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Urothelial antigen-specific CD4+ T cells function as direct effector cells and induce bladder autoimmune inflammation independent of CD8+ T cells.

Liu W, Chen X, Evanoff DP, Luo Y - Mucosal Immunol (2011)

Bottom Line: Adoptive transfer of naive OVA-specific CD4(+) T cells led to exogenous T-cell proliferation, activation, and bladder infiltration but no inflammatory induction.In contrast, adoptive transfer of preactivated OVA-specific CD4(+) T cells induced bladder inflammation.These results indicate that urothelial Ag-specific CD4(+) T cells can function as direct effector cells to induce bladder autoimmune inflammation independent of CD8(+) T cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, University of Iowa, Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa, USA.

ABSTRACT
The role of CD4(+) T cells in bladder autoimmune inflammation has not been identified because of the lack of a proper animal model. We investigated CD4(+) T-cell responses to bladder urothelial ovalbumin (OVA), a model self-antigen (Ag), in transgenic URO-OVA mice. The expression of bladder urothelial OVA rendered mice unresponsive to OVA and resulted in quick clearance of Ag-specific CD4(+) T cells. Adoptive transfer of naive OVA-specific CD4(+) T cells led to exogenous T-cell proliferation, activation, and bladder infiltration but no inflammatory induction. In contrast, adoptive transfer of preactivated OVA-specific CD4(+) T cells induced bladder inflammation. Studies further demonstrated that CD4(+) T cells induced bladder inflammation in URO-OVA mice depleted of CD8(+) T cells or deficient in the recombinase activating gene-1 (Rag-1(-/-)). These results indicate that urothelial Ag-specific CD4(+) T cells can function as direct effector cells to induce bladder autoimmune inflammation independent of CD8(+) T cells.

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URO-OVA mice exhibit quick clearance of adoptively transferred OVA-specific CD4+ T cells. OT-II splenocytes (Thy1.1) were prepared and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse; both Thy1.2 background) at day 0. Mice were sacrificed for analysis at days 1, 4 and 7. The spleens and BLNs were collected and analyzed for CD4+Thy1.1+ T cells by flow cytometry. A gate was set on lymphocytes according to scatter criteria. The percent of CD4+Thy1.1+ T cells in total lymphocytes is indicated. (a) A representative flow cytometric analysis for URO-OVA mice vs. B6 mice. (b) The kinetics of clearance of OT-II CD4+Thy1.1+ T cells in URO-OVA mice vs. B6 mice. n = 6-10 mice for each timepoint.
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Figure 2: URO-OVA mice exhibit quick clearance of adoptively transferred OVA-specific CD4+ T cells. OT-II splenocytes (Thy1.1) were prepared and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse; both Thy1.2 background) at day 0. Mice were sacrificed for analysis at days 1, 4 and 7. The spleens and BLNs were collected and analyzed for CD4+Thy1.1+ T cells by flow cytometry. A gate was set on lymphocytes according to scatter criteria. The percent of CD4+Thy1.1+ T cells in total lymphocytes is indicated. (a) A representative flow cytometric analysis for URO-OVA mice vs. B6 mice. (b) The kinetics of clearance of OT-II CD4+Thy1.1+ T cells in URO-OVA mice vs. B6 mice. n = 6-10 mice for each timepoint.

Mentions: To determine whether the expression of bladder urothelial OVA could affect host tolerance to self-Ag specific CD4+ T cells, we adoptively transferred URO-OVA mice (Thy1.2) with naive splenocytes from OT-II mice (Thy1.1), a transgenic line that expressed CD4 TCR (Vα2Vβ5) specific for I-Ab/OVA323-339 epitope.33,34 Mice were sacrificed for analysis 1, 4 and 7 days after cell transfer. B6 mice (Thy1.2) were included for comparison. Flow cytometry was used to analyze exogenous OT-II CD4+Thy1.1+ T cells in the spleens and bladder draining lymph nodes (BLNs) (Figure 2a and 2b). In B6 mice the highest number of CD4+Thy1.1+ T cells was observed at day 1. Then, the number of CD4+Thy1.1+ T cells returned to a baseline level at days 4 and 7. In contrast to B6 mice, in URO-OVA mice the number of CD4+Thy1.1+ T cells was low (baseline levels) at day 1, peaked at day 4 (20 fold increase in the spleens and 10 fold increase in the BLNs), and returned to the baseline levels at day 7. The observed increased numbers of CD4+Thy1.1+ T cells in the spleens and BLNs of URO-OVA mice suggested that the bladder urothelial OVA was antigenic and could access the immune system for CD4+ T cell stimulation. However, the numbers of CD4+Thy1.1+ T cells at day 1 was 5 fold less in URO-OVA mice than in B6 mice, suggesting that these urothelial Ag-specific CD4+ T cells encountered host clearance within 24 hours after transfer in the former mice. The quick decrease in CD4+Thy1.1+ T cell numbers from day 4 to day 7 in URO-OVA mice further suggested the ability of these mice to quick clear self-Ag specific CD4+ T cells. The CD4+Thy1.1+ T cells became less in URO-OVA mice than in B6 mice at day 14 (0.06% vs. 0.18% in the spleens and 0.07% vs. 0.17% in the BLNs) and negligible in both mice at day 21 and thereafter.


Urothelial antigen-specific CD4+ T cells function as direct effector cells and induce bladder autoimmune inflammation independent of CD8+ T cells.

Liu W, Chen X, Evanoff DP, Luo Y - Mucosal Immunol (2011)

URO-OVA mice exhibit quick clearance of adoptively transferred OVA-specific CD4+ T cells. OT-II splenocytes (Thy1.1) were prepared and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse; both Thy1.2 background) at day 0. Mice were sacrificed for analysis at days 1, 4 and 7. The spleens and BLNs were collected and analyzed for CD4+Thy1.1+ T cells by flow cytometry. A gate was set on lymphocytes according to scatter criteria. The percent of CD4+Thy1.1+ T cells in total lymphocytes is indicated. (a) A representative flow cytometric analysis for URO-OVA mice vs. B6 mice. (b) The kinetics of clearance of OT-II CD4+Thy1.1+ T cells in URO-OVA mice vs. B6 mice. n = 6-10 mice for each timepoint.
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Related In: Results  -  Collection

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Figure 2: URO-OVA mice exhibit quick clearance of adoptively transferred OVA-specific CD4+ T cells. OT-II splenocytes (Thy1.1) were prepared and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse; both Thy1.2 background) at day 0. Mice were sacrificed for analysis at days 1, 4 and 7. The spleens and BLNs were collected and analyzed for CD4+Thy1.1+ T cells by flow cytometry. A gate was set on lymphocytes according to scatter criteria. The percent of CD4+Thy1.1+ T cells in total lymphocytes is indicated. (a) A representative flow cytometric analysis for URO-OVA mice vs. B6 mice. (b) The kinetics of clearance of OT-II CD4+Thy1.1+ T cells in URO-OVA mice vs. B6 mice. n = 6-10 mice for each timepoint.
Mentions: To determine whether the expression of bladder urothelial OVA could affect host tolerance to self-Ag specific CD4+ T cells, we adoptively transferred URO-OVA mice (Thy1.2) with naive splenocytes from OT-II mice (Thy1.1), a transgenic line that expressed CD4 TCR (Vα2Vβ5) specific for I-Ab/OVA323-339 epitope.33,34 Mice were sacrificed for analysis 1, 4 and 7 days after cell transfer. B6 mice (Thy1.2) were included for comparison. Flow cytometry was used to analyze exogenous OT-II CD4+Thy1.1+ T cells in the spleens and bladder draining lymph nodes (BLNs) (Figure 2a and 2b). In B6 mice the highest number of CD4+Thy1.1+ T cells was observed at day 1. Then, the number of CD4+Thy1.1+ T cells returned to a baseline level at days 4 and 7. In contrast to B6 mice, in URO-OVA mice the number of CD4+Thy1.1+ T cells was low (baseline levels) at day 1, peaked at day 4 (20 fold increase in the spleens and 10 fold increase in the BLNs), and returned to the baseline levels at day 7. The observed increased numbers of CD4+Thy1.1+ T cells in the spleens and BLNs of URO-OVA mice suggested that the bladder urothelial OVA was antigenic and could access the immune system for CD4+ T cell stimulation. However, the numbers of CD4+Thy1.1+ T cells at day 1 was 5 fold less in URO-OVA mice than in B6 mice, suggesting that these urothelial Ag-specific CD4+ T cells encountered host clearance within 24 hours after transfer in the former mice. The quick decrease in CD4+Thy1.1+ T cell numbers from day 4 to day 7 in URO-OVA mice further suggested the ability of these mice to quick clear self-Ag specific CD4+ T cells. The CD4+Thy1.1+ T cells became less in URO-OVA mice than in B6 mice at day 14 (0.06% vs. 0.18% in the spleens and 0.07% vs. 0.17% in the BLNs) and negligible in both mice at day 21 and thereafter.

Bottom Line: Adoptive transfer of naive OVA-specific CD4(+) T cells led to exogenous T-cell proliferation, activation, and bladder infiltration but no inflammatory induction.In contrast, adoptive transfer of preactivated OVA-specific CD4(+) T cells induced bladder inflammation.These results indicate that urothelial Ag-specific CD4(+) T cells can function as direct effector cells to induce bladder autoimmune inflammation independent of CD8(+) T cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, University of Iowa, Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa, USA.

ABSTRACT
The role of CD4(+) T cells in bladder autoimmune inflammation has not been identified because of the lack of a proper animal model. We investigated CD4(+) T-cell responses to bladder urothelial ovalbumin (OVA), a model self-antigen (Ag), in transgenic URO-OVA mice. The expression of bladder urothelial OVA rendered mice unresponsive to OVA and resulted in quick clearance of Ag-specific CD4(+) T cells. Adoptive transfer of naive OVA-specific CD4(+) T cells led to exogenous T-cell proliferation, activation, and bladder infiltration but no inflammatory induction. In contrast, adoptive transfer of preactivated OVA-specific CD4(+) T cells induced bladder inflammation. Studies further demonstrated that CD4(+) T cells induced bladder inflammation in URO-OVA mice depleted of CD8(+) T cells or deficient in the recombinase activating gene-1 (Rag-1(-/-)). These results indicate that urothelial Ag-specific CD4(+) T cells can function as direct effector cells to induce bladder autoimmune inflammation independent of CD8(+) T cells.

Show MeSH
Related in: MedlinePlus