Different mucins are produced by the surface epithelium and the submucosa in human trachea: identification of MUC5AC as a major mucin from the goblet cells.
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ABSTRACT:
SUBMITTER: Hovenberg HW
PROVIDER: S-EPMC1217624 | biostudies-other | 1996 Aug
REPOSITORIES: biostudies-other
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Project description:To understand the mucociliary clearance system, mucins were visualized by light, confocal and electron microscopy, and mucus was stained by Alcian blue and tracked by video microscopy on tracheal explants of newborn piglets. We observed long linear mucus bundles that appeared at the submucosal gland openings and were transported cephalically. The mucus bundles were shown by mass spectrometry and immunostaining to have a core made of MUC5B mucin and were coated with MUC5AC mucin produced by surface goblet cells. The transport speed of the bundles was slower than the airway surface liquid flow. We suggest that the goblet cell MUC5AC mucin anchors the mucus bundles and thus controls their transport. Normal clearance of the respiratory tree of pigs and humans, both rich in submucosal glands, is performed by thick and long mucus bundles.
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Project description:An antibody (PGM2B) recognizing a pig gastric-mucin apoprotein reacts with the surface epithelium of pig gastric mucosa. Virtually no reactivity was observed over the mucin-producing cells in the glands, which were recognized by the GlcNAc-selective Griffonia simplicifolia II (GSA-II) lectin. Mucins from the glandular tissue of the cardiac region, corpus and antrum were purified using isopycnic density-gradient centrifugation in CsCl/guanidinium chloride. In the cardiac region, two major mucin populations at 1.5 and 1.4 g/ml were identified. The high-density population reacted preferentially with the PGM2B antibody and resembled mucins from the surface epithelium of this region, whereas the low-density population reacted strongly with the GSA-II lectin and appeared to originate from the glands. In the glandular tissue of corpus, a component with strong GSA-II lectin reactivity, which was distinctly different from the surface mucins from this region, was found at 1.4 g/ml, thus resembling the gland component from the cardiac region. Mucins from antrum glandular tissue contained at least two GSA-II lectin-reactive populations banding at 1.5 and 1.4 g/ml, respectively. Gland mucins from all regions were large oligomeric glycoproteins and heterogeneous with respect to charge properties, as shown by using rate-zonal centrifugation and ion-exchange HPLC, respectively. Gel chromatography of mucin glycopeptides showed that gland mucins from antrum and corpus contained significantly longer glycosylated domains than those from the surface mucosa. Thus, mucins from pig gastric glandular tissue comprise a number of large and oligomeric glycoproteins that differ from those from the surface epithelium in buoyant density, apoprotein structure and carbohydrate substitution.
| S-EPMC1219406 | biostudies-other
Project description:Bovine trachea in organ culture secretes mucus containing a 'high-density' (1.46 g/ml) and a 'low-density' (1.37 g/ml) mucin similar to those identified previously in bovine respiratory secretions [Hovenberg, Carlstedt and Davies (1997) Biochem. J. 321, 117-123]. After pulse-labelling, autoradiography showed uptake of [35S]sulphate by both epithelial goblet cells and submucosal glands, while [3H]proline was mainly incorporated into the ciliated surface epithelial cells. After 24 h of radiolabelling, neither the high- nor the low-density mucin in the secreted mucus gel was heavily radiolabelled with the precursors. In contrast, a population of molecules banding at 1.50 g/ml was heavily radiolabelled with [35S]sulphate. This component was smaller than the high-density mucin from the mucus gel and was insensitive to reduction or digestion with chondroitin ABC lyase or heparan sulphate lyase. The molecules yielded two populations of high-Mr glycopeptides upon trypsin digestion, were sensitive to keratanase and endo-beta-galactosidase digestion and contained O-linked glycans. Extracts of the surface epithelium and submucosal tissue after radiolabelling showed that the high- and low-density mucins in the tissue were also poorly radiolabelled. Thus, under these conditions, the radiolabelled precursors were not effectively incorporated into the large oligomeric mucins but into a high-Mr monomeric species. This study suggests that data obtained in investigations where mucins are radiolabelled and studied without further separation into distinct components may rather reflect the turnover of this 'novel' monomeric species than the large oligomeric mucins.
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Project description:ObjectiveCystic fibrosis (CF) is a genetic condition that causes abnormal mucus secretions in affected organs. MUC5AC and MUC5B are gel-forming mucins and frequent targets for investigations in CF tissues. Our objective was to qualify MUC5AC and MUC5B immunohistochemical techniques to provide a useful tool to identify, localize and interpret mucin expression in ferret tissues.ResultsMUC5AC and MUC5B mucins were detected most commonly in large airways and least in small airways, consistent with reported goblet cell density in airway surface epithelia. We evaluated whether staining method affected the detection of goblet cell mucins in serial sections of bronchial surface epithelia. Significant differences between stains were not observed suggesting common co-expression MUC5AC and MUC5B proteins in goblet cells of airway surface epithelia. Gallbladder and stomach tissues are reported to have differential mucin enrichment, so we tested these tissues in wildtype ferrets. Stomach tissues were enriched in MUC5AC and gallbladder tissues enriched in MUC5B, mucin enrichment similar to human tissues. Mucin immunostaining techniques were further qualified for specificity using lung tissue from recently generated MUC5AC-/- and MUC5B-/- ferrets. Qualified techniques for MUC5AC and MUC5B immunohistochemistry will be useful tools for mucin tissue studies in CF and other ferret models.
| S-EPMC10286488 | biostudies-literature
Project description:Some mucin genes have been detected during human embryonic and fetal organ development; however, little is known about mucin expression in epidermal development, neither in humans nor in other species. The present research was developed to explore Muc5ac skin expression during prenatal and postnatal rat development. Immunohistochemistry (IHC), Western blotting (WB) and RT-PCR were employed. By IHC, Muc5ac protein was found early in embryonic epidermis from day 13 of gestation until seven days after birth when the surface epidermis became negative and the reaction was restricted to secreting sebum cells. In coincidence with IHC findings, WB analysis showed a band at approximately 200KDa at the same periods of development. Results were also confirmed by RT-PCR. Muc5ac expression in rat embryonic epidermis suggests that Muc5ac may play a protective role in embryonic skin previous to birth which may be replaced by pile covering. To our knowledge, this is the first report which confirmed Muc5ac expression during skin development. Muc5ac expression in rat embryonic epidermis suggests that Muc5ac may play a protective role in embryonic skin previous to birth which may be replaced by pile covering. To our knowledge, this is the first report which confirmed Muc5ac expression during skin development.
| S-EPMC4378216 | biostudies-literature
Project description:The secreted mucins MUC5AC and MUC5B are large glycoproteins that play critical defensive roles in pathogen entrapment and mucociliary clearance. Their respective genes contain polymorphic and degenerate protein-coding variable number tandem repeats (VNTRs) that make the loci difficult to investigate with short reads. We characterize the structural diversity of MUC5AC and MUC5B by long-read sequencing and assembly of 206 human and 20 nonhuman primate (NHP) haplotypes. We find that human MUC5B is largely invariant (5,761-5,762 amino acids [aa]); however, seven haplotypes have expanded VNTRs (6,291-7,019 aa). In contrast, 30 allelic variants of MUC5AC encode 16 distinct proteins (5,249-6,325 aa) with cysteine-rich domain and VNTR copy-number variation. We group MUC5AC alleles into three phylogenetic clades: H1 (46%, ∼5,654 aa), H2 (33%, ∼5,742 aa), and H3 (7%, ∼6,325 aa). The two most common human MUC5AC variants are smaller than NHP gene models, suggesting a reduction in protein length during recent human evolution. Linkage disequilibrium and Tajima's D analyses reveal that East Asians carry exceptionally large blocks with an excess of rare variation (p < 0.05) at MUC5AC. To validate this result, we use Locityper for genotyping MUC5AC haplogroups in 2,600 unrelated samples from the 1000 Genomes Project. We observe a signature of positive selection in H1 among East Asians and a depletion of the likely ancestral haplogroup (H3). In Europeans, H3 alleles show an excess of common variation and deviate from Hardy-Weinberg equilibrium (p < 0.05), consistent with heterozygote advantage and balancing selection. This study provides a generalizable strategy to characterize complex protein-coding VNTRs for improved disease associations.
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Project description:RationaleMUC5AC and MUC5B are the predominant gel-forming mucins in the mucus layer of human airways. Each mucin has distinct functions and site-specific expression. However, the regional distribution of expression and cell types that secrete each mucin in normal/healthy human airways are not fully understood.ObjectivesTo characterize the regional distribution of MUC5B and MUC5AC in normal/healthy human airways and assess which cell types produce these mucins, referenced to the club cell secretory protein (CCSP).MethodsMultiple airway regions from 16 nonsmoker lungs without a history of lung disease were studied. MUC5AC, MUC5B, and CCSP expression/colocalization were assessed by RNA in situ hybridization and immunohistochemistry in five lungs with histologically healthy airways. Droplet digital PCR and cell cultures were performed for absolute quantification of MUC5AC/5B ratios and protein secretion, respectively.Measurements and main resultsSubmucosal glands expressed MUC5B, but not MUC5AC. However, MUC5B was also extensively expressed in superficial epithelia throughout the airways except for the terminal bronchioles. Morphometric calculations revealed that the distal airway superficial epithelium was the predominant site for MUC5B expression, whereas MUC5AC expression was concentrated in proximal, cartilaginous airways. RNA in situ hybridization revealed MUC5AC and MUC5B were colocalized with CCSP-positive secretory cells in proximal superficial epithelia, whereas MUC5B and CCSP-copositive cells dominated distal regions.ConclusionsIn normal/healthy human airways, MUC5B is the dominant secretory mucin in the superficial epithelium and glands, with distal airways being a major site of expression. MUC5B and MUC5AC expression is a property of CCSP-positive secretory cells in superficial airway epithelia.
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Project description:In asthma, airflow obstruction is thought to result primarily from inflammation-triggered airway smooth muscle (ASM) contraction. However, anti-inflammatory and smooth muscle-relaxing treatments are often temporary or ineffective. Overproduction of the mucin MUC5AC is an additional disease feature that, while strongly associated pathologically, is poorly understood functionally. Here we show that Muc5ac is a central effector of allergic inflammation that is required for airway hyperreactivity (AHR) to methacholine (MCh). In mice bred on two well-characterized strain backgrounds (C57BL/6 and BALB/c) and exposed to two separate allergic stimuli (ovalbumin and Aspergillus extract), genetic removal of Muc5ac abolishes AHR. Residual MCh responses are identical to unchallenged controls, and although inflammation remains intact, heterogeneous mucous occlusion decreases by 74%. Thus, whereas inflammatory effects on ASM alone are insufficient for AHR, Muc5ac-mediated plugging is an essential mechanism. Inhibiting MUC5AC may be effective for treating asthma and other lung diseases where it is also overproduced.
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