| 1. |
- Arike, Liisa, et al.
(författare)
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Protein Turnover in Epithelial Cells and Mucus along the Gastrointestinal Tract Is Coordinated by the Spatial Location and Microbiota
- 2020
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Ingår i: Cell Reports. - : Elsevier BV. - 2211-1247. ; 30:4, s. 1077-1087
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Tidskriftsartikel (refereegranskat)abstract
- The gastrointestinal tract is covered by a single layer of epithelial cells that, together with the mucus layers, protect the underlying tissue from microbial invasion. The epithelium has one of the highest turnover rates in the body. Using stable isotope labeling, high-resolution mass spectrometry, and computational analysis, we report a comprehensive dataset of the turnover of more than 3,000 and the expression of more than 5,000 intestinal epithelial cell proteins, analyzed under conventional and germ-free conditions across five different segments in mouse intestine. The median protein half-life is shorter in the small intestine than in the colon. Differences in protein turnover rates along the intestinal tract can be explained by distinct physiological and immune-related functions between the small and large intestine. An absence of microbiota results in an approximately 1 day longer protein half-life in germ-free animals.
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| 2. |
- Bos, Meike F., et al.
(författare)
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Goblet cell interactions reorient bundled mucus strands for efficient airway clearance
- 2023
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Ingår i: PNAS Nexus. - 2752-6542. ; 2:11
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Tidskriftsartikel (refereegranskat)abstract
- The respiratory tract of larger animals is cleared by sweeping bundled strands along the airway surface. These bundled strands can be millimetric in length and consist of MUC5B mucin. They are produced by submucosal glands, and upon emerging from these glands, the long axis of the bundled strands is oriented along the cilia-mediated flow toward the oral cavity. However, after release, the bundled strands are found to have turned orthogonal to the flow, which maximizes their clearance potential. How this unexpected reorientation is accomplished is presently not well understood. Recent experiments suggest that the reorientation process involves bundled strands sticking to MUC5AC mucus threads, which are tethered to the goblet cells. Such goblet cells are present in small numbers throughout the airway epithelium. Here, we develop a minimal model for reorientation of bundled mucus strands through adhesive interactions with surface goblet cells. Our simulations reveal that goblet cell interactions can reorient the bundled strands within 10 mm of release-making reorientation on the length scale of the tracheal tube feasible-and can stabilize the orthogonal orientation. Our model also reproduces other experimental observations such as strong velocity fluctuations and significant slow-down of the bundled strand with respect to the cilia-mediated flow. We further provide insight into the strand turning mechanism by examining the effect of strand shape on the impulse exerted by a single goblet cell. We conclude that goblet cell-mediated reorientation is a viable route for bundled strand reorientation, which should be further validated in future experiment.
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| 3. |
- Dolan, Brendan, et al.
(författare)
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Clearance of small intestinal crypts involves goblet cell mucus secretion by intracellular granule rupture and enterocyte ion transport
- 2022
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Ingår i: Science Signaling. - : American Association for the Advancement of Science (AAAS). - 1945-0877 .- 1937-9145. ; 15:752
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Tidskriftsartikel (refereegranskat)abstract
- Goblet cells in the small intestinal crypts contain large numbers of mucin granules that are rapidly discharged to clean bacteria from the crypt. Because acetylcholine released by neuronal and nonneuronal cells controls many aspects of intestinal epithelial function, we used tissue explants and organoids to investigate the response of the small intestinal crypt to cholinergic stimulation. The activation of muscarinic acetylcholine receptors initiated a coordinated and rapid emptying of crypt goblet cells that flushed the crypt contents into the intestinal lumen. Cholinergic stimulation induced an expansion of the granule contents followed by intracellular rupture of the mucin granules. The mucus expanded intracellularly before the rupture of the goblet cell apical membrane and continued to expand after its release into the crypt lumen. The goblet cells recovered from membrane rupture and replenished their stores of mucin granules. Mucus secretion from the goblet cells depended on Ca2+ signaling and the expansion of the mucus in the crypt depended on gap junctions and on ion and water transport by enterocytes adjacent to the goblet cells. This distinctive mode of mucus secretion, which we refer to as “expanding secretion,” efficiently cleans the small intestine crypt through coordinated mucus, ion, and fluid secretion by goblet cells and enterocytes.
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| 4. |
- Ermund, Anna, et al.
(författare)
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Mucus threads from surface goblet cells clear particles from the airways
- 2021
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Ingår i: Respiratory Research. - : Springer Science and Business Media LLC. - 1465-993X. ; 22:1
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Tidskriftsartikel (refereegranskat)abstract
- Background The mucociliary clearance system driven by beating cilia protects the airways from inhaled microbes and particles. Large particles are cleared by mucus bundles made in submucosal glands by parallel linear polymers of the MUC5B mucins. However, the structural organization and function of the mucus generated in surface goblet cells are poorly understood. Methods The origin and characteristics of different mucus structures were studied on live tissue explants from newborn wild-type (WT), cystic fibrosis transmembrane conductance regulator (CFTR) deficient (CF) piglets and weaned pig airways using video microscopy, Airyscan imaging and electron microscopy. Bronchoscopy was performed in juvenile pigs in vivo. Results We have identified a distinct mucus formation secreted from the surface goblet cells with a diameter less than two micrometer. This type of mucus was named mucus threads. With time mucus threads gathered into larger mucus assemblies, efficiently collecting particles. The previously observed Alcian blue stained mucus bundles were around 10 times thicker than the threads. Together the mucus bundles, mucus assemblies and mucus threads cleared the pig trachea from particles. Conclusions These results demonstrate that normal airway mucus is more complex and has a more variable structural organization and function than was previously understood. These observations emphasize the importance of studying young objects to understand the function of a non-compromised lung.
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| 5. |
- Fakih, Dalia, et al.
(författare)
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Normal murine respiratory tract has its mucus concentrated in clouds based on the Muc5b mucin
- 2020
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Ingår i: American Journal of Physiology-Lung Cellular and Molecular Physiology. - : American Physiological Society. - 1040-0605 .- 1522-1504. ; 318:6
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Tidskriftsartikel (refereegranskat)abstract
- The organization of the normal airway mucus system differs in small experimental animals from that in humans and large mammals. To address normal murine airway mucociliary clearance, Alcian blue-stained mucus transport was measured ex vivo on tracheal tissues of naive C57BL/6, Muc5b(-/-), Muc5ac(-/-), and EGFP-tagged Muc5b reporter mice. Close to the larynx with a few submucosal glands, the mucus appeared as thick bundles. More distally in the trachea and in large bronchi, Alcian blue-stained mucus was organized in cloud-like formations based on the Muc5b mucin. On tilted tissue, the mucus clouds moved upward toward the larynx with an average velocity of 12 mu m/s compared with 20 mu m/s for beads not associated with clouds. In Muc5ac(-/-) mice, Muc5b formed mucus strands attached to the tissue surface, while in Muc5b(-/-) mice, Muc5ac had a more variable appearance. The normal mouse lung mucus thus appears as discontinuous clouds, clearly different from the stagnant mucus layer in diseased lungs.
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| 6. |
- Giorgetti, Melania, et al.
(författare)
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New generation ENaC inhibitors detach cystic fibrosis airway mucus bundles via sodium/hydrogen exchanger inhibition
- 2021
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Ingår i: European Journal of Pharmacology. - : Elsevier BV. - 0014-2999. ; 904
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Tidskriftsartikel (refereegranskat)abstract
- Cystic fibrosis (CF) is a recessive inherited disease caused by mutations affecting anion transport by the epithelial ion channel cystic fibrosis transmembrane conductance regulator (CFTR). The disease is characterized by mucus accumulation in the airways and intestine, but the major cause of mortality in CF is airway mucus accumulation, leading to bacterial colonization, inflammation and respiratory failure. Several drug targets are under evaluation to alleviate airway mucus obstruction in CF and one of these targets is the epithelial sodium channel ENaC. To explore effects of ENaC inhibitors on mucus properties, we used two model systems to investigate mucus characteristics, mucus attachment in mouse ileum and mucus bundle transport in piglet airways. We quantified mucus attachment in explants from CFTR null (CF) mice and tracheobronchial explants from newborn CFTR null (CF) piglets to evaluate effects of ENaC or sodium/hydrogen exchanger (NHE) inhibitors on mucus attachment. ENaC inhibitors detached mucus in the CF mouse ileum, although the ileum lacks ENaC expression. This effect was mimicked by two NHE inhibitors. Airway mucus bundles were immobile in untreated newborn CF piglets but were detached by the therapeutic drug candidate AZD5634 (patent WO, 2015140527). These results suggest that the ENaC inhibitor AZD5634 causes detachment of CF mucus in the ileum and airway via NHE inhibition and that drug design should focus on NHE instead of ENaC inhibition.
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| 7. |
- Hoang, Oanh N, et al.
(författare)
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Mucins MUC5AC and MUC5B Are Variably Packaged in the Same and in Separate Secretory Granules.
- 2022
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Ingår i: American journal of respiratory and critical care medicine. - 1073-449X .- 1535-4970. ; 206:9, s. 1081-1095
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Tidskriftsartikel (refereegranskat)abstract
- Rationale: MUC5AC (mucin 5AC, oligomeric gel-forming) and MUC5B (mucin 5B, oligomeric gel-forming) are the predominant secreted polymeric mucins in mammalian airways. They contribute differently to the pathogenesis of various muco-obstructive and interstitial lung diseases, and their genes are separately regulated, but whether they are packaged together or in separate secretory granules is not known. Objectives: To determine the packaging of MUC5AC and MUC5B within individual secretory granules in mouse and human airways under varying conditions of inflammation and along the proximal-distal axis. Methods: Lung tissue was obtained from mice stimulated to upregulate mucin production by the cytokines IL-1β and IL-13 or by porcine pancreatic elastase. Human lung tissue was obtained from donated normal lungs, biopsy samples of transplanted lungs, and explanted lungs from subjects with chronic obstructive pulmonary disease. MUC5AC and MUC5B were labeled with antibodies from different animal species or, in mice only, by transgenic chimeric mucin-fluorescent proteins and imaged using widefield deconvolution or Airyscan fluorescence microscopy. Measurements and Main Results: In both mouse and human airways, most secretory granules contained both mucins interdigitating within the granules. Smaller numbers of granules contained MUC5B alone, and even fewer contained MUC5AC alone. Conclusions: MUC5AC and MUC5B are variably stored both in the same and in separate secretory granules of both mice and humans. The high fraction of granules containing both mucins under a variety of conditions makes it unlikely that their secretion can be differentially controlled as a therapeutic strategy. This work also advances knowledge of the packaging of mucins within secretory granules to understand mechanisms of epithelial stress in the pathogenesis of chronic lung diseases.
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| 8. |
- Jabbar, Karolina S., et al.
(författare)
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Association between Brachyspira and irritable bowel syndrome with diarrhoea
- 2021
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Ingår i: Gut. - : BMJ. - 0017-5749 .- 1468-3288. ; 70, s. 1117-1129
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Tidskriftsartikel (refereegranskat)abstract
- Objective: The incidence of IBS increases following enteric infections, suggesting a causative role for microbial imbalance. However, analyses of faecal microbiota have not demonstrated consistent alterations. Here, we used metaproteomics to investigate potential associations between mucus-resident microbiota and IBS symptoms. Design: Mucus samples were prospectively collected from sigmoid colon biopsies from patients with IBS and healthy volunteers, and their microbial protein composition analysed by mass spectrometry. Observations were verified by immunofluorescence, electron microscopy and real-Time PCR, further confirmed in a second cohort, and correlated with comprehensive profiling of clinical characteristics and mucosal immune responses. Results: Metaproteomic analysis of colon mucus samples identified peptides from potentially pathogenic Brachyspira species in a subset of patients with IBS. Using multiple diagnostic methods, mucosal Brachyspira colonisation was detected in a total of 19/62 (31%) patients with IBS from two prospective cohorts, versus 0/31 healthy volunteers (p<0.001). The prevalence of Brachyspira colonisation in IBS with diarrhoea (IBS-D) was 40% in both cohorts (p=0.02 and p=0.006 vs controls). Brachyspira attachment to the colonocyte apical membrane was observed in 20% of patients with IBS and associated with accelerated oro-Anal transit, mild mucosal inflammation, mast cell activation and alterations of molecular pathways linked to bacterial uptake and ion-fluid homeostasis. Metronidazole treatment paradoxically promoted Brachyspira relocation into goblet cell secretory granules-possibly representing a novel bacterial strategy to evade antibiotics. Conclusion: Mucosal Brachyspira colonisation was significantly more common in IBS and associated with distinctive clinical, histological and molecular characteristics. Our observations suggest a role for Brachyspira in the pathogenesis of IBS, particularly IBS-D. © Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.
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| 9. |
- Rodriguez-Piñeiro, Ana, et al.
(författare)
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Proteome of airway surface liquid and mucus in newborn wildtype and cystic fibrosis piglets
- 2023
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Ingår i: Respiratory Research. - : Springer Science and Business Media LLC. - 1465-9921 .- 1465-993X. ; 24
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Tidskriftsartikel (refereegranskat)abstract
- Background: The respiratory tract is protected from inhaled particles and microbes by mucociliary clearance, mediated by the mucus and the cilia creating a flow to move the mucus cephalad. Submucosal glands secrete linear MUC5B mucin polymers and because they pass through the gland duct before reaching the airway surface, bundled strands of 1000–5000 parallel molecules exit the glands. In contrast, the surface goblet cells secrete both MUC5AC and MUC5B. Methods: We used mass-spectrometry based proteomic analysis of unstimulated and carbachol stimulated newborn wild-type (WT) and cystic fibrosis transmembrane conductance regulator (CFTR) null (CF) piglet airways to study proteins in the airway surface liquid and mucus, to investigate if levels of MUC5AC and MUC5B were affected by carbachol stimulation and whether the proteins clustered according to function. Results: Proteins in the first four extracted fractions clustered together and the fifth fraction contained the mucus cluster, mucins and other proteins known to associate with mucins, whereas the traditional airway surface liquid proteins clustered to fraction 1–4 and were absent from the mucus fraction. Carbachol stimulation resulted in increased MUC5AC and MUC5B. Conclusions: These results indicate a distinct separation between proteins in the washable surface liquid and the mucus fraction. In fractions 1–4 from newborn CF piglets an additional cluster containing acute phase proteins was observed, suggesting an early inflammatory response in CF piglets. Alternatively, increased levels of these proteins could indicate altered lung development in the CF piglets. This observation suggests that CF airway disease is present at birth and thus, treatment should commence directly after diagnosis.
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