IL-4 and IL-13 both contribute to the homeostasis of human conjunctival goblet cells in vitro.
Ontology highlight
ABSTRACT:
SUBMITTER: Hansen PM
PROVIDER: S-EPMC9542340 | biostudies-literature | 2022 Aug
REPOSITORIES: biostudies-literature
Publications
IL-4 and IL-13 both contribute to the homeostasis of human conjunctival goblet cells in vitro.
Hansen Pernille M PM Tollenaere Maxim A X MAX Hedengran Anne A Heegaard Steffen S Amoudruz Petra P Røpke Mads M Thyssen Jacob P JP Kolko Miriam M Norsgaard Hanne H
Allergy 20220506 8
Similar Datasets
Project description:PurposeTo investigate the effects of IL-13 on goblet cell proliferation, differentiation, and expression of mucin and immunomodulatory genes.MethodsExplants were excised from the conjunctiva of young C57BL/6 mice. Cultures received 200 μL per week of either Keratinocyte media (KSFM) or KSFM supplemented with 10 ng/mL IL-13 and were incubated for 3 (D3), 7 (D7), or 14 (D14) days. Subsequently, cell proliferation was assessed or cultures were immunostained, collected for dot blot, or for reverse transcription (RT) and quantitative real-time PCR (qPCR) or for RT-PCR gene array.ResultsThe cultured conjunctival epithelium expressed goblet cell associated keratin 7 and mucins MUC5AC and MUC2 and when stimulated with IL-13 showed increased proliferation at D3 and D7 (P < 0.05) compared with control. MUC5AC expression was increased in the IL-13-treated group at D3 and D14 (P < 0.05). IL-13-treated cultures showed increased chemokine ligand 26 (CCL26), chloride channel calcium activated channel 3 (CLCA3), fas ligand (FasL), and Relm-β at D7. All conjunctival cultures expressed MUC2, and its expression was decreased at D3 (P < 0.05) and increased at D14 (P < 0.05) with IL-13 treatment.ConclusionsThis study demonstrated that conjunctival goblet cells are IL-13 responsive cells that produce factors known to maintain epithelial barrier, stimulate mucin production, and modulate immune response in nonocular mucosa when treated with IL-13. The functional significance of IL-13-stimulated factors remains to be determined.
| S-EPMC4495812 | biostudies-literature
Project description:Conjunctival goblet cells (GCs) are specialized epithelial cells that secrete mucins onto the ocular surface to maintain the wet environment. Assessment of GCs is important because various ocular surface diseases are associated with their loss. Although there are GC assessment methods available, the current methods are either invasive or difficult to use. In this report, we developed a simple and non-invasive GC assessment method based on fluorescence imaging. Moxifloxacin ophthalmic solution was used to label GCs via topical administration, and then various fluorescence microscopies could image GCs in high contrasts. Fluorescence imaging of GCs in the mouse conjunctiva was confirmed by both confocal reflection microscopy and histology with Periodic acid-Schiff (PAS) labeling. Real-time in-vivo conjunctival GC imaging was demonstrated in a rat model by using both confocal fluorescence microscopy and simple wide-field fluorescence microscopy. Different GC densities were observed in the forniceal and bulbar conjunctivas of the rat eye. Moxifloxacin based fluorescence imaging provides high-contrast images of conjunctival GCs non-invasively and could be useful for the study or diagnosis of GC related ocular surface diseases.
| S-EPMC6820527 | biostudies-literature
Project description:The angiotensin-converting enzyme 2 (ACE2) receptor has been proved for SARS-CoV-2 cell entry after auxiliary cellular protease priming by transmembrane protease serine 2 (TMPRSS2), but the co-effect of this molecular mechanism was unknown. Here, single-cell sequencing was performed with human conjunctiva and the results have shown that ACE2 and TMPRSS2 were highly co-expressed in the goblet cells with genes involved in immunity process. This identification of conjunctival cell types which are permissive to virus entry would help to understand the process by which SARS-CoV-2 infection was established. These finding might be suggestive for COVID-19 control and protection.
| S-EPMC7883706 | biostudies-literature
Project description:Solithromycin is a novel fluoroketolide antibiotic belonging to the class of macrolide antibiotics. Activation of the interleukin (IL)-13 receptor leads to STAT6 activation and subsequent induction of SAM pointed domain containing ETS transcription factor (SPDEF), chloride channel accessory 1 (CLCA1), and anoctamin-1 (ANO1), all of which are associated with the induction of MUC5AC. We examined the effects of solithromycin on mucin production led by IL-13 signaling. Normal human bronchial epithelial cells were grown at the air-liquid interface with IL-13 with/without solithromycin for 14 days. Histochemical analysis was performed using hematoxylin and eosin staining and MUC5AC immunostaining. MUC5AC, SPDEF, CLCA1, and ANO1 mRNA expressions were examined using real-time polymerase chain reaction. Western blot analysis was performed to assess CLCA1 and ANO1 proteins, and phosphorylation of STAT6 and ERK. Solithromycin attenuated IL-13 induction of goblet cell hyperplasia and MUC5AC, CLCA1 and ANO1 mRNA and protein expression induced by IL-13, but had no effect on the phosphorylation of STAT6 and ERK. Our results indicate that solithromycin could attenuate goblet cell hyperplasia and MUC5AC induced by IL-13 through inhibition of CLCA1 and ANO1 mRNA and protein expression. However, much more information is required to clarify the molecular mechanisms underlying the inhibition of CLCA1 and ANO1 by solithromycin.
| S-EPMC9854378 | biostudies-literature
Project description:Goblet cell metaplasia, a disabling hallmark of chronic lung disease, lacks curative treatments at present. To identify novel therapeutic targets for goblet cell metaplasia, we studied the transcriptional response profile of IL-13-exposed primary human airway epithelia in vitro and asthmatic airway epithelia in vivo. A perturbation-response profile connectivity approach identified geldanamycin, an inhibitor of heat shock protein 90 (HSP90) as a candidate therapeutic target. Our experiments confirmed that geldanamycin and other HSP90 inhibitors prevented IL-13-induced goblet cell metaplasia in vitro and in vivo. Geldanamycin also reverted established goblet cell metaplasia. Geldanamycin did not induce goblet cell death, nor did it solely block mucin synthesis or IL-13 receptor-proximal signaling. Geldanamycin affected the transcriptome of airway cells when exposed to IL-13, but not when exposed to vehicle. We hypothesized that the mechanism of action probably involves TGF-β, ERBB, or EHF, which would predict that geldanamycin would also revert IL-17-induced goblet cell metaplasia, a prediction confirmed by our experiments. Our findings suggest that persistent airway goblet cell metaplasia requires HSP90 activity and that HSP90 inhibitors will revert goblet cell metaplasia, despite active upstream inflammatory signaling. Moreover, HSP90 inhibitors may be a therapeutic option for airway diseases with goblet cell metaplasia of unknown mechanism.
| S-EPMC6355221 | biostudies-literature
Project description:We used cultured human conjunctival goblet cells to determine (i) whether the toxigenic S. aureus- induced activation of the epithelial goblet cells requires two signals to activate the NLRP3 inflammasome, (ii) if one signal is mediated by TLR1, TLR2, or TLR6, and (iii) if the S. aureus toxin α toxin is another signal for the activation of the inflammasome and secretion of mature IL-1β. Cultured cells were incubated with siRNA to knock down the different TLRs. After stimulation with toxigenic S. aureus RN6390, pro-IL-1β synthesis, caspase-1 activity, and mature IL-1β secretion were measured. In a separate set of experiments, the cells were incubated with toxigenic S. aureus RN6390 or mutant S. aureus ALC837 that does not express α toxin with or without exogenous α toxin. A gentamicin protection assay was used to determine if intracellular bacteria were active. We conclude that α toxin from toxigenic S. aureus triggers two separate mechanisms required for the activation of the NLRP3 inflammasome and secretion of mature IL-1β. In the first mechanism, α toxin secreted from internalized S. aureus produces a pore, allowing the internalized bacteria and associated pathogen-associated molecular patterns to interact with intracellular TLR2 and, to a lesser extent, TLR1. In the second mechanism, α toxin forms a pore in the plasma membrane, leading to an efflux of cytosolic K+ and influx of Ca2+. We conclude that α toxin by these two different mechanisms triggers the synthesis of pro-IL-1β and NLRP3 components, activation of capase-1, and secretion of mature IL-1β to defend against bacterial infection.
| S-EPMC10711068 | biostudies-literature
Project description:BackgroundIL-13 is a critical effector cytokine for allergic inflammation. It is produced by several cell types, including mast cells, basophils, and TH2 cells. In mast cells and basophils its induction can be stimulated by cross-linkage of immunoglobulin receptors or cytokines. The IL-1 family members IL-33 and IL-18 have been linked to induction of IL-13 production by mast cells and basophils. In CD4 TH2 cells IL-33-mediated production of IL-13 requires simultaneous signal transducer and activator of transcription (STAT) 5 activation.ObjectiveHere we have addressed whether cytokine-induced IL-13 production in mast cells and basophils follows the same logic as in TH2 cells: requirement of 2 separate signals.MethodsBy generating a bacterial artificial chromosome (BAC) transgenic IL-13 reporter mouse, we measured IL-13 production in mast cells and basophils.ResultsIn mast cells harvested from peritoneal cavities, 2 cytokine signals are required for IL-13 production: IL-33 and IL-3. In bone marrow mast cells IL-13 production requires IL-33, but the requirement for a STAT5 inducer is difficult to evaluate because these cells require the continuous presence of IL-3 (a STAT5 activator) for survival. Poorer STAT5 inducers in culture (IL-4 or stem cell factor) result in less IL-13 production on IL-33 challenge, but the addition of exogenous IL-3 enhances IL-13 production. This implies that bone marrow-derived mast cells, like peritoneal mast cells and TH2 cells, require stimulation both by an IL-1 family member and a STAT5 inducer to secrete IL-13. Basophils follow the same rule; splenic basophils produce IL-13 in response to IL-18 or IL-33 plus IL-3.ConclusionOptimal IL-13 production from mast cells and basophils requires 2 cytokine signals.
| S-EPMC3782850 | biostudies-literature
Project description:BackgroundIL-13 is a central mediator of airway responsiveness and mucus expression in patients with allergic airway inflammation, and IL-13 is currently a therapeutic target for asthma. However, little is known about how IL-13 regulates human CD4(+) T-cell lineages because IL-13 receptor (IL-13R) α1, a subunit of IL-13R, has not previously been reported to exist on human T cells.ObjectiveWe sought to determine whether human CD4(+) T(H)17 cells express IL-13Rα1 and whether IL-13 regulates T(H)17 cytokine production.MethodsNaive human CD4(+) cells were isolated from whole blood, activated with anti-CD3 and anti-CD28, and polarized to T(H)1, T(H)2, T(H)17, or induced regulatory T cells in the presence of IL-13 (0-10 ng/mL). Cell supernatants, total RNA, or total protein was examined 4 days after T(H)17 polarization.ResultsT(H)17 cells, but not T(H)0, T(H)1, T(H)2, or induced regulatory T cells, expressed IL-13Rα1. IL-13 attenuated IL-17A production, as well as expression of retinoic acid-related orphan receptor, runt-related transcription factor-1, and interferon regulatory factor 4 in T(H)17-polarized cells. IL-13 neither inhibited IFN-γ production from T(H)1 cells nor inhibited IL-4 production from T(H)2 cells. Furthermore, attenuation of IL-17A production only occurred when IL-13 was present within 24 hours of T-cell activation or at the time of restimulation.ConclusionsIL-13Rα1 is expressed on human CD4(+) T(H)17 cells, and IL-13 attenuates IL-17A production at polarization and restimulation. Although IL-13 is an attractive therapeutic target for decreasing symptoms associated with asthma, these results suggest that therapies inhibiting IL-13 production could have adverse side effects by increasing IL-17A production.
| S-EPMC3916096 | biostudies-literature
Project description:We previously reported anti-miR-328 therapy for dry eye disease (DED). Since decreased mucin secretion is a risk factor for DED, we aimed to explore whether anti-miR-328 affects mucin expression and goblet cells. MiR-328 was increased in goblet cells when they were under desiccating stress or treated with benzalkonium chloride (BAC), both of which are risk factors for DED. Based on bioinformatics tool results, miR-328 was predicted to directly target the transcription factor CREB1 that has been known to promote the expression of mucin5AC. The inhibitory effect of miR-328 on CREB1 was confirmed by the transfection assay. A miR-328 binding site on the CREB1 gene was confirmed by the luciferase assay. Furthermore, anti-miR-328 increased CREB1 and mucin5AC in cultured goblet cells according to qPCR, Western blot, and IF staining experiments. Anti-miR-328 increased mucin5AC secretion from the cultured goblet cells based on an ELISA assay for the cultured medium. Finally, impression cytology data revealed anti-miR-328 increased conjunctival goblet cells in the DED rabbits induced by BAC. In conclusion, anti-miR-328 increases CREB1 expression leading to an increase in mucin5AC production and secretion. Furthermore, anti-miR-328 also increases conjunctival goblet cells. These results warrant the further development of anti-miR-328 therapy for DED.
| S-EPMC9953597 | biostudies-literature
Project description:PurposeTo investigate the presence of uridine-5'-triphosphate (UTP)-activated P2Y1-like nucleotide receptors (P2Y2R, P2Y4R, and P2Y6R) in conjunctival goblet cells (CGCs) and determine if they increase intracellular Ca2+ concentration ([Ca2+]i) and induce mucin secretion.MethodsAdult, male rat conjunctiva was used for culture of CGCs. To investigate the expression of P2YRs, mRNA was extracted from CGCs and used for reverse transcription PCR (RT-PCR) with commercially obtained primers specific to P2Y2R, P2Y4R, and P2Y6R. Immunofluorescence (IF) and western blot (WB) analyses were performed using first-passage CGCs and stained with antibodies specific to each P2YR. Furthermore, CGCs were incubated with fura-2/AM, and [Ca2+]i was measured after stimulation with the P2YR selective agonists UTP, uridine 5'-diphosphate (UDP), or UDP-glucose and agonists specific to P2Y2R (MRS 2768), P2Y4R (MRS 4062), and P2Y6R (MRS 2693). [Ca2+]i measurements after P2Y2R and P2Y6R siRNA treatment were performed. Mucin secretion was measured after stimulation of P2Y2R, P2Y4R, and P2Y6R.ResultsmRNA for all pyrimidinergic P2Y1-like receptors was found as single bands of expected base pair number with RT-PCR. The presence of these P2YRs was confirmed with IF microscopy and WB analysis. UTP and UDP elicited concentration-dependent increases in [Ca2+]i. The receptor-specific agonists and UDP-glucose increased [Ca2+]i, although these responses were substantially lower than those elicited by UTP and UDP at 10-4 M and 10-3 M and did not show similar dose dependency. P2Y2R- and P2Y6R-depleted CGCs responded with reduced peak [Ca2+]i. UTP, MRS 2768 (P2Y2R), and UDP each stimulated mucin secretion from CGCs.ConclusionsP2Y2R, P2Y4R, and P2Y6R are present and functional in rat CGCs and may represent novel therapeutic targets for dry eye treatment and other types of ocular surface disease.
| S-EPMC11753474 | biostudies-literature