Enzymatic treatment of whey proteins in cow's milk results in differential inhibition of IgE-mediated mast cell activation compared to T-cell activation.

BACKGROUND: Cow's milk (CM) hydrolysates are frequently used as milk substitutes for children with CM allergy. In hydrolysates, allergenic epitopes within CM proteins are diminished by enzymatic treatment. The aim of this study was to examine the allergenic and immunogenic properties of whey proteins during hydrolysis. METHODS: During hydrolysis, samples were obtained at 0, 10, 15, 30, 45, 60, 75 and 90 min. Degradation was checked by HPLC and SDS-PAGE. Allergenic potential was analyzed by IgE crosslinking capacity of human Fcε receptor type 1-transduced rat basophilic leukemia cells sensitized with serum of CM-allergic patients. Whey-sensitized C3H/HeOuJ mice were ear challenged intracutaneously with the hydrolysates. Immunogenicity was tested using whey-specific human T-cell clones and T-cell lines at the level of proliferation and release of IL-4, IL-10, IL-13 and IFN-γ. RESULTS: After 15 min of hydrolysis, the majority of the proteins were degraded. Hydrolysis for 15 min resulted in 92% inhibition of mast cell degranulation and in 82% reduction of ear swelling in the mouse model. In contrast, T-cell-stimulatory capacity was less affected by hydrolysis: reduction of human T-cell proliferation was only 9%. This was further reduced to 57 and 74% after 30 and 45 min of hydrolysis, respectively. Cytokine production followed the pattern of T-cell proliferation. CONCLUSION: Via differential analysis of allergenic versus immunogenic properties of the time kinetics of hydrolysis of whey proteins, we have demonstrated specific hydrolysis conditions with reduced IgE-crosslinking responses but retained T-cell activating properties. This approach might be useful in better defining CM hydrolysates.

Cow's milk-specific T-cell reactivity of children with and without persistent cow's milk allergy: key role for IL-10.

BACKGROUND: The role of antigen-specific T cells in the mechanism of food allergy or maintenance of tolerance toward an innocuous antigen, such as cow's milk, is not yet fully understood. OBJECTIVE: The cow's milk-specific T-cell response of donors with various allergic backgrounds was investigated. METHODS: Cow's milk-specific T-cell clones (TCCs) were generated from the blood of children with persistent cow's milk allergy (CMA) and the blood of cow's milk-tolerant allergic and nonallergic control subjects. The TCCs were characterized by their antigen-specific proliferation, cytokine production, and activation status. RESULTS: Cow's milk-specific TCCs of children with persistent CMA were T(H)2 skewed, and the production of IL-4 and IL-13 was significantly correlated with the expression of the activation marker CD25. TCCs of the allergic control subjects were characterized by a high production of IL-10, which was positively correlated with the production of IL-4 and IFN-gamma and with the expression of CD25. TCCs derived from nonallergic control subjects had an attenuated response toward cow's milk in that they did not produce high levels of cytokines nor did they express high levels of surface markers. As in the allergic control subjects, in the nonallergic control subjects IL-10 production was positively correlated with the expression of CD25. CONCLUSION: The activation status of T cells derived from persistent donors with CMA was associated with the production of IL-4 and IL-13, whereas activated TCCs of cow's milk-tolerant control subjects were characterized by the production of IL-10 and, to a lesser extent, IFN-gamma. These findings suggest that activated CD4(+) T cells (characterized by a high CD25 expression) might contribute to the tolerogenic immune response toward an antigen, such as cow's milk, through the production of IL-10.

Cell-surface expression of CD25, CD26, and CD30 by allergen-specific T cells is intrinsically different in cow's milk allergy.

BACKGROUND: The release of T(H)2 cytokines by food-specific T cells is thought to be important in the etiology of food allergy. It has been suggested that the activation state of food-specific T cells also plays a significant role, but this has not yet been studied at the single-cell level. OBJECTIVE: Differences in the expression of cell-surface markers by cow's milk protein (CMP)-specific T cells between infants with and without cow's milk allergy (CMA) were evaluated at the clonal level. In addition, expression after the spontaneous development of tolerance of cow's milk in infants with CMA was analyzed. METHODS: We established CMP-specific T-cell clones (TCCs) from blood of infants with CMA and atopic dermatitis, from atopic controls with atopic dermatitis but without CMA, and from nonatopic controls. In addition, we established TCCs from infants with CMA after they had spontaneously developed tolerance to cow's milk. Expression levels of CD25, CD26, and CD30 by each TCC were analyzed by use of flow cytometry. RESULTS: Cow's milk protein-specific T cells from infants with CMA expressed much higher levels of CD25 and CD30 than CMP-specific T cells from infants without CMA. Expression of CD26 was much lower than in normal controls. After development of tolerance for cow's milk, expression of CD25 and CD30 was decreased, whereas the expression of CD26 was increased to normal levels. CONCLUSION: Antigen-specific T cells from patients with food allergy display an increased expression of cell-surface markers of activation compared with cells of patients without food allergy. This suggests an intrinsically stronger food-specific T-cell response in food-allergic patients, and points to the key role of food-specific T cells in the pathogenesis of food allergy.