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Uteroglobin represses allergen-induced inflammatory response by blocking PGD2 receptor-mediated functions.

Mandal AK, Zhang Z, Ray R, Choi MS, Chowdhury B, Pattabiraman N, Mukherjee AB - J. Exp. Med. (2004)

Bottom Line: These effects are abrogated by recombinant UG treatment.Here we report that DP signaling is mediated via p38 mitogen-activated protein kinase, p44/42 mitogen-activated protein kinase, and protein kinase C pathways in a cell type-specific manner leading to nuclear factor-kappaB activation stimulating COX-2 gene expression.Further, we found that recombinant UG blocks DP-mediated nuclear factor-kappaB activation and suppresses COX-2 gene expression.

View Article: PubMed Central - PubMed

Affiliation: Bldg. 10, Rm. 9S241, National Institutes of Health, 9000 Rockville Pike, Bethesda, MD 20892, USA.

ABSTRACT
Uteroglobin (UG) is an antiinflammatory protein secreted by the epithelial lining of all organs communicating with the external environment. We reported previously that UG-knockout mice manifest exaggerated inflammatory response to allergen, characterized by increased eotaxin and Th2 cytokine gene expression, and eosinophil infiltration in the lungs. In this study, we uncovered that the airway epithelia of these mice also express high levels of cyclooxygenase (COX)-2, a key enzyme for the production of proinflammatory lipid mediators, and the bronchoalveolar lavage fluid (BALF) contain elevated levels of prostaglandin D2. These effects are abrogated by recombinant UG treatment. Although it has been reported that prostaglandin D2 mediates allergic inflammation via its receptor, DP, neither the molecular mechanism(s) of DP signaling nor the mechanism by which UG suppresses DP-mediated inflammatory response are clearly understood. Here we report that DP signaling is mediated via p38 mitogen-activated protein kinase, p44/42 mitogen-activated protein kinase, and protein kinase C pathways in a cell type-specific manner leading to nuclear factor-kappaB activation stimulating COX-2 gene expression. Further, we found that recombinant UG blocks DP-mediated nuclear factor-kappaB activation and suppresses COX-2 gene expression. We propose that UG is an essential component of a novel innate homeostatic mechanism in the mammalian airways to repress allergen-induced inflammatory responses.

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UG Inhibits proinflammatory cytokine expression and eosinophil infiltration. (A) The expression of IL-4 (i), IL-5 (ii), IL-13 (iii), and eotaxin (iv) mRNA was determined by real-time quantitative RT-PCR using total RNA from the lungs of PBS-treated WT mice (bar 1), OVA-sensitized and -challenged WT mice (bar 2), UG-KO mice (bar 3), OVA-sensitized and -challenged UG-KO mice without rUG treatment (bar 4), and OVA-sensitized UG-KO mice that were pretreated with rUG (250 μg of rUG in 200 μl PBS administered i.v.) 30 min before OVA challenge (bar 5). Asterisks indicate significance at P < 0.05. (B) UG-mRNA expression in the lungs of PBS-challenged WT and OVA-sensitized and -challenged WT mice by semiquantitative RT-PCR. (C) UG protein expression (immunofluorescence) in bronchiolar epithelial cells of WT mouse challenged with PBS (left) or sensitized and challenged with OVA (right). (D) Total cell counts and number of individual leukocytes per milliliter of BALF from PBS (control) and OVA-sensitized and -challenged WT (left) and UG-KO (right) mice and the results from OVA-sensitized and challenged (middle) and OVA-sensitized UG-KO mice treated with rUG before OVA challenge (extreme right). Values are expressed as the mean ± SD (n = 5 per treatment group). E, eosinophil; N, neutrophil; L, lymphocyte; M, monocyte. (E) Eosinophil infiltration in nonsensitized and unchallenged WT control (frame 1); nonsensitized and unchallenged UG-KO control (frame 2); WT mouse sensitized and challenged with OVA (frame 3); UG-KO mouse sensitized and challenged with OVA (frame 4); UG-KO mouse sensitized and treated with rUG before OVA challenge (frame 5). b, bronchiole. Magnification, ×400.
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fig1: UG Inhibits proinflammatory cytokine expression and eosinophil infiltration. (A) The expression of IL-4 (i), IL-5 (ii), IL-13 (iii), and eotaxin (iv) mRNA was determined by real-time quantitative RT-PCR using total RNA from the lungs of PBS-treated WT mice (bar 1), OVA-sensitized and -challenged WT mice (bar 2), UG-KO mice (bar 3), OVA-sensitized and -challenged UG-KO mice without rUG treatment (bar 4), and OVA-sensitized UG-KO mice that were pretreated with rUG (250 μg of rUG in 200 μl PBS administered i.v.) 30 min before OVA challenge (bar 5). Asterisks indicate significance at P < 0.05. (B) UG-mRNA expression in the lungs of PBS-challenged WT and OVA-sensitized and -challenged WT mice by semiquantitative RT-PCR. (C) UG protein expression (immunofluorescence) in bronchiolar epithelial cells of WT mouse challenged with PBS (left) or sensitized and challenged with OVA (right). (D) Total cell counts and number of individual leukocytes per milliliter of BALF from PBS (control) and OVA-sensitized and -challenged WT (left) and UG-KO (right) mice and the results from OVA-sensitized and challenged (middle) and OVA-sensitized UG-KO mice treated with rUG before OVA challenge (extreme right). Values are expressed as the mean ± SD (n = 5 per treatment group). E, eosinophil; N, neutrophil; L, lymphocyte; M, monocyte. (E) Eosinophil infiltration in nonsensitized and unchallenged WT control (frame 1); nonsensitized and unchallenged UG-KO control (frame 2); WT mouse sensitized and challenged with OVA (frame 3); UG-KO mouse sensitized and challenged with OVA (frame 4); UG-KO mouse sensitized and treated with rUG before OVA challenge (frame 5). b, bronchiole. Magnification, ×400.

Mentions: We reported previously that UG-KO mice, sensitized and challenged with OVA, manifest exaggerated allergic response characterized by elevated eotaxin and Th2 cytokine expression and an increased number of eosinophils in BALF (6). In this study, we examined the effects of rUG on these parameters. We first treated the OVA-sensitized UG-KO mice with rUG before OVA challenge and determined the expression of eotaxin and representative Th2 cytokine mRNAs by real-time quantitative RT-PCR. Our results confirm that OVA sensitization and challenge increase the level of these mRNAs in the lungs of UG-KO mice (Fig. 1 A) as reported previously (6). More importantly, we demonstrate that rUG treatment of OVA-sensitized UG-KO mice before OVA challenge inhibits the expression of IL-4 (Fig. 1 A, i), IL-5 (Fig. 1 A, ii), IL-13 (Fig. 1 A, iii), and eotaxin (Fig. 1 A, iv), all of which are recognized markers of allergic inflammatory response. The reduced level of inflammatory response in the lungs of OVA-sensitized and challenged WT mice may be due to the stimulation of UG expression in response to increased levels of Th2 cytokines. This hypothesis is at least partially supported by the fact that IL-13 treatment stimulates UG production in rat lungs (23). Therefore, we analyzed UG mRNA and UG protein in the lungs of OVA-sensitized and challenged WT mice by semiquantitative RT-PCR and immunofluorescence, respectively. The results show that OVA sensitization and challenge of WT littermates of UG-KO mice expressed higher levels of UG mRNA (Fig. 1 B) and UG protein (Fig. 1 C) compared with their nonsensitized and nonchallenged counterparts. These results raise the possibility that the observed OVA-induced elevation of UG production in WT mouse lungs is likely to be a homeostatic response to counteract inadvertent stimulation of allergen-induced inflammation.


Uteroglobin represses allergen-induced inflammatory response by blocking PGD2 receptor-mediated functions.

Mandal AK, Zhang Z, Ray R, Choi MS, Chowdhury B, Pattabiraman N, Mukherjee AB - J. Exp. Med. (2004)

UG Inhibits proinflammatory cytokine expression and eosinophil infiltration. (A) The expression of IL-4 (i), IL-5 (ii), IL-13 (iii), and eotaxin (iv) mRNA was determined by real-time quantitative RT-PCR using total RNA from the lungs of PBS-treated WT mice (bar 1), OVA-sensitized and -challenged WT mice (bar 2), UG-KO mice (bar 3), OVA-sensitized and -challenged UG-KO mice without rUG treatment (bar 4), and OVA-sensitized UG-KO mice that were pretreated with rUG (250 μg of rUG in 200 μl PBS administered i.v.) 30 min before OVA challenge (bar 5). Asterisks indicate significance at P < 0.05. (B) UG-mRNA expression in the lungs of PBS-challenged WT and OVA-sensitized and -challenged WT mice by semiquantitative RT-PCR. (C) UG protein expression (immunofluorescence) in bronchiolar epithelial cells of WT mouse challenged with PBS (left) or sensitized and challenged with OVA (right). (D) Total cell counts and number of individual leukocytes per milliliter of BALF from PBS (control) and OVA-sensitized and -challenged WT (left) and UG-KO (right) mice and the results from OVA-sensitized and challenged (middle) and OVA-sensitized UG-KO mice treated with rUG before OVA challenge (extreme right). Values are expressed as the mean ± SD (n = 5 per treatment group). E, eosinophil; N, neutrophil; L, lymphocyte; M, monocyte. (E) Eosinophil infiltration in nonsensitized and unchallenged WT control (frame 1); nonsensitized and unchallenged UG-KO control (frame 2); WT mouse sensitized and challenged with OVA (frame 3); UG-KO mouse sensitized and challenged with OVA (frame 4); UG-KO mouse sensitized and treated with rUG before OVA challenge (frame 5). b, bronchiole. Magnification, ×400.
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fig1: UG Inhibits proinflammatory cytokine expression and eosinophil infiltration. (A) The expression of IL-4 (i), IL-5 (ii), IL-13 (iii), and eotaxin (iv) mRNA was determined by real-time quantitative RT-PCR using total RNA from the lungs of PBS-treated WT mice (bar 1), OVA-sensitized and -challenged WT mice (bar 2), UG-KO mice (bar 3), OVA-sensitized and -challenged UG-KO mice without rUG treatment (bar 4), and OVA-sensitized UG-KO mice that were pretreated with rUG (250 μg of rUG in 200 μl PBS administered i.v.) 30 min before OVA challenge (bar 5). Asterisks indicate significance at P < 0.05. (B) UG-mRNA expression in the lungs of PBS-challenged WT and OVA-sensitized and -challenged WT mice by semiquantitative RT-PCR. (C) UG protein expression (immunofluorescence) in bronchiolar epithelial cells of WT mouse challenged with PBS (left) or sensitized and challenged with OVA (right). (D) Total cell counts and number of individual leukocytes per milliliter of BALF from PBS (control) and OVA-sensitized and -challenged WT (left) and UG-KO (right) mice and the results from OVA-sensitized and challenged (middle) and OVA-sensitized UG-KO mice treated with rUG before OVA challenge (extreme right). Values are expressed as the mean ± SD (n = 5 per treatment group). E, eosinophil; N, neutrophil; L, lymphocyte; M, monocyte. (E) Eosinophil infiltration in nonsensitized and unchallenged WT control (frame 1); nonsensitized and unchallenged UG-KO control (frame 2); WT mouse sensitized and challenged with OVA (frame 3); UG-KO mouse sensitized and challenged with OVA (frame 4); UG-KO mouse sensitized and treated with rUG before OVA challenge (frame 5). b, bronchiole. Magnification, ×400.
Mentions: We reported previously that UG-KO mice, sensitized and challenged with OVA, manifest exaggerated allergic response characterized by elevated eotaxin and Th2 cytokine expression and an increased number of eosinophils in BALF (6). In this study, we examined the effects of rUG on these parameters. We first treated the OVA-sensitized UG-KO mice with rUG before OVA challenge and determined the expression of eotaxin and representative Th2 cytokine mRNAs by real-time quantitative RT-PCR. Our results confirm that OVA sensitization and challenge increase the level of these mRNAs in the lungs of UG-KO mice (Fig. 1 A) as reported previously (6). More importantly, we demonstrate that rUG treatment of OVA-sensitized UG-KO mice before OVA challenge inhibits the expression of IL-4 (Fig. 1 A, i), IL-5 (Fig. 1 A, ii), IL-13 (Fig. 1 A, iii), and eotaxin (Fig. 1 A, iv), all of which are recognized markers of allergic inflammatory response. The reduced level of inflammatory response in the lungs of OVA-sensitized and challenged WT mice may be due to the stimulation of UG expression in response to increased levels of Th2 cytokines. This hypothesis is at least partially supported by the fact that IL-13 treatment stimulates UG production in rat lungs (23). Therefore, we analyzed UG mRNA and UG protein in the lungs of OVA-sensitized and challenged WT mice by semiquantitative RT-PCR and immunofluorescence, respectively. The results show that OVA sensitization and challenge of WT littermates of UG-KO mice expressed higher levels of UG mRNA (Fig. 1 B) and UG protein (Fig. 1 C) compared with their nonsensitized and nonchallenged counterparts. These results raise the possibility that the observed OVA-induced elevation of UG production in WT mouse lungs is likely to be a homeostatic response to counteract inadvertent stimulation of allergen-induced inflammation.

Bottom Line: These effects are abrogated by recombinant UG treatment.Here we report that DP signaling is mediated via p38 mitogen-activated protein kinase, p44/42 mitogen-activated protein kinase, and protein kinase C pathways in a cell type-specific manner leading to nuclear factor-kappaB activation stimulating COX-2 gene expression.Further, we found that recombinant UG blocks DP-mediated nuclear factor-kappaB activation and suppresses COX-2 gene expression.

View Article: PubMed Central - PubMed

Affiliation: Bldg. 10, Rm. 9S241, National Institutes of Health, 9000 Rockville Pike, Bethesda, MD 20892, USA.

ABSTRACT
Uteroglobin (UG) is an antiinflammatory protein secreted by the epithelial lining of all organs communicating with the external environment. We reported previously that UG-knockout mice manifest exaggerated inflammatory response to allergen, characterized by increased eotaxin and Th2 cytokine gene expression, and eosinophil infiltration in the lungs. In this study, we uncovered that the airway epithelia of these mice also express high levels of cyclooxygenase (COX)-2, a key enzyme for the production of proinflammatory lipid mediators, and the bronchoalveolar lavage fluid (BALF) contain elevated levels of prostaglandin D2. These effects are abrogated by recombinant UG treatment. Although it has been reported that prostaglandin D2 mediates allergic inflammation via its receptor, DP, neither the molecular mechanism(s) of DP signaling nor the mechanism by which UG suppresses DP-mediated inflammatory response are clearly understood. Here we report that DP signaling is mediated via p38 mitogen-activated protein kinase, p44/42 mitogen-activated protein kinase, and protein kinase C pathways in a cell type-specific manner leading to nuclear factor-kappaB activation stimulating COX-2 gene expression. Further, we found that recombinant UG blocks DP-mediated nuclear factor-kappaB activation and suppresses COX-2 gene expression. We propose that UG is an essential component of a novel innate homeostatic mechanism in the mammalian airways to repress allergen-induced inflammatory responses.

Show MeSH
Related in: MedlinePlus