed DCs was 3-fold higher than that from Escherichia coli-pulsed DCs. This may account for the significantly (p < 0.05) lower proliferation of autologous CD4(+) T cells and reduced release of IFN-gamma elicited by P. gingivalis-pulsed DCs. Taken together, these findings suggest a previously unreported mechanism for the pathophysiology of CP, involving the activation and in situ maturation of DCs by the oral pathogen P. gingivalis, leading to release of counterregulatory cytokines and the formation of T cell-DC foci.

Permeation of polysucrose 15000 across the human nasal mucosa in vivo.

Studies of the nasal permeation of small molecules (< 1000 Da) have yielded important information about the integrity of the human airway mucosa in health and disease. In this study, we used a much larger tracer molecule, polysucrose (PS) 15,000 (approx. 14,700 Da), to predict the mucosal permeation of inhalational allergens. PS 15,000 (50 mg/ml; 15 ml), with or without a detergent type of permeation enhancer (dioctyl sodium sulfosuccinate 10 mg/ml), was maintained for 15 min in one nasal cavity of 12 healthy nonatopic subjects by employment of a nasal-pool device. Permeation as determined by the 24-h urine recovery of PS (micro-ELISA analysis assay) was expressed as percentage of nasal instillate. Mean baseline permeation was 0.044% (range 0.009-0.250%). In the presence of the detergent, permeation increased to 0.600% (range 0.007-2.260%) (P < 0.01). After oral intake of 750 mg of PS 15,000 (50 micrograms/ml; 15 ml), the 24-h urinary recovery was 0.013% (range 0.004-0.023%). Our study thus demonstrates a measurable baseline permeation of PS 15,000, an elevated permeation rate in the presence of an epithelium-damaging detergent molecule, and a negligible permeation by the oral route. These properties support the utility of PS 15,000 as a nasal airway permeation tracer. Its size further suggests that its permeation may reflect mucosal perviousness to many allergens.

The "nasal pool"-device for challenge and lavage of the nasal mucosa in children: histamine-induced plasma exudation responses.

The accessibility of the nasal airway allows important examination of the airway mucosa in health and disease. However, the current methods for nasal challenge and lavage in children suffer from several shortcomings. In the present study, we have assessed the utility of a recently developed "nasal pool"-device in 7-9 year old children, and explored the ability of the nasal mucosa of these school-children to mount a plasma exudation response (lumenal entry of bulk plasma). Isotonic saline was instilled and maintained (1 min) as a "pool" in the unilateral nasal cavity. Recovery of the "pool" (lavage fluids) was determined. Concomitant challenge and lavage was then performed by exposing the nasal mucosa to a "pool" of isotonic saline and histamine (40-400 micrograms/ml) for 2 min. "Pool" fluids were analysed for alpha 2-macroglobulin as an index of microvascular-epithelial exudation of bulk plasma. The school-children successfully managed to carry out nasal lavages as well as concomitant histamine challenges and lavages with the "nasal pool"-device. The recovery of the nasal lavage fluids was almost quantitative (> 85%) and thus well reproducible. Histamine produced significant exudation of bulk plasma (alpha 2-macroglobulin). We suggest that the "nasal pool"-device is well suited for challenge and lavage of the nasal airway mucosa in children above 6 years of age, and conclude that lumenal entry of multipotent humoural protein-systems may be an important first line respiratory defence mechanism in children.

Expression of TNF and the 55-kDa TNF receptor in epidermis, oral mucosa, lichen planus and squamous cell carcinoma.

TNF has diverse biological effects including a role in the immune response and growth regulatory effects. The aim of this study was to determine the nature and location of cells able to synthesise and respond to TNF in oral mucosa in health and disease. The location of immunoreactive TNF and 55-kDa and 75-kDa TNF receptors was demonstrated using immunohistology. We also used RT-PCR to help determine the sites of synthesis of this cytokine in oral mucosa. Only occasional TNF-positive cells were detected in normal epidermis and oral mucosa. However, this cytokine was found throughout the epithelia in oral lichen planus, leukoplakia and squamous cell carcinoma (SCC). The possible cellular sources and biological effects of TNF in these disease processes is discussed. The 55-kDa TNFR was expressed at the cell membrane throughout the epidermis, but confined to the lower cell layers on oral mucosa. Ten of 22 SCC did not express this receptor. A significant correlation was observed between the velocity of tumour growth and absence of TNF receptors. The growth inhibitory and immunoregulatory effects of TNF may be modulated by changes in receptor expression and alterations in synthesis of this cytokine in a subgroup of oral SCC.

Intragastric nitric oxide production in humans: measurements in expelled air.

High values (800-6000 parts per billion) of nitric oxide (NO) in expelled air from the stomach were shown in humans by chemiluminescence technique. These NO values were more than 100 times higher than those found in orally exhaled air. Intragastric NO production is probably non-enzymatic, requiring an acidic environment, as NO in expelled air was reduced by 95% after pretreatment with the proton pump inhibitor omeprazole. Furthermore, large amounts of NO were formed in vitro from lettuce and saliva when placed in hydrogen chloride (pH < 2). In conclusion, large amounts of NO are formed intragastrically in humans and this source of NO may be of importance for the integrity of the gastric mucosa in health and disease. Measurements of NO in expelled air might be of value as a non-invasive method for estimation of gastric acidity.

Metabolism of large bowel mucosa in health and disease

Attitudes to colonic mucosal diseases have changed since the knowledge that the microflora act by its metabolites on the metabolic activity of the colonocytes, thereby influencing their functions and growth. The primary actions of the flora are twofold. They produce first metabolites which act as energy-producing substrates for the colonocytes, and second substances which may interfere with the enzymatic activities of the mucosa. Study of the action of microfloral metabolites on colonocyte metabolism and of the consequences following surgery offer an opportunity to obtain further information on the pathogenesis and therapy of large bowel diseases.