Jeremy C Parker1, Surendran Thavagnanam1, Grzegorz Skibinski1, Michael McBrien2, Liam G Heaney1, Michael D Shields3. 1. Centre for Infection and Immunity, Health Sciences Building, Queen's University Belfast, 97 Lisburn Road, Belfast BT9 7BL, Northern Ireland, United Kingdom. 2. Department of Anaesthetics, Belfast Health and Social Care Trust, Royal Victoria Hospital, Belfast BT12 6BE, Northern Ireland, United Kingdom. 3. Centre for Infection and Immunity, Health Sciences Building, Queen's University Belfast, 97 Lisburn Road, Belfast BT9 7BL, Northern Ireland, United Kingdom ; Department of Child Health, Royal Belfast Hospital for Sick Children, Belfast Health and Social Care Trust, Royal Victoria Hospital, Belfast BT12 6BE, Northern Ireland, United Kingdom.
Abstract
BACKGROUND: IL-31 is a novel cytokine that has been implicated in allergic diseases such as atopic dermatitis and more recently asthma. While IL-31 has been well studied in skin conditions such as atopic dermatitis, little is known about the role IL-31 plays in asthma and specifically the differentiation process of the bronchial epithelium, which is central to the pathogenesis of allergic asthma. METHODS: We examined the effects of IL-13 (20 ng/ml), IL-31 (20 ng/ml) and an IL-13/IL-31 combination stimulation (20 ng/ml each) on the in vitro mucociliary differentiation of paediatric bronchial epithelial cells (PBECs) from healthy patients (n=6). IL-31 receptor (IL-31-RA) expression, markers of differentiation (goblet and ciliated cells), transepithelial electrical resistance (TEER), quantification of goblet and ciliated cells, real time PCR for MUC5AC, ELISA for VEGF, EGF and MCP-1 (CCL-2) and ELISA for MUC5AC were assessed. RESULTS: We found that well-differentiated PBECs expressed IL-31-RA however it's expression did not increase upon stimulation with IL-31 or either of the other treatments. TEER indicated good formation of tight junctions which was found to be similar across all treatment groups (p=0.9). We found that IL-13 alone significantly reduced the number of ciliated cells compared with unstimulated (IL-13 stimuation: mean=4.8% (SD=2.5); unstimulated: mean=15.9%, (SD=7.4), p<0.01). IL-31 stimulation alone had no effect on ciliated cells whereas the IL-13/IL-31 combination stimulation significantly reduced the number of ciliated cells compared with control (IL-13/IL-31 combination: mean=5.1% (SD=4.6); unstimulated: mean=15.9%, (SD=7.4), p<0.01). We did not find that the combination of IL-13 and IL-31 had any additional effects to that of IL-13 alone. MUC5AC mRNA and secreted mucin was found in similar levels between unstimulated and all treatments, however IL-13 increased levels of MUC5AC mRNA by a factor of 2.84, albeit not significantly, compared with unstimulated cultures (IL-13 stimulation: mean=2.84 (SD=3.79); unstimulated: mean=1.0). CONCLUSIONS: IL-31RA receptor is present on well-differentiated paediatric bronchial epithelial cells. IL-31 does not exhibit any detrimental effects on mucociliary differentiation. IL-31 does not appear to have a synergistic effect when combined in culture with IL-13, in the differentiation process.
BACKGROUND:IL-31 is a novel cytokine that has been implicated in allergic diseases such as atopic dermatitis and more recently asthma. While IL-31 has been well studied in skin conditions such as atopic dermatitis, little is known about the role IL-31 plays in asthma and specifically the differentiation process of the bronchial epithelium, which is central to the pathogenesis of allergic asthma. METHODS: We examined the effects of IL-13 (20 ng/ml), IL-31 (20 ng/ml) and an IL-13/IL-31 combination stimulation (20 ng/ml each) on the in vitro mucociliary differentiation of paediatric bronchial epithelial cells (PBECs) from healthy patients (n=6). IL-31 receptor (IL-31-RA) expression, markers of differentiation (goblet and ciliated cells), transepithelial electrical resistance (TEER), quantification of goblet and ciliated cells, real time PCR for MUC5AC, ELISA for VEGF, EGF and MCP-1 (CCL-2) and ELISA for MUC5AC were assessed. RESULTS: We found that well-differentiated PBECs expressed IL-31-RA however it's expression did not increase upon stimulation with IL-31 or either of the other treatments. TEER indicated good formation of tight junctions which was found to be similar across all treatment groups (p=0.9). We found that IL-13 alone significantly reduced the number of ciliated cells compared with unstimulated (IL-13 stimuation: mean=4.8% (SD=2.5); unstimulated: mean=15.9%, (SD=7.4), p<0.01). IL-31 stimulation alone had no effect on ciliated cells whereas the IL-13/IL-31 combination stimulation significantly reduced the number of ciliated cells compared with control (IL-13/IL-31 combination: mean=5.1% (SD=4.6); unstimulated: mean=15.9%, (SD=7.4), p<0.01). We did not find that the combination of IL-13 and IL-31 had any additional effects to that of IL-13 alone. MUC5AC mRNA and secreted mucin was found in similar levels between unstimulated and all treatments, however IL-13 increased levels of MUC5AC mRNA by a factor of 2.84, albeit not significantly, compared with unstimulated cultures (IL-13 stimulation: mean=2.84 (SD=3.79); unstimulated: mean=1.0). CONCLUSIONS: IL-31RA receptor is present on well-differentiated paediatric bronchial epithelial cells. IL-31 does not exhibit any detrimental effects on mucociliary differentiation. IL-31 does not appear to have a synergistic effect when combined in culture with IL-13, in the differentiation process.
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