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TSLP Enhances Barrier Function in Human Conjunctival Epithelial Cells
AB126 Abstracts
475
SUNDAY
Plasma Thymic Stromal Lymphopoietin Concentration is related to Allergen Sensitization in Young Children G. R. Bloomberg1, S. Demehri1, C. M. Visness2, G. O’Connor3, M. Kattan4, R. Wood5, J. E. Gern6, R. Kopan1; 1Washington University School of Medicine, St. Louis, MO, 2Rho Federal Systems Division, Inc.,, Chapel Hill, NC, 3Boston University School of Medicine, Boston, MA, 4Columbia University School of Medicine, New York, NY, 5Johns Hopkins University School of Medicine, Baltimore, MD, 6University of Wisconsin School of Medicine and Public Health, Madison, WI. RATIONALE: Studies in mice have shown that elevated levels in blood of the cytokine, Thymic Stromal Lymphopoietin (TSLP), lead to allergen sensitization and airway hyperresponsiveness to allergen challenge. We completed a pilot study to measure plasma TSLP concentration in young children enrolled in the Urban Environment and Childhood Asthma (URECA) study, an inner-city birth cohort, and the relation to eczema, wheeze, and allergy. METHODS: Newborns of atopic parents, enrolled at birth, were followed through 3 years of age. Features examined were: eczema (EASI>1), any allergen-specific IgE > 0.35 kU/L, and recurrent wheezing episodes. We measured plasma TSLP levels in a pilot study of 50 children, 35 at 24 months of age, 15 at 36 months. (Three with samples at both time points.) Plasma TLSP concentration was measured by ELISA. Specific IgE for Der p, Der f, cat, dog, mouse, and Alternaria were available for the 36-month samples. RESULTS: TSLP was detectable in plasma in 23 of 50 children examined. Average TSLP concentrations were higher for children with detectable allergen-specific IgE, compared to those with undetectable specific IgE, for all allergens except Alternaria. These differences were significant (p<0.05) for milk, peanut, and cat allergens, and were borderline significant (p50.06) for mouse, Der p and Der f. Plasma TSLP concentration was not significantly related to the presence of eczema or recurrent wheezing. CONCLUSIONS: This pilot study demonstrated an association between plasma TSLP concentration and allergen sensitization in young children. We will undertake further study of this relationship in the full URECA cohort.
476
TSLP Enhances Barrier Function in Human Conjunctival Epithelial Cells E. B. Cook, J. L. Stahl, E. A. Schwantes, F. M. Graziano, S. K. Mathur; University Wisconsin, Madison, WI. RATIONALE: The conjunctival epithelium forms a barrier that isolates the eye from the external environment and regulates the active and passive movement of macromolecules, bacterial components, allergens and cells through both paracellular and transcellular pathways. Thymic stromal lymphopoietin (TSLP) is an epithelial-derived cytokine that plays a critical role in allergic diseases and has recently been shown to affect respiratory epithelial barrier function. The purpose of this study was to examine the effect of TSLP on human conjunctival epithelial cell (HCE) barrier function and mediator release. METHODS: For barrier function studies, HCE (IOBA-NHC cell line) were cultured (multi-layers) on cell culture inserts and stimulated with various concentrations of TSLP (30 min) after which horseradish peroxidase (HRP) was added to the upper compartment. Aliquots were collected from the basal compartment 60 min after HRP addition. Concentration of HRP in the basal compartment was measured fluorometrically using a commercially available kit. For mediator release, HCE were cultured in 24 well plates with various concentrations of TSLP for 48 hours and release of TNFa and IL-8 was measured by ELISA. RESULTS: TSLP stimulation decreased HRP permeability of HCE multilayers at 10 ng/ml (n54, p< 0.05), but had minimal effects at higher concentrations. However, stimulation of HCE with TSLP (10 - 1000 ng/ml) did not result in release of either TNFa or IL-8. CONCLUSIONS: TSLP may play a role in maintenance of conjunctival epithelial barrier integrity and function through regulation of permeability.
J ALLERGY CLIN IMMUNOL FEBRUARY 2011
477
Effect of Thymic Stromal Lymphopoietin (TSLP) on Cord Blood (CB) Progenitor Cell Hemopoietic Cytokine Receptors (HCR) D. Heroux1, I. Asher1, J. Denburg1, G. Delespesse2, Z. Allakhverdi2; 1 McMaster Universtiy, Hamilton, ON, CANADA, 2CRCHUM, NotreDame Hospital,, Montreal, QC, CANADA. RATIONALE: Since CB CD341 cells of atopic at-risk infants have been previously shown to express phenotypic alterations of HCR, we investigated the expression of IL-3Ra, IL-5Ra and GMCSFRa on these cells after overnight TSLP incubation. METHODS: Fresh and frozen CB cells, highly purified for CD 341 through a magnetic-activated cell sorting technique, were stimulated for 24 hours at varying doses of TSLP (0.01, 0.1, 1, and 10 ng/ml), with and without IL-33 (10ng/ml), then stained for surface expression of IL-3Ra, IL-5Ra and GMCSFRa. RESULTS: After overnight stimulation with TSLP, mean expression by CB CD341 cells of IL-5Ra, but not IL-3Ra or GM-CSFRa, was significantly enhanced by TSLP stimulation at 0.1 ng/mL TSLP (n58, p50.04). In addition, TSLP, combined with HCR, induced eosinophil/ basophil colony-forming cells in methylcellulose cultures. CONCLUSIONS: TSLP stimulation increases the expression of IL -5Ra on CB CD341 cells, and co-stimulation with TSLP promotes Eo/B differentiation. These findings may reflect an in utero pathway influencing CB progenitor cell eosinophilic lineage commitment, eventuating in allergic inflammation and disease in early life.
478
Changes in CXCL9, CXCL10 and CXCL11 Concentrations after Nasal Allergen Challenge D. Tworek1, M. Bochenska-Marciniak1, I. Kuprys-Lipinska1, Z. Kurmanowska1, W. Mlynarski2, P. Gorski1, P. Kuna1, A. Antczak1; 1Barlicki University Hospital, Medical University of Lodz, Lodz, POLAND, 2Medical University of Lodz, Lodz, POLAND. RATIONALE: The role of CXCL9, CXCL10 and CXCL11 in allergic inflammation has not been explored in details. Therefore we examined changes in concentrations of these chemokines in nasal lavages after nasal allergen challenge in patients allergic to grass pollen. METHODS: 21 subjects allergic to grass pollen and 19 non-allergic controls were included. All subjects underwent nasal allergen challenge outside the pollen season. Nasal lavages were collected before and 30 minutes after control solution (diluents for the allergen) and allergen administration. Concentrations of chemokines in nasal lavages were determined using ELISA. RESULTS: We observed significantly higher concentrations of CXCL10 in allergic patients in subsequent timepoints (354,496329,24 vs 164,626175,94; p50,036; 420,36421,28 vs 246,886353,24; p50,021 and 403,286359,29 vs 162,686148,69; p50,025; respectively). CXCL10 levels did not change 30 minutes after allergen provocation. In contrast, CXCL9 levels were similar in both groups before and after application of placebo. However, 30 minutes after allergen challenge significant rise in CXCL9 concentration was noticed in allergic patients (138,886109,59 vs 395,86301,2; p50,00026). CXCL11 concentrations increased after application of allergen in both groups, however it seems that it is rather the effect of preservative (phenol) than allergen itself as the significant rise in CXCL11 was also noticed after application of control solution. CONCLUSIONS: CXCL10 concentrations are increased in nasal lavages in allergic patients outside of pollen season and it seems that it may play a role in chronic allergic inflammation. CXCL9 levels increase rapidly after allergen application which may suggest its role in early allergic response. Results on CXCL11 concentrations remain inconclusive.
475
SUNDAY
Plasma Thymic Stromal Lymphopoietin Concentration is related to Allergen Sensitization in Young Children G. R. Bloomberg1, S. Demehri1, C. M. Visness2, G. O’Connor3, M. Kattan4, R. Wood5, J. E. Gern6, R. Kopan1; 1Washington University School of Medicine, St. Louis, MO, 2Rho Federal Systems Division, Inc.,, Chapel Hill, NC, 3Boston University School of Medicine, Boston, MA, 4Columbia University School of Medicine, New York, NY, 5Johns Hopkins University School of Medicine, Baltimore, MD, 6University of Wisconsin School of Medicine and Public Health, Madison, WI. RATIONALE: Studies in mice have shown that elevated levels in blood of the cytokine, Thymic Stromal Lymphopoietin (TSLP), lead to allergen sensitization and airway hyperresponsiveness to allergen challenge. We completed a pilot study to measure plasma TSLP concentration in young children enrolled in the Urban Environment and Childhood Asthma (URECA) study, an inner-city birth cohort, and the relation to eczema, wheeze, and allergy. METHODS: Newborns of atopic parents, enrolled at birth, were followed through 3 years of age. Features examined were: eczema (EASI>1), any allergen-specific IgE > 0.35 kU/L, and recurrent wheezing episodes. We measured plasma TSLP levels in a pilot study of 50 children, 35 at 24 months of age, 15 at 36 months. (Three with samples at both time points.) Plasma TLSP concentration was measured by ELISA. Specific IgE for Der p, Der f, cat, dog, mouse, and Alternaria were available for the 36-month samples. RESULTS: TSLP was detectable in plasma in 23 of 50 children examined. Average TSLP concentrations were higher for children with detectable allergen-specific IgE, compared to those with undetectable specific IgE, for all allergens except Alternaria. These differences were significant (p<0.05) for milk, peanut, and cat allergens, and were borderline significant (p50.06) for mouse, Der p and Der f. Plasma TSLP concentration was not significantly related to the presence of eczema or recurrent wheezing. CONCLUSIONS: This pilot study demonstrated an association between plasma TSLP concentration and allergen sensitization in young children. We will undertake further study of this relationship in the full URECA cohort.
476
TSLP Enhances Barrier Function in Human Conjunctival Epithelial Cells E. B. Cook, J. L. Stahl, E. A. Schwantes, F. M. Graziano, S. K. Mathur; University Wisconsin, Madison, WI. RATIONALE: The conjunctival epithelium forms a barrier that isolates the eye from the external environment and regulates the active and passive movement of macromolecules, bacterial components, allergens and cells through both paracellular and transcellular pathways. Thymic stromal lymphopoietin (TSLP) is an epithelial-derived cytokine that plays a critical role in allergic diseases and has recently been shown to affect respiratory epithelial barrier function. The purpose of this study was to examine the effect of TSLP on human conjunctival epithelial cell (HCE) barrier function and mediator release. METHODS: For barrier function studies, HCE (IOBA-NHC cell line) were cultured (multi-layers) on cell culture inserts and stimulated with various concentrations of TSLP (30 min) after which horseradish peroxidase (HRP) was added to the upper compartment. Aliquots were collected from the basal compartment 60 min after HRP addition. Concentration of HRP in the basal compartment was measured fluorometrically using a commercially available kit. For mediator release, HCE were cultured in 24 well plates with various concentrations of TSLP for 48 hours and release of TNFa and IL-8 was measured by ELISA. RESULTS: TSLP stimulation decreased HRP permeability of HCE multilayers at 10 ng/ml (n54, p< 0.05), but had minimal effects at higher concentrations. However, stimulation of HCE with TSLP (10 - 1000 ng/ml) did not result in release of either TNFa or IL-8. CONCLUSIONS: TSLP may play a role in maintenance of conjunctival epithelial barrier integrity and function through regulation of permeability.
J ALLERGY CLIN IMMUNOL FEBRUARY 2011
477
Effect of Thymic Stromal Lymphopoietin (TSLP) on Cord Blood (CB) Progenitor Cell Hemopoietic Cytokine Receptors (HCR) D. Heroux1, I. Asher1, J. Denburg1, G. Delespesse2, Z. Allakhverdi2; 1 McMaster Universtiy, Hamilton, ON, CANADA, 2CRCHUM, NotreDame Hospital,, Montreal, QC, CANADA. RATIONALE: Since CB CD341 cells of atopic at-risk infants have been previously shown to express phenotypic alterations of HCR, we investigated the expression of IL-3Ra, IL-5Ra and GMCSFRa on these cells after overnight TSLP incubation. METHODS: Fresh and frozen CB cells, highly purified for CD 341 through a magnetic-activated cell sorting technique, were stimulated for 24 hours at varying doses of TSLP (0.01, 0.1, 1, and 10 ng/ml), with and without IL-33 (10ng/ml), then stained for surface expression of IL-3Ra, IL-5Ra and GMCSFRa. RESULTS: After overnight stimulation with TSLP, mean expression by CB CD341 cells of IL-5Ra, but not IL-3Ra or GM-CSFRa, was significantly enhanced by TSLP stimulation at 0.1 ng/mL TSLP (n58, p50.04). In addition, TSLP, combined with HCR, induced eosinophil/ basophil colony-forming cells in methylcellulose cultures. CONCLUSIONS: TSLP stimulation increases the expression of IL -5Ra on CB CD341 cells, and co-stimulation with TSLP promotes Eo/B differentiation. These findings may reflect an in utero pathway influencing CB progenitor cell eosinophilic lineage commitment, eventuating in allergic inflammation and disease in early life.
478
Changes in CXCL9, CXCL10 and CXCL11 Concentrations after Nasal Allergen Challenge D. Tworek1, M. Bochenska-Marciniak1, I. Kuprys-Lipinska1, Z. Kurmanowska1, W. Mlynarski2, P. Gorski1, P. Kuna1, A. Antczak1; 1Barlicki University Hospital, Medical University of Lodz, Lodz, POLAND, 2Medical University of Lodz, Lodz, POLAND. RATIONALE: The role of CXCL9, CXCL10 and CXCL11 in allergic inflammation has not been explored in details. Therefore we examined changes in concentrations of these chemokines in nasal lavages after nasal allergen challenge in patients allergic to grass pollen. METHODS: 21 subjects allergic to grass pollen and 19 non-allergic controls were included. All subjects underwent nasal allergen challenge outside the pollen season. Nasal lavages were collected before and 30 minutes after control solution (diluents for the allergen) and allergen administration. Concentrations of chemokines in nasal lavages were determined using ELISA. RESULTS: We observed significantly higher concentrations of CXCL10 in allergic patients in subsequent timepoints (354,496329,24 vs 164,626175,94; p50,036; 420,36421,28 vs 246,886353,24; p50,021 and 403,286359,29 vs 162,686148,69; p50,025; respectively). CXCL10 levels did not change 30 minutes after allergen provocation. In contrast, CXCL9 levels were similar in both groups before and after application of placebo. However, 30 minutes after allergen challenge significant rise in CXCL9 concentration was noticed in allergic patients (138,886109,59 vs 395,86301,2; p50,00026). CXCL11 concentrations increased after application of allergen in both groups, however it seems that it is rather the effect of preservative (phenol) than allergen itself as the significant rise in CXCL11 was also noticed after application of control solution. CONCLUSIONS: CXCL10 concentrations are increased in nasal lavages in allergic patients outside of pollen season and it seems that it may play a role in chronic allergic inflammation. CXCL9 levels increase rapidly after allergen application which may suggest its role in early allergic response. Results on CXCL11 concentrations remain inconclusive.