Background-Animal models are essential for analyzing the allergenic potential of food proteins and for investigating mechanisms underlying food allergy. Based on previous studies revealing acid-suppression medication as risk factor for food allergy induction, we aimed to establish a mouse model mimicking the natural route of sensitization in patients.Methods-The effect of acid-suppressing medication on murine gastric pH was assessed by intragastric pH measurements after two injections of a proton pump inhibitor (PPI). To investigate dose-dependency, mice were fed different concentrations of ovalbumin (OVA; 0.2, 0.5, 1.0, 2.5 or 5.0 mg) either with or without anti-ulcer medication. Additionally, different routes of exposure (i.p. vs. oral) were compared in a second immunization experiment. Sera were screened for OVAspecific antibody titers (IgG1, IgG2a and IgE) in ELISA and RBL assay. Clinical reactivity was evaluated by measuring rectal temperature after oral challenge and by type I skin tests.Results-Two intravenous injections of PPI significantly elevated the gastric pH from 2.97 to 5.3. Only oral immunization with 0.2 mg OVA under anti-acid medication rendered elevated IgG1, IgG2a and IgE titers compared to all other concentrations. Protein feeding alone altered antibody titers only marginally. Even though also i.p. immunizations induced high levels of specific IgE, only oral immunizations under anti-acids induced anaphylactic reactions evidenced by a significant decrease of body temperature.Conclusion-Only low-dosage ovalbumin feedings under anti-acid medication resulted in IgE mediated food allergy. Based on this knowledge we have established a suitable food allergy model for further investigations of food adverse reactions.

We investigated the effects of 1α,25-dihydroxyvitamin D3[1,25(OH)2D3] on paracellular intestinal Ca2+absorption by determination of transepithelial electric resistance (TEER), as a measure of tight-junction ion permeability and bidirectional transepithelial45Ca2+fluxes in confluent Caco-2 cell cultures. The rise of TEER to steady-state levels of ∼2,000 Ω ⋅ cm2 was significantly attenuated by 1,25(OH)2D3(by up to 50%) in a dose-dependent fashion between 10−11 and 10−8 M. Synthetic analogs of 1,25(OH)2D3, namely, 1α,25-dihydroxy-16-ene,23-yne-vitamin D3 and 1α,25-dihydroxy-26,27-hexafluoro-16-ene,23-yne-vitamin D3, exhibited similar biopotency, whereas their genomically inactive 1-deoxy congeners were only marginally effective. Enhancement of transepithelial conductance of Caco-2 cell monolayers by vitamin D was accompanied by a significant increase in bidirectional transepithelial45Ca2+fluxes. Although 1,25(OH)2D3also induced cellular45Ca2+uptake from the apical aspect of Caco-2 cell layers and upregulated the expression of calbindin-9kDa mRNA, no significant contribution of the Ca2+-adenosinetriphosphatase-mediated transcellular pathway to transepithelial Ca2+ transport could be detected. Therefore stimulation of Ca2+fluxes across confluent Caco-2 cells very likely results from a genomic effect of vitamin D sterols on assembly and permeability of tight-junctional complexes.

Indirect evidence suggests that hot spices may interact with epithelial cells of the gastrointestinal tract to modulate their transport properties. Using HCT-8 cells, a cell line from a human ileocoecal carcinoma, we studied the effects of spices on transepithelial electrical resistance (TER), permeability for fluorescein isothiocyanate (FITC)-labeled dextrans with graded molecular weight, and morphological alterations of tight junctions by immunofluorescence using an anti-ZO-1 antibody, a marker for tight junction integrity. Two different reactivity patterns were observed: paprika and cayenne pepper significantly decreased the TER and increased permeability for 10-, 20- and 40-kDa dextrans but not for -70 kDa dextrans. Simultaneously, tight junctions exhibited a discontinuous pattern. Applying extracts from black or green pepper, bay leaf or nutmeg increased the TER and macromolecular permeability remained low. Immunofluorescence ZO-1 staining was preserved. In accordance with the above findings, capsaicin transiently reduced resistance and piperine increased resistance, making them candidates for causing the effects seen with crude spice extracts. The observation that Solanaceae spices (paprika, cayenne pepper) increase permeability for ions and macromolecules might be of pathophysiological importance, particularly with respect to food allergy and intolerance.

The major group human rhinovirus type 8 can enter cells via heparan sulphate. When internalized into ICAM-1 negative rhabdomyosarcoma (RD) cells, HRV8 accumulated in the cells but caused CPE only after 3 days when used at high MOI. Adaptation by three blind passages alternating between RD and HeLa cells resulted in variant HRV8v with decreased stability at acidic pH allowing for productive infection in the absence of ICAM-1. HRV8v produced CPE at 10 times lower MOI within 1 day. Confocal fluorescence microscopy colocalization and the use of pharmacological and dominant negative inhibitors revealed that viral uptake is clathrin, caveolin, and flotillin independent. However, it is blocked by dynasore, amiloride, and EIPA. Furthermore, HRV8v induced FITC-dextran uptake and colocalized with this fluid phase marker. Except for the complete inhibition by dynasore, the entry pathway of HRV8v via HS is similar to that of HRV14 in RD cells that overexpress ICAM-1.

Intercellular adhesion molecule 1 (ICAM-1) mediates binding and entry of major group human rhinoviruses (HRVs). Whereas the entry pathway of minor group HRVs has been studied in detail and is comparatively well understood, the pathway taken by major group HRVs is largely unknown. Use of immunofluorescence microscopy, colocalization with specific endocytic markers, dominant negative mutants, and pharmacological inhibitors allowed us to demonstrate that the major group virus HRV14 enters rhabdomyosarcoma cells transfected to express human ICAM-1 in a clathrin-, caveolin-, and flotillin-independent manner. Electron microscopy revealed that many virions accumulated in long tubular structures, easily distinguishable from clathrin-coated pits and caveolae. Virus entry was strongly sensitive to the Na ؉ /H ؉ ion exchange inhibitor amiloride and moderately sensitive to cytochalasin D. Thus, cellular uptake of HRV14 occurs via a pathway exhibiting some, but not all, characteristics of macropinocytosis and is similar to that recently described for adenovirus 3 entry via ␣ v integrin/CD46 in HeLa cells.