Intestinal goblet cell mucus: isolation and identification by immunofluorescence of a goblet cell glycoprotein.
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Animals↗
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Density of goblet cells in the human Eustachian tube. Relationship between goblet-cell density in the tube and number of mucous glands in the middle ear.
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Autopsy↗
Activation of lamina propria T cells induces crypt epithelial proliferation and goblet cell depletion in cultured human fetal colon.
An organ culture model has been used to study the effects of T cell activation in the human colon. Lamina propria T cells in explant cultures of human fetal colon (11 to 23 weeks gestation) were activated in situ using pokeweed mitogen or an anti-CD3 monoclonal antibody, and compared with unstimulated controls. After three days of culture, there was a two to four-fold increase in crypt epithelial cell proliferation in T cell stimulated explants of more than 15 weeks gestation, associated with a fall in crypt goblet cell numbers of up to 20-fold. By three days, the surface epithelium of stimulated explants appeared thin with loss of goblet cells, and by day 7, severe and extensive mucosal damage was observed by light and electron microscopy. These changes did not occur in control cultures and explants deficient in T cells (less than 16 weeks gestation), and were inhibited by cyclosporin A. These experiments indicate that the increase in epithelial cell proliferation and accompanying goblet cell depletion observed in colorectal crypts in chronic inflammatory bowel disease may be mediated by activated T cells.
Cell Division↗
Isolation and characterization of cultured human conjunctival goblet cells.
PURPOSE: To isolate and characterize goblet cells from normal human conjunctival tissue to determine whether epidermal growth factor (EGF) receptors are present and whether EGF can influence goblet cell proliferation. METHODS: Goblet cells were isolated from explant cultures established from normal conjunctival tissue harvested from patients during periocular surgery. The cells were grown in RPMI culture medium supplemented with 10% fetal bovine serum and characterized using morphology, histochemistry, indirect immunofluorescence microscopy, molecular biology, and biochemistry. Proliferation was determined with a MTT proliferation assay after exposing goblet cells, which had been serum deprived for 48 hours, to increasing concentrations of epidermal growth factor (EGF; 0-80 ng/mL) for 24 hours. RESULTS: Goblet cells were isolated from conjunctival explants by scraping nongoblet cells from the culture dish. Human goblet cells exhibited positive reactivity with alcian blue-periodic acid Schiff (PAS) reagent, goblet cell-specific cytokeratin-7, HPA lectin, and MUC5AC, but negative reactivity to the stratified squamous epithelial cell marker, cytokeratin-4. The mRNA for MUC5AC was detected using RT-PCR. The presence of the EGF receptors EGFR, ErbB2, and ErbB3 was confirmed through Western blot analysis of cell lysates. EGF elicited a concentration-dependent increase in goblet cell proliferation of 160% +/- 0.5%, 188% +/- 0.45%, 293% +/- 1.3%, and 220% +/- 0.5% of control values with 10, 20, 40, and 80 ng/mL EGF, respectively. CONCLUSIONS: Human goblet cells that retain characteristics of goblet cells in vivo can be cultured. EGF receptors are present in human goblet cells, and EGF stimulates their proliferation.
Adult↗
Intratracheal priming with ovalbumin- and ovalbumin 323-339 peptide-pulsed dendritic cells induces airway hyperresponsiveness, lung eosinophilia, goblet cell hyperplasia, and inflammation.
Dendritic cells (DC) are the primary APC responsible for the capture of allergens in the airways and the shuttling of processed allergens to the draining lymph nodes where Ag presentation and T cell activation take place. The mechanism of this Ag handling and presentation in asthma is poorly understood. In addition, the feasibility of asthma induction by DC priming has not been directly tested. In this report an asthma model using intratracheally (i.t.) injected splenic DC was used to address these issues. DC pulsed with a model Ag OVA or the major MHC class II-restricted OVA T epitope peptide OVA(323-339) and instilled i.t. primed mice to exhibit asthma-like diseases. With OVA as the Ag, mice exhibit airway hyperresponsiveness (AHR), lung eosinophilia and inflammation, and pulmonary goblet cell hyperplasia. In OVA(323-339)-immunized mice, AHR and goblet cell hyperplasia were noted, with little eosinophilia and parenchymal inflammation. The latter finding provides evidence for dissociating AHR from eosinophilia. In both cases mediastinal node hypertrophy occurred, and high levels of Th2 cytokines were produced by the lung and mediastinal lymph node cells (LNC). Interestingly, mediastinal LNC also produced high levels of Th1 cytokines. Lung cells produced much less Th1 cytokines than these LNC. These results demonstrate that DC when introduced i.t. are potent in inducing asthma-like diseases by recruiting lymphocytes to the lung-draining lymph nodes and by stimulating Th2 responses and also suggest that the lung environment strongly biases T cell responses to Th2.
Animals↗
Mouse intestinal goblet cells expressing SV40 T antigen directed by the MUC2 mucin gene promoter undergo apoptosis upon migration to the villi.
Mucinous colorectal cancers exhibit a characteristic set of molecular genetic alterations and may be derived from progenitor cells committed to the goblet cell lineage. Previously, we demonstrated that the MUC2 mucin gene promoter drives transgene reporter expression with high specificity in small intestinal goblet cells of transgenic mice. On the basis of these experiments, we reasoned that the MUC2 promoter could be used to drive SV40 T antigen (Tag) expression in the same cell type, decoupling them from their normal antiproliferative controls. A line of mice was established (MUCTag6) that expressed Tag in intestinal goblet cells as determined by RNA blot and immunohistochemical analysis. These goblet cells were markedly involuted however, most notably in the villi. Endogenous intestinal MUC2 message levels were reduced to about one third the normal level in these mice. However, absorptive cell lineage markers were comparable with nontransgenics. Bromodeoxyuridine-positive S-phase cells are limited to crypts in nontransgenic intestine but are present in both crypts and villi in MUCTag6. In contrast, mitotic cells were not present in the villi, indicating that MUCTag6 villi goblet cells do not progress into M phase. Apoptotic cells positive for terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling were increased more than fourfold in MUCTag6 villi (P < 0.0001), and apoptotic goblet cells were evident. Electron microscopic examination of MUCTag6 intestinal villi revealed the presence of degraded cell remnants containing mucin goblets together with other cell debris, further indicating apoptosis of the goblet cell lineage. Thus, the expression of Tag in intestinal goblet cells releases them from normal antiproliferative controls, causing their inappropriate entry into S phase even after they transverse the crypt/villus junction. They do not, however, progress to M phase. Instead, they undergo apoptosis with a high degree of efficiency in S or G(2) phase. These experiments demonstrate that apoptosis effectively blocks inappropriate goblet cell proliferation in the intestine, supporting its proposed role as an antineoplastic mechanism.
Animals↗
The influence of different management systems and age on intestinal morphology, immune cell numbers and mucin production from goblet cells in post-weaning pigs.
At time of weaning, the immune system in piglets is not fully mature resulting in reduced growth and increased mortality. Early-weaned pigs transported to a segregated early weaning (SEW) facility have enhanced performance and gut development compared to conventional (CONV) pigs which may be due, in part, to decreased pathogen challenge. To gain further insight into SEW enhanced performance and gut development, gut samples from pigs weaned at 19+/-2 days were assessed during the post-weaning (PW) period. The numbers of cells expressing CD2, CD4, CD8, and CD172 (the 74-22-15 (SWC3) antibody is now known to be specific for CD172), MHC class II, and CD25 were quantified using immunohistochemistry. Additionally, samples of duodenum, jejunum, and ileum were evaluated for the production of neutral, acidic, and sulfuric mucins from goblet cells and morphological measurements were also made. No effects due to the management systems alone were observed for any of the parameters. However, there were interactive effects of age/time post-weaning and management system on the immune cells as well as on the mucin secreting goblet cells. There were no differences in gut morphology between SEW and CONV reared pigs.
Age Factors↗
Goblet cell number in the ileum of rats denervated during suckling and weaning.
The enteric nervous system plays a role on the stimulation of secretory cells of intestinal epithelia. We have demonstrated that ablation of ENS stimulates epithelial cell proliferation. As goblet cells are important constituents of the epithelial sheet, it is mandatory to investigate separately this cell type. The myenteric plexus of the ileum of rats in postnatal development was partially removed by the serosal application of benzalkonium chloride (BAC). Three groups of animals were used: those where BAC application was at 13 days and sacrifice was at 15 (13/28-day-old) or 23 days after treatment (13/36-day-old), and those where BAC was applied at 21 days and rats were killed 15 days after treatment (21/36-day-old) . The number of goblet cells in the ileum was estimated in sections stained by periodic acid-Schiff (PAS) histochemistry. In the 13/28 and 21/36 groups, the number of goblet cells was significantly higher after BAC treatment. These results suggest that the myenteric denervation may have an acute effect on the number of goblet cell in suckling and weanling rats, probably through submucous plexus.
Animals↗
Notch/gamma-secretase inhibition turns proliferative cells in intestinal crypts and adenomas into goblet cells.
The self-renewing epithelium of the small intestine is ordered into stem/progenitor crypt compartments and differentiated villus compartments. Recent evidence indicates that the Wnt cascade is the dominant force in controlling cell fate along the crypt-villus axis. Here we show a rapid, massive conversion of proliferative crypt cells into post-mitotic goblet cells after conditional removal of the common Notch pathway transcription factor CSL/RBP-J. We obtained a similar phenotype by blocking the Notch cascade with a gamma-secretase inhibitor. The inhibitor also induced goblet cell differentiation in adenomas in mice carrying a mutation of the Apc tumour suppressor gene. Thus, maintenance of undifferentiated, proliferative cells in crypts and adenomas requires the concerted activation of the Notch and Wnt cascades. Our data indicate that gamma-secretase inhibitors, developed for Alzheimer's disease, might be of therapeutic benefit in colorectal neoplastic disease.
Adenoma↗
[Further contribution on the fine structure of fetal goblet cells. Investigations on the small intestine of cattle].
Early in the fetal development, i.e. at the time both of the growth of intestinal villi and crypts and the epithelial cell differentiation the goblet cells also appear. The maturation of goblet cells progresses during their migration from the base of the villi respectively of the crypts to the villous top. As in the bovine large intestine there are vesicles and capsulated vacuoles, which contain vesicles, as single elements or as conglomerates in the immature goblet cells of the small intestine. These images deliver a scenario of the mechanism of secretory granule production and probably play a role in the formation of mucus.
Animals↗
Ca2+ dependency of 'Ca2+-independent' exocytosis in SPOC1 airway goblet cells.
SPOC1 airway goblet cells secrete mucin in response to P2Y2 receptor agonists and to secretagogues, phorbol 12-myristate 13-acetate (PMA) and ionomycin, which mobilize elements of the phospholipase C pathway, PKC and Ca2+, respectively. Previous studies demonstrated that mucin secretion from SLO-permeabilized, EGTA-buffered SPOC1 cells was stimulated by PMA at low Ca2+ levels (< 0.1 microm), consistent with the notion that regulated exocytosis may occur by Ca2+-independent pathways. We tested the alternative hypothesis that PMA-induced mucin secretion is, in fact, a Ca2+-dependent process under the conditions of low bulk Ca2+, one that is permitted in the typical SLO-permeabilized cell model by the slow binding kinetics of EGTA. Both IP3 and elevated bulk Ca2+ activated mucin secretion in SPOC1 cells buffered by EGTA, suggesting that IP3 generates a local Ca2+ gradient in the vicinity of the secretory granules to the degree necessary to trigger exocytosis. BAPTA, which binds Ca2+ approximately 100-fold faster than EGTA, diminished IP3-induced mucin release over a range of concentrations by > or = 69%, yet maintained an essentially normal mucin secretory response to elevated bulk Ca2+ in permeabilized SPOC1 cells. BAPTA also diminished the mucin secretory response of permeabilized cells to PMA, relative to the EGTA-buffered control: at PMA below 30 nm, BAPTA abolished the secretory response, and at higher concentrations it was reduced significantly relative to the EGTA-buffered controls. PMA-induced secretion in EGTA was insensitive to heparin. These results suggest that Ca2+ is released locally during PMA-induced exocytosis, by an IP3-independent mechanism.
Animals↗
Effects of streptozotocin-diabetes on rat intestinal mucin and goblet cells.
Intestinal mucin and goblet cells were examined in streptozotocin-diabetic rats and age-matched controls. Mucin (tissue content and secretion) was measured using a highly specific enzyme-linked immunoassay. In contrast to the increased protein to deoxyribonucleic acid ratio, an absolute decrease was observed in the mucin to deoxyribonucleic acid ratio in mucosal homogenates of the diabetic intestine. This was not due to a loss of goblet cells as their numbers per crypt-villus unit increased in diabetic rats (in proportion to the rise in enterocyte numbers and crypt-villus length). Histochemically, goblet cell mucin was unchanged in diabetes. After a 90-min incubation of everted intestinal segments in Krebs' buffer, pH 7.4, at 37 degrees C, the amount of mucin released into the medium was the same in diabetic and control rats when expressed relative to tissue deoxyribonucleic acid. However, secreted mucin represented a significantly larger proportion of the total tissue mucin content in diabetic animals. Thus, to maintain mucin output at normal levels, the rate of mucin secretion is apparently increased in the diabetic intestine, despite (or perhaps causing) a large decrease in the tissue mucin content.
Animals↗
MUC5B is the prominent mucin in human gallbladder and is also expressed in a subset of colonic goblet cells.
To elucidate the roles of human gallbladder mucin (HGBM), such as in gallstone formation and cytoprotection, it is essential to identify HGBM and study its expression. This was performed by metabolic labeling, Western blotting, immunohistochemistry, and RT-PCR. In a large number of individuals, antibodies against purified HGBM and against MUC5B detected a mucin precursor (approximately 470 kDa) in the gallbladder and colon, but not in the small intestine. In the gallbladder, Western blotting using specific anti-MUC5B antibodies showed that this mucin precursor represented an identical mucin, MUC5B. RT-PCR experiments demonstrated a similar tissue distribution pattern of MUC5B mRNA. Immunohistochemistry with anti-HGBM and anti-MUC5B showed staining in gallbladder epithelial cells and colonic goblet cells in the crypt base, but not in the small intestine; double labeling showed that HGBM was located in small granules within goblet cells, colocalizing to MUC2-containing goblet cells. Metabolic labeling demonstrated the secretion of mature MUC5B in the colon. Conclusively, MUC5B is identified as the prominent HGBM and is also expressed and secreted in the colon.
Blotting, Western↗
Colitogenic Environment Suppresses Notch/Mechanistic Target of Rapamycin-Mediated Epithelial Proliferation to Promote Goblet Cell Differentiation in the Colon.
Epithelial barrier dysfunction is a hallmark of inflammatory bowel diseases; however, the mechanisms underlying such impairment remain incompletely understood. In the present study, a dextran sulfate sodium-induced colitis model was used to investigate how the inflammatory environment damages the intestinal mucosa. The results demonstrated that colitogenic ambiance enhances intestinal epithelial cell death, delays epithelial cell proliferation, and exacerbates mucosal erosion. Unexpectedly, this work identified a previously unrecognized role for Notch signaling in mediating these effects. Specifically, the colitogenic milieu reduces Notch/mechanistic target of rapamycin complex 1 (mTORC1)-mediated intestinal epithelial cell proliferation to promote goblet cell differentiation. Chemical activation of Notch signaling stimulated intestinal epithelial cell proliferation and reduced goblet cell differentiation in the colitic mucosa, further aggravating mucosal damage. Conversely, inhibition of Notch or mTORC1 signaling during mucosal repair reduced intestinal epithelial cell proliferation and enhanced goblet cell differentiation, corroborating the implication of Notch and mTORC1 signaling in both processes. Collectively, these findings uncover a context-dependent role for the Notch-mechanistic target of rapamycin axis in regulating intestinal epithelial cell proliferation and differentiation in the colitic mucosa and suggest that its targeted modulation may hold therapeutic potential in inflammatory bowel diseases.
Animals↗
Elimination of IL-13 reverses established goblet cell metaplasia into ciliated epithelia in airway epithelial cell culture.
BACKGROUND: Interleukin (IL)-13 induces goblet cell metaplasia and plays an important role in mucus hypersecretion in asthma. We previously reported that IL-13 induced goblet cell differentiation along with less ciliated cell differentiation in guinea pig tracheal epithelial cells in vitro. In this study, we asked whether elimination of IL-13 could reverse the established goblet cell metaplasia into ciliated epithelia. METHODS: Primary epithelial cells from guinea pig tracheas were cultured at an air-liquid interface with the medium containing human recombinant IL-13 for 14 days, and continuously cultured with IL-13-eliminated medium, or cultured under the condition of neutralization of IL-13 with anti IL-13 antibody. RESULTS: 2 days after elimination of IL-13, the periodic acid-Schiff-positive area as well as MUC5AC protein level rapidly decreased. After 4 days, the number of goblet cells dramatically decreased, while that of ciliated cells inversely increased. The total number of epithelial cells did not change, and 5-bromo-2'-deoxyuridine uptake decreased after IL-13 elimination. Transitional cells with cilia and secretory granules increased after IL-13 elimination. Similarly, the neutralization of IL-13 with anti-IL-13 antibody for 5 days reversed the goblet cell metaplasia into ciliated epithelia, and transitional cells also increased. CONCLUSIONS: Elimination of IL-13 reverses goblet cell metaplasia into ciliated epithelia in vitro, and transition of goblet cells to other phenotypes, especially ciliated cells, may be involved in this phenomenon. IL-13 inhibition may be a therapeutic strategy of established goblet cell metaplasia in asthma.
Animals↗
PAF mediates cigarette smoke-induced goblet cell metaplasia in guinea pig airways.
Goblet cell metaplasia is an important morphological feature in the airways of patients with chronic airway diseases; however, the precise mechanisms that cause this feature are unknown. We investigated the role of endogenous platelet-activating factor (PAF) in airway goblet cell metaplasia induced by cigarette smoke in vivo. Guinea pigs were exposed repeatedly to cigarette smoke for 14 consecutive days. The number of goblet cells in each trachea was determined with Alcian blue-periodic acid-Schiff staining. Differential cell counts and PAF levels in bronchoalveolar lavage fluid were also evaluated. Cigarette smoke exposure significantly increased the number of goblet cells. Eosinophils, neutrophils, and PAF levels in bronchoalveolar lavage fluid were also significantly increased after cigarette smoke. Treatment with a specific PAF receptor antagonist, E-6123, significantly attenuated the increases in the number of airway goblet cells, eosinophils, and neutrophils observed after cigarette smoke exposure. These results suggest that endogenous PAF may play a key role in goblet cell metaplasia induced by cigarette smoke and that potential roles exist for inhibitors of PAF receptor in the treatment of hypersecretory airway diseases.
Animals↗
The effects of irradiation at different times of the day on rat intestinal goblet cells.
Quantitative changes in jejunal goblet cells were studied in control and whole body irradiated rats using PAS-Alcian blue staining of crypt sections. A circadian dependence was observed when control animals were killed at different times during the light/dark cycle. Irradiation with 3 Gy produced a 2-3-fold increase within 36 h in goblet cells relative to controls, followed by a reduction to very low levels. There was a return to pre-treatment levels later than was observed for the columnar cells. The present results on the pattern of response of goblet cells and those of brush border enzyme activity are consistent with the hypothesis that ionizing radiation can influence differentiation. In fact during the first hours after irradiation an early induction of differentiation is evident while during the early repopulation phase columnar cells prevailed relative to the goblet cells. Only at later times were normal differentiation patterns seen. Groups of animals exposed to the same dose of radiation at different times of the day showed similar general patterns of behaviour even if the group irradiated at midnight showed a more marked and longer lasting injury.
Animals↗
Distribution and density of conjunctival goblet cells.
There is a regional variation in conjunctival goblet cell distribution and density per unit of measurement. By secreting mucin goblet cells are involved in formation of tear film. Tear film instability is the basic sign of dry eye syndrome. Although dry eye syndrome can be a side effect of a disease, the most common cause is normal aging. The objective of the research was to determine goblet cell density in bulbar and lower forniceal conjunctiva and possible changes in goblet cell density related to age and sex. The research was conducted on 30 conjunctival samples of patients with no clinical changes of the conjunctiva. Tissue sections were stained with hematoxylin eosin (HE), periodic acid-schiff (PAS), alcian blue (AB)/PAS, Giemsa methods. Goblet cell density was 1.24 +/- 1.62 in bulbar conjunctiva and 30.21 +/- 14.32 in lower forniceal conjunctiva. Goblet cell distribution in the conjunctiva was unequal. A correlation between goblet cell density and age was not determined. Goblet cell density doesn't decrease with aging. No significant difference in goblet cell density between men and women was established.
Adult↗