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Negative regulation of immunoglobulin E-dependent allergic responses by Lyn kinase.

Odom S, Gomez G, Kovarova M, Furumoto Y, Ryan JJ, Wright HV, Gonzalez-Espinosa C, Hibbs ML, Harder KW, Rivera J - J. Exp. Med. (2004)

Bottom Line: Contrary to this belief, Lyn kinase was found to have an important role as a negative regulator of the allergic response.This became apparent from the hyperresponsive degranulation of lyn-/- bone marrow-derived mast cells, which is driven by hyperactivation of Fyn kinase that occurs, in part, through the loss of negative regulation by COOH-terminal Src kinase (Csk) and the adaptor, Csk-binding protein.This was mirrored by increased circulating histamine, increased mast cell numbers, increased cell surface expression of the high affinity IgE receptor (FcepsilonRI), and eosinophilia.

View Article: PubMed Central - PubMed

Affiliation: Molecular Inflammation Section, Molecular Immunology and Inflammation Branch, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
A role for Lyn kinase as a positive regulator of immunoglobulin (Ig)E-dependent allergy has long been accepted. Contrary to this belief, Lyn kinase was found to have an important role as a negative regulator of the allergic response. This became apparent from the hyperresponsive degranulation of lyn-/- bone marrow-derived mast cells, which is driven by hyperactivation of Fyn kinase that occurs, in part, through the loss of negative regulation by COOH-terminal Src kinase (Csk) and the adaptor, Csk-binding protein. This phenotype is recapitulated in vivo as young lyn-/- mice showed an enhanced anaphylactic response. In vivo studies also demonstrated that as lyn-/- mice aged, their serum IgE increased as well as occupancy of the high affinity IgE receptor (FcepsilonRI). This was mirrored by increased circulating histamine, increased mast cell numbers, increased cell surface expression of the high affinity IgE receptor (FcepsilonRI), and eosinophilia. The increased IgE production was not a consequence of increased Fyn kinase activity in lyn-/- mice because both lyn-/- and lyn-/- fyn-/- mice showed high IgE levels. Thus, lyn-/- mice and mast cells thereof show multiple allergy-associated traits, causing reconsideration of the possible efficacy in therapeutic targeting of Lyn in allergic disease.

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Lyn−/− mice have increased numbers of peritoneal mast cells with increased cell surface FcεRI expression and eosinophilia. Fyn is not required for increased serum IgE. (a) Cells from a peritoneal lavage of wt or lyn−/− mice were stained with antibody to KIT and IgE after incubation with IgE to ensure FcεRI saturation. As a negative control, isotype-specific IgG was also used to stain the cells. Inset numbers reflect the percentage of KIT/FcεRI+ cells. (b) The percentage of mast cells in the peritoneum and the mean fluorescence intensity of FITC-IgE staining (reflecting mast cell FcεRI expression) of all the mice analyzed (n = 10). (c) Wright-Giemsa stain of peritoneal lavage cells from wt or lyn−/− mice. Mast cells are indicated by intense granule staining (dark cells, marked with arrow head), whereas eosinophils show a light red/pink stain and multilobed/segmented nucleus (arrow, see inset in lyn−/− sample). Graph is a quantitation of the percentage of eosinophils found in the peritoneal lavage of all wt and lyn−/− mice analyzed (n = 10). (d) The serum IgE concentration of the indicated genotypes was measured by ELISA from eight sex- and age-matched mice as described in Materials and Methods. Mean of the observed IgE concentrations for lyn−/− or lyn−/− fyn−/− mice is indicated by the line. No significant difference was found between these two populations, but both were significantly different from wt or fyn−/− mice. Significance was determined by an unpaired t test. ***, a p-value of ≤0.006; *, a p-value of ≤0.03.
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fig6: Lyn−/− mice have increased numbers of peritoneal mast cells with increased cell surface FcεRI expression and eosinophilia. Fyn is not required for increased serum IgE. (a) Cells from a peritoneal lavage of wt or lyn−/− mice were stained with antibody to KIT and IgE after incubation with IgE to ensure FcεRI saturation. As a negative control, isotype-specific IgG was also used to stain the cells. Inset numbers reflect the percentage of KIT/FcεRI+ cells. (b) The percentage of mast cells in the peritoneum and the mean fluorescence intensity of FITC-IgE staining (reflecting mast cell FcεRI expression) of all the mice analyzed (n = 10). (c) Wright-Giemsa stain of peritoneal lavage cells from wt or lyn−/− mice. Mast cells are indicated by intense granule staining (dark cells, marked with arrow head), whereas eosinophils show a light red/pink stain and multilobed/segmented nucleus (arrow, see inset in lyn−/− sample). Graph is a quantitation of the percentage of eosinophils found in the peritoneal lavage of all wt and lyn−/− mice analyzed (n = 10). (d) The serum IgE concentration of the indicated genotypes was measured by ELISA from eight sex- and age-matched mice as described in Materials and Methods. Mean of the observed IgE concentrations for lyn−/− or lyn−/− fyn−/− mice is indicated by the line. No significant difference was found between these two populations, but both were significantly different from wt or fyn−/− mice. Significance was determined by an unpaired t test. ***, a p-value of ≤0.006; *, a p-value of ≤0.03.

Mentions: As stated above, increased concentrations of monomeric IgE were shown to increase mast cell survival, but these conditions also resulted in increased cell surface expression of mast cell FcεRI (for review see reference 12), two factors associated with increased allergic responses. Therefore, we investigated whether these factors might be observed in 7-wk-old lyn−/− mice given their increased serum IgE. Fig. 6 a shows that mast cells in a peritoneal lavage of lyn−/− mice are increased, compared with wild-type mice of the same age. Analysis on all tested mice revealed that peritoneal mast cells of wild-type mice comprised, on average, 2.2% of the total cells in the peritoneal lavage (Fig. 6 b). In contrast, lyn−/− mice showed, on average, ∼5.3% as mast cells (ranging from a low of 2.1 to a high of ∼11%). Regardless of whether the numbers of peritoneal mast cells in a given mouse were enhanced or not, all mast cells showed increased cell surface FcεRI expression (Fig. 6 b). It should be noted that lyn−/− mice also showed increased numbers of dermal mast cells, with 8.0 ± 0.4 mast cells per field (a magnification of 1,000) versus wild-type mice, which had 4.5 ± 0.3 mast cells (P ≤ 0.001; not depicted). Importantly, 4-wk-old wild-type and lyn−/− mice did not differ significantly in peritoneal mast cell numbers with 1.3 ± 0.4 and 1.7 ± 0.3% of total cells as mast cells, respectively (not depicted), even though the lyn−/− mice showed a slight trend toward increased mast cell numbers.


Negative regulation of immunoglobulin E-dependent allergic responses by Lyn kinase.

Odom S, Gomez G, Kovarova M, Furumoto Y, Ryan JJ, Wright HV, Gonzalez-Espinosa C, Hibbs ML, Harder KW, Rivera J - J. Exp. Med. (2004)

Lyn−/− mice have increased numbers of peritoneal mast cells with increased cell surface FcεRI expression and eosinophilia. Fyn is not required for increased serum IgE. (a) Cells from a peritoneal lavage of wt or lyn−/− mice were stained with antibody to KIT and IgE after incubation with IgE to ensure FcεRI saturation. As a negative control, isotype-specific IgG was also used to stain the cells. Inset numbers reflect the percentage of KIT/FcεRI+ cells. (b) The percentage of mast cells in the peritoneum and the mean fluorescence intensity of FITC-IgE staining (reflecting mast cell FcεRI expression) of all the mice analyzed (n = 10). (c) Wright-Giemsa stain of peritoneal lavage cells from wt or lyn−/− mice. Mast cells are indicated by intense granule staining (dark cells, marked with arrow head), whereas eosinophils show a light red/pink stain and multilobed/segmented nucleus (arrow, see inset in lyn−/− sample). Graph is a quantitation of the percentage of eosinophils found in the peritoneal lavage of all wt and lyn−/− mice analyzed (n = 10). (d) The serum IgE concentration of the indicated genotypes was measured by ELISA from eight sex- and age-matched mice as described in Materials and Methods. Mean of the observed IgE concentrations for lyn−/− or lyn−/− fyn−/− mice is indicated by the line. No significant difference was found between these two populations, but both were significantly different from wt or fyn−/− mice. Significance was determined by an unpaired t test. ***, a p-value of ≤0.006; *, a p-value of ≤0.03.
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Related In: Results  -  Collection

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fig6: Lyn−/− mice have increased numbers of peritoneal mast cells with increased cell surface FcεRI expression and eosinophilia. Fyn is not required for increased serum IgE. (a) Cells from a peritoneal lavage of wt or lyn−/− mice were stained with antibody to KIT and IgE after incubation with IgE to ensure FcεRI saturation. As a negative control, isotype-specific IgG was also used to stain the cells. Inset numbers reflect the percentage of KIT/FcεRI+ cells. (b) The percentage of mast cells in the peritoneum and the mean fluorescence intensity of FITC-IgE staining (reflecting mast cell FcεRI expression) of all the mice analyzed (n = 10). (c) Wright-Giemsa stain of peritoneal lavage cells from wt or lyn−/− mice. Mast cells are indicated by intense granule staining (dark cells, marked with arrow head), whereas eosinophils show a light red/pink stain and multilobed/segmented nucleus (arrow, see inset in lyn−/− sample). Graph is a quantitation of the percentage of eosinophils found in the peritoneal lavage of all wt and lyn−/− mice analyzed (n = 10). (d) The serum IgE concentration of the indicated genotypes was measured by ELISA from eight sex- and age-matched mice as described in Materials and Methods. Mean of the observed IgE concentrations for lyn−/− or lyn−/− fyn−/− mice is indicated by the line. No significant difference was found between these two populations, but both were significantly different from wt or fyn−/− mice. Significance was determined by an unpaired t test. ***, a p-value of ≤0.006; *, a p-value of ≤0.03.
Mentions: As stated above, increased concentrations of monomeric IgE were shown to increase mast cell survival, but these conditions also resulted in increased cell surface expression of mast cell FcεRI (for review see reference 12), two factors associated with increased allergic responses. Therefore, we investigated whether these factors might be observed in 7-wk-old lyn−/− mice given their increased serum IgE. Fig. 6 a shows that mast cells in a peritoneal lavage of lyn−/− mice are increased, compared with wild-type mice of the same age. Analysis on all tested mice revealed that peritoneal mast cells of wild-type mice comprised, on average, 2.2% of the total cells in the peritoneal lavage (Fig. 6 b). In contrast, lyn−/− mice showed, on average, ∼5.3% as mast cells (ranging from a low of 2.1 to a high of ∼11%). Regardless of whether the numbers of peritoneal mast cells in a given mouse were enhanced or not, all mast cells showed increased cell surface FcεRI expression (Fig. 6 b). It should be noted that lyn−/− mice also showed increased numbers of dermal mast cells, with 8.0 ± 0.4 mast cells per field (a magnification of 1,000) versus wild-type mice, which had 4.5 ± 0.3 mast cells (P ≤ 0.001; not depicted). Importantly, 4-wk-old wild-type and lyn−/− mice did not differ significantly in peritoneal mast cell numbers with 1.3 ± 0.4 and 1.7 ± 0.3% of total cells as mast cells, respectively (not depicted), even though the lyn−/− mice showed a slight trend toward increased mast cell numbers.

Bottom Line: Contrary to this belief, Lyn kinase was found to have an important role as a negative regulator of the allergic response.This became apparent from the hyperresponsive degranulation of lyn-/- bone marrow-derived mast cells, which is driven by hyperactivation of Fyn kinase that occurs, in part, through the loss of negative regulation by COOH-terminal Src kinase (Csk) and the adaptor, Csk-binding protein.This was mirrored by increased circulating histamine, increased mast cell numbers, increased cell surface expression of the high affinity IgE receptor (FcepsilonRI), and eosinophilia.

View Article: PubMed Central - PubMed

Affiliation: Molecular Inflammation Section, Molecular Immunology and Inflammation Branch, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
A role for Lyn kinase as a positive regulator of immunoglobulin (Ig)E-dependent allergy has long been accepted. Contrary to this belief, Lyn kinase was found to have an important role as a negative regulator of the allergic response. This became apparent from the hyperresponsive degranulation of lyn-/- bone marrow-derived mast cells, which is driven by hyperactivation of Fyn kinase that occurs, in part, through the loss of negative regulation by COOH-terminal Src kinase (Csk) and the adaptor, Csk-binding protein. This phenotype is recapitulated in vivo as young lyn-/- mice showed an enhanced anaphylactic response. In vivo studies also demonstrated that as lyn-/- mice aged, their serum IgE increased as well as occupancy of the high affinity IgE receptor (FcepsilonRI). This was mirrored by increased circulating histamine, increased mast cell numbers, increased cell surface expression of the high affinity IgE receptor (FcepsilonRI), and eosinophilia. The increased IgE production was not a consequence of increased Fyn kinase activity in lyn-/- mice because both lyn-/- and lyn-/- fyn-/- mice showed high IgE levels. Thus, lyn-/- mice and mast cells thereof show multiple allergy-associated traits, causing reconsideration of the possible efficacy in therapeutic targeting of Lyn in allergic disease.

Show MeSH
Related in: MedlinePlus