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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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Naïve OVA-specific CD4+ T cells gain proliferation, activation and homing to the bladder in URO-OVA mice. (a) CD4+ T cell proliferation in URO-OVA mice. OT-II splenocytes were prepared, labeled with CFSE, and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse) at day 0. The BLNs were collected at day 4 and analyzed for CFSE-positive cells by flow cytometry. A gate was set on CD4+ T cells. A representative of 3 mice for each group is shown. (b) CD4+ T cell homing to the bladders of URO-OVA mice. The bladders of the same mice were collected at day 4. Single-cell suspensions were prepared and analyzed for CFSE-positive cells by flow cytometry. Total CFSE-positive cells in each bladder are indicated. A representative of 3 bladders for each group is shown. (c) CD4+ T cell activation in URO-OVA mice. 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. The BLNs were collected at day 4 and analyzed for CD44, CD45RB, CD62L and CD69 by flow cytometry. A gate was set on CD4+Thy1.1+ T cells. A representative of 3 mice for each group is shown. Hollow curve: B6 mice; Filled curve: URO-OVA mice.
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Figure 3: Naïve OVA-specific CD4+ T cells gain proliferation, activation and homing to the bladder in URO-OVA mice. (a) CD4+ T cell proliferation in URO-OVA mice. OT-II splenocytes were prepared, labeled with CFSE, and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse) at day 0. The BLNs were collected at day 4 and analyzed for CFSE-positive cells by flow cytometry. A gate was set on CD4+ T cells. A representative of 3 mice for each group is shown. (b) CD4+ T cell homing to the bladders of URO-OVA mice. The bladders of the same mice were collected at day 4. Single-cell suspensions were prepared and analyzed for CFSE-positive cells by flow cytometry. Total CFSE-positive cells in each bladder are indicated. A representative of 3 bladders for each group is shown. (c) CD4+ T cell activation in URO-OVA mice. 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. The BLNs were collected at day 4 and analyzed for CD44, CD45RB, CD62L and CD69 by flow cytometry. A gate was set on CD4+Thy1.1+ T cells. A representative of 3 mice for each group is shown. Hollow curve: B6 mice; Filled curve: URO-OVA mice.

Mentions: To further assess Ag-specific CD4+ T cell responses to bladder urothelial OVA, we adoptively transferred URO-OVA mice with naïve OT-II splenocytes labeled with carboxyfluoroscein succinimidyl ester (CFSE), followed by flow cytometric analysis of transferred OT-II CD4+ T cells. B6 mice were included for comparison. At day 4 after cell transfer a clear T cell division was observed in the BLNs (49.3%) of URO-OVA mice but not in the BLNs of B6 mice (Figure 3a). In addition, the bladders of the same URO-OVA mice also showed increased CFSE-positive infiltrating cells compared to the bladders of B6 mice (1,257 cells vs. 87 cells) (Figure 3b). These observations supported the antigenicity of the bladder urothelial OVA and its accessibility to the immune system for CD4+ T cell stimulation. Also, since OT-II CD4+ T cells accumulated more in the bladders of URO-OVA mice than in the bladders of B6 mice, it appeared that the bladder urothelial OVA served as a target for OVA-specific CD4+ T cells.


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)

Naïve OVA-specific CD4+ T cells gain proliferation, activation and homing to the bladder in URO-OVA mice. (a) CD4+ T cell proliferation in URO-OVA mice. OT-II splenocytes were prepared, labeled with CFSE, and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse) at day 0. The BLNs were collected at day 4 and analyzed for CFSE-positive cells by flow cytometry. A gate was set on CD4+ T cells. A representative of 3 mice for each group is shown. (b) CD4+ T cell homing to the bladders of URO-OVA mice. The bladders of the same mice were collected at day 4. Single-cell suspensions were prepared and analyzed for CFSE-positive cells by flow cytometry. Total CFSE-positive cells in each bladder are indicated. A representative of 3 bladders for each group is shown. (c) CD4+ T cell activation in URO-OVA mice. 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. The BLNs were collected at day 4 and analyzed for CD44, CD45RB, CD62L and CD69 by flow cytometry. A gate was set on CD4+Thy1.1+ T cells. A representative of 3 mice for each group is shown. Hollow curve: B6 mice; Filled curve: URO-OVA mice.
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Figure 3: Naïve OVA-specific CD4+ T cells gain proliferation, activation and homing to the bladder in URO-OVA mice. (a) CD4+ T cell proliferation in URO-OVA mice. OT-II splenocytes were prepared, labeled with CFSE, and transferred i.v. into URO-OVA mice and B6 mice (2 × 107 cells/mouse) at day 0. The BLNs were collected at day 4 and analyzed for CFSE-positive cells by flow cytometry. A gate was set on CD4+ T cells. A representative of 3 mice for each group is shown. (b) CD4+ T cell homing to the bladders of URO-OVA mice. The bladders of the same mice were collected at day 4. Single-cell suspensions were prepared and analyzed for CFSE-positive cells by flow cytometry. Total CFSE-positive cells in each bladder are indicated. A representative of 3 bladders for each group is shown. (c) CD4+ T cell activation in URO-OVA mice. 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. The BLNs were collected at day 4 and analyzed for CD44, CD45RB, CD62L and CD69 by flow cytometry. A gate was set on CD4+Thy1.1+ T cells. A representative of 3 mice for each group is shown. Hollow curve: B6 mice; Filled curve: URO-OVA mice.
Mentions: To further assess Ag-specific CD4+ T cell responses to bladder urothelial OVA, we adoptively transferred URO-OVA mice with naïve OT-II splenocytes labeled with carboxyfluoroscein succinimidyl ester (CFSE), followed by flow cytometric analysis of transferred OT-II CD4+ T cells. B6 mice were included for comparison. At day 4 after cell transfer a clear T cell division was observed in the BLNs (49.3%) of URO-OVA mice but not in the BLNs of B6 mice (Figure 3a). In addition, the bladders of the same URO-OVA mice also showed increased CFSE-positive infiltrating cells compared to the bladders of B6 mice (1,257 cells vs. 87 cells) (Figure 3b). These observations supported the antigenicity of the bladder urothelial OVA and its accessibility to the immune system for CD4+ T cell stimulation. Also, since OT-II CD4+ T cells accumulated more in the bladders of URO-OVA mice than in the bladders of B6 mice, it appeared that the bladder urothelial OVA served as a target for OVA-specific CD4+ T cells.

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