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Role of T cells in a gp91phox knockout murine model of acute allergic asthma.

Banerjee ER, Henderson WR - Allergy Asthma Clin Immunol (2013)

Bottom Line: Molecular regulation of inflammation, especially, the role of effector cells in NADPH oxidase-mediated redox reactions for producing O2- (superoxide anion) is a critical step.Materials and Methods and Treatment: To clarify the role of NADPH oxidase in the pathophysiology of T cell-initiatedmacrophage-associated allergic asthma, we induced allergen dependent inflammation in a gp91phox-/- SKO (single knockout) and a gp91phox-/- MMP-12-/- DKO (double knockout) mouse and analysed trafficking and functionality of various cell types, the T cell function and T cell-macrophage interaction being given special emphasis.Composite asthma symptoms expressed in a more aggravated manner in both the KO (SKO and DKO) mice compared to WT indicating that some redundancy may exist in the response pathways of gp91phox and MMP-12.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medicine, Division of Allergy and Infectious Diseases, Center for Allergy and Inflammation, University of Washington, Room 254, 850 Republican Street, Seattle, WA 98109, USA. enarb1@gmail.com.

ABSTRACT

Objective: Molecular regulation of inflammation, especially, the role of effector cells in NADPH oxidase-mediated redox reactions for producing O2- (superoxide anion) is a critical step. This study explores the roles of macrophages and neutrophils and their cross-talk with extra-cellular matrix components in the light of the role essayed by T cells. Materials and Methods and Treatment: To clarify the role of NADPH oxidase in the pathophysiology of T cell-initiatedmacrophage-associated allergic asthma, we induced allergen dependent inflammation in a gp91phox-/- SKO (single knockout) and a gp91phox-/- MMP-12-/- DKO (double knockout) mouse and analysed trafficking and functionality of various cell types, the T cell function and T cell-macrophage interaction being given special emphasis.

Results: Composite asthma symptoms expressed in a more aggravated manner in both the KO (SKO and DKO) mice compared to WT indicating that some redundancy may exist in the response pathways of gp91phox and MMP-12. On the one hand, upregulation in macrophage functions such as proliferation, mixed lymphocyte reaction, and MCP-1 directed chemotaxis, may indicate that a regulatory cross-talk is switched on between T cell and macrophage and on the other, downregulation of respiratory burst response hints at a dichotomy in their signaling pathways. Increased B7.1 but reduced B7.2 and MHC class II expression on KO alveolar macrophages may suggest that a switching on-off mechanism is operative where alteration of co-stimulatory molecule expression selectively activating T cell is a critical step.

Inference: T cell mediated functions such as Th2 cytokine secretion, and T cell proliferation in response to OVA were upregulated synchronous with the overall robustness of the asthma phenotype.

Conclusions: As far as cell-cell interaction is concerned, the data is indicative of the existence of a plethora of networks where molecular switches may exist that selectively induce activation and deactivation of regulatory pathways that ultimately manifest in the overall response. gp91phox and MMP-12 either redundantly or synergistically but not additively, provide a regulatory checkpoint for restricting T cell cross-talk with macrophages and keep excessive tissue damage and ECM degradation during acute allergic inflammation under control.

No MeSH data available.


Related in: MedlinePlus

Upregulation of MMP-12 gene in post-OVA in KO lung. Real time PCR analysis was used to quantitate expression of mRNA for the particular genes as calculated by relative index of Ct values normalized to GAPDH by real time PCR. PCR was carried out using the comparative Ct method (Applied Biosystems software) with SYBR Green PCRcore reagents (Applied Biosystems) and anlysed using Applied Biosystems 7900HT Real-Time PCR System software SDS 2.2.1. All primers used were specific to mouse. * denotes p value<0.01 compared to WT+OVA values. # denotes p value<0.01 compared to WT+alum (control baseline values). n=5/group pooled from 2 experiments. Expression of the gene of interest was expressed in relative values normalized to the values obtained for mouse GAPDH. There was 10-fold increase in the gene expression of MMP-12 in OVA-treated WT vs. control WT whereas saline-treated NOX KO lungs showed 6.5-fold increase over untreated control mouse lung. However, post-OVA treatment, NOX KO mouse lung showed MMP-12 shooting up 4.7-fold over untreated KO. # denotes p value <0.01 compared to control untreated, * denotes p value <0.01 compares untreated KO versus OVA-treated KO lung.
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Figure 3: Upregulation of MMP-12 gene in post-OVA in KO lung. Real time PCR analysis was used to quantitate expression of mRNA for the particular genes as calculated by relative index of Ct values normalized to GAPDH by real time PCR. PCR was carried out using the comparative Ct method (Applied Biosystems software) with SYBR Green PCRcore reagents (Applied Biosystems) and anlysed using Applied Biosystems 7900HT Real-Time PCR System software SDS 2.2.1. All primers used were specific to mouse. * denotes p value<0.01 compared to WT+OVA values. # denotes p value<0.01 compared to WT+alum (control baseline values). n=5/group pooled from 2 experiments. Expression of the gene of interest was expressed in relative values normalized to the values obtained for mouse GAPDH. There was 10-fold increase in the gene expression of MMP-12 in OVA-treated WT vs. control WT whereas saline-treated NOX KO lungs showed 6.5-fold increase over untreated control mouse lung. However, post-OVA treatment, NOX KO mouse lung showed MMP-12 shooting up 4.7-fold over untreated KO. # denotes p value <0.01 compared to control untreated, * denotes p value <0.01 compares untreated KO versus OVA-treated KO lung.

Mentions: Since this is an inflammatory response to the allergen, we hypothesized that other pro-inflammatory kinases and proteases may also be intimately involved in the pathway. FigureĀ 2 and 3 shows RT-PCR analysis of gene expression of Rho kinase, known to have anti-inflammatory functions. Genes for MMP7, 10 and 28 were similarly downregulated as well.


Role of T cells in a gp91phox knockout murine model of acute allergic asthma.

Banerjee ER, Henderson WR - Allergy Asthma Clin Immunol (2013)

Upregulation of MMP-12 gene in post-OVA in KO lung. Real time PCR analysis was used to quantitate expression of mRNA for the particular genes as calculated by relative index of Ct values normalized to GAPDH by real time PCR. PCR was carried out using the comparative Ct method (Applied Biosystems software) with SYBR Green PCRcore reagents (Applied Biosystems) and anlysed using Applied Biosystems 7900HT Real-Time PCR System software SDS 2.2.1. All primers used were specific to mouse. * denotes p value<0.01 compared to WT+OVA values. # denotes p value<0.01 compared to WT+alum (control baseline values). n=5/group pooled from 2 experiments. Expression of the gene of interest was expressed in relative values normalized to the values obtained for mouse GAPDH. There was 10-fold increase in the gene expression of MMP-12 in OVA-treated WT vs. control WT whereas saline-treated NOX KO lungs showed 6.5-fold increase over untreated control mouse lung. However, post-OVA treatment, NOX KO mouse lung showed MMP-12 shooting up 4.7-fold over untreated KO. # denotes p value <0.01 compared to control untreated, * denotes p value <0.01 compares untreated KO versus OVA-treated KO lung.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 3: Upregulation of MMP-12 gene in post-OVA in KO lung. Real time PCR analysis was used to quantitate expression of mRNA for the particular genes as calculated by relative index of Ct values normalized to GAPDH by real time PCR. PCR was carried out using the comparative Ct method (Applied Biosystems software) with SYBR Green PCRcore reagents (Applied Biosystems) and anlysed using Applied Biosystems 7900HT Real-Time PCR System software SDS 2.2.1. All primers used were specific to mouse. * denotes p value<0.01 compared to WT+OVA values. # denotes p value<0.01 compared to WT+alum (control baseline values). n=5/group pooled from 2 experiments. Expression of the gene of interest was expressed in relative values normalized to the values obtained for mouse GAPDH. There was 10-fold increase in the gene expression of MMP-12 in OVA-treated WT vs. control WT whereas saline-treated NOX KO lungs showed 6.5-fold increase over untreated control mouse lung. However, post-OVA treatment, NOX KO mouse lung showed MMP-12 shooting up 4.7-fold over untreated KO. # denotes p value <0.01 compared to control untreated, * denotes p value <0.01 compares untreated KO versus OVA-treated KO lung.
Mentions: Since this is an inflammatory response to the allergen, we hypothesized that other pro-inflammatory kinases and proteases may also be intimately involved in the pathway. FigureĀ 2 and 3 shows RT-PCR analysis of gene expression of Rho kinase, known to have anti-inflammatory functions. Genes for MMP7, 10 and 28 were similarly downregulated as well.

Bottom Line: Molecular regulation of inflammation, especially, the role of effector cells in NADPH oxidase-mediated redox reactions for producing O2- (superoxide anion) is a critical step.Materials and Methods and Treatment: To clarify the role of NADPH oxidase in the pathophysiology of T cell-initiatedmacrophage-associated allergic asthma, we induced allergen dependent inflammation in a gp91phox-/- SKO (single knockout) and a gp91phox-/- MMP-12-/- DKO (double knockout) mouse and analysed trafficking and functionality of various cell types, the T cell function and T cell-macrophage interaction being given special emphasis.Composite asthma symptoms expressed in a more aggravated manner in both the KO (SKO and DKO) mice compared to WT indicating that some redundancy may exist in the response pathways of gp91phox and MMP-12.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medicine, Division of Allergy and Infectious Diseases, Center for Allergy and Inflammation, University of Washington, Room 254, 850 Republican Street, Seattle, WA 98109, USA. enarb1@gmail.com.

ABSTRACT

Objective: Molecular regulation of inflammation, especially, the role of effector cells in NADPH oxidase-mediated redox reactions for producing O2- (superoxide anion) is a critical step. This study explores the roles of macrophages and neutrophils and their cross-talk with extra-cellular matrix components in the light of the role essayed by T cells. Materials and Methods and Treatment: To clarify the role of NADPH oxidase in the pathophysiology of T cell-initiatedmacrophage-associated allergic asthma, we induced allergen dependent inflammation in a gp91phox-/- SKO (single knockout) and a gp91phox-/- MMP-12-/- DKO (double knockout) mouse and analysed trafficking and functionality of various cell types, the T cell function and T cell-macrophage interaction being given special emphasis.

Results: Composite asthma symptoms expressed in a more aggravated manner in both the KO (SKO and DKO) mice compared to WT indicating that some redundancy may exist in the response pathways of gp91phox and MMP-12. On the one hand, upregulation in macrophage functions such as proliferation, mixed lymphocyte reaction, and MCP-1 directed chemotaxis, may indicate that a regulatory cross-talk is switched on between T cell and macrophage and on the other, downregulation of respiratory burst response hints at a dichotomy in their signaling pathways. Increased B7.1 but reduced B7.2 and MHC class II expression on KO alveolar macrophages may suggest that a switching on-off mechanism is operative where alteration of co-stimulatory molecule expression selectively activating T cell is a critical step.

Inference: T cell mediated functions such as Th2 cytokine secretion, and T cell proliferation in response to OVA were upregulated synchronous with the overall robustness of the asthma phenotype.

Conclusions: As far as cell-cell interaction is concerned, the data is indicative of the existence of a plethora of networks where molecular switches may exist that selectively induce activation and deactivation of regulatory pathways that ultimately manifest in the overall response. gp91phox and MMP-12 either redundantly or synergistically but not additively, provide a regulatory checkpoint for restricting T cell cross-talk with macrophages and keep excessive tissue damage and ECM degradation during acute allergic inflammation under control.

No MeSH data available.


Related in: MedlinePlus