Limits...
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.

Show MeSH

Related in: MedlinePlus

Pre-activated OVA-specific CD4+ T cells induce bladder inflammation independent of CD8+ T cells in URO-OVA mice. URO-OVA mice were injected i.p. with 2.43 antibody for CD8+ T cell depletion in 3-day intervals from day −3 to day 9. OT-II CD4+ T cells were prepared, activated with OVA323-339 peptide in vitro, and transferred i.v. into URO-OVA mice (1 × 107 cells/mouse) for cystitis induction at day 0. Mice were sacrificed for analysis at day 10. (a) Flow cytometric analysis of CD8+ T cell depletion in the spleen (left panel) and BLNs (right panel) of URO-OVA mice treated with 2.43 antibody. URO-OVA mice without 2.43 treatment were included for comparison. A gate was set on lymphocytes according to scatter criteria. The percent of both CD4+ and CD8+ T cell subsets in total lymphocytes is indicated. (b) Histological H&E staining of the bladders of URO-OVA mice treated with (right panel) or without (middle panel) 2.43 antibody at day 10. The normal bladders of URO-OVA mice (left panel) are included for comparison. Cellular infiltration and edema are indicated by green and red arrows, respectively. A representative of 3 bladders for each group is shown. (c) RT-PCR analysis of TNF-α, NGF and substance P precursor (pre-SP) mRNAs in the bladders of URO-OVA mice treated with or without 2.43 antibody at day 10. Specific band intensity is presented after normalization with GAPDH. 1 and 2: the normal bladders; 3 and 4: the inflamed bladders of mice with no 2.43 treatment; 5 and 6: the inflamed bladders of mice treated with 2.43 antibodies.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3118865&req=5

Figure 4: Pre-activated OVA-specific CD4+ T cells induce bladder inflammation independent of CD8+ T cells in URO-OVA mice. URO-OVA mice were injected i.p. with 2.43 antibody for CD8+ T cell depletion in 3-day intervals from day −3 to day 9. OT-II CD4+ T cells were prepared, activated with OVA323-339 peptide in vitro, and transferred i.v. into URO-OVA mice (1 × 107 cells/mouse) for cystitis induction at day 0. Mice were sacrificed for analysis at day 10. (a) Flow cytometric analysis of CD8+ T cell depletion in the spleen (left panel) and BLNs (right panel) of URO-OVA mice treated with 2.43 antibody. URO-OVA mice without 2.43 treatment were included for comparison. A gate was set on lymphocytes according to scatter criteria. The percent of both CD4+ and CD8+ T cell subsets in total lymphocytes is indicated. (b) Histological H&E staining of the bladders of URO-OVA mice treated with (right panel) or without (middle panel) 2.43 antibody at day 10. The normal bladders of URO-OVA mice (left panel) are included for comparison. Cellular infiltration and edema are indicated by green and red arrows, respectively. A representative of 3 bladders for each group is shown. (c) RT-PCR analysis of TNF-α, NGF and substance P precursor (pre-SP) mRNAs in the bladders of URO-OVA mice treated with or without 2.43 antibody at day 10. Specific band intensity is presented after normalization with GAPDH. 1 and 2: the normal bladders; 3 and 4: the inflamed bladders of mice with no 2.43 treatment; 5 and 6: the inflamed bladders of mice treated with 2.43 antibodies.

Mentions: We next tested whether OVA-specific CD4+ T cells upon activation could induce bladder inflammation in URO-OVA mice. To test this, OT-II CD4+ T cells were prepared, activated in vitro with OVA323-339 peptide, and transferred i.v. into URO-OVA mice for cystitis induction at day 0. Mice were sacrificed for analysis at day 10. The in vitro activation of OT-II CD4+ T cells increased the clonal phenotype Vβ5-positive CD4+ T cells from 38% to 92% (data not shown). To further test whether CD4+ T cells had an ability to induce bladder inflammation in the absence of CD8+ T cells, URO-OVA mice were injected intraperitoneally (i.p.) with a CD8+ T cell-depleting antibody (clone: 2.43) in 3-day intervals from days −3 to 9. Mice were transferred i.v. with pre-activated OT-II CD4+ T cells for cystitis induction at day 0 and sacrificed for analysis at day 10. Flow cytometry showed nearly complete CD8+ T cell depletion in both spleen (1% vs. 10%) and BLNs (2% vs. 25%) at day 10 (Figure 4a). However, despite this CD8+ T cell depletion, mice developed histological bladder inflammation (Figure 4b, right panel; score: 2+) although its severity was less than that of mice without CD8+ T cell depletion (Figure 4b, middle panel; score: 2-3+). Both inflamed bladders of 2.43-treated and non-treated mice showed clear edema and cellular infiltration by histological hematoxylin and eosin (H&E) staining. In addition, the inflamed bladders of 2.43-treated mice also produced increased neuroinflammatory factor mRNAs such as tumor necrosis factor (TNF)-α, nerve growth factor (NGF) and substance P precursor (Figure 4c), although the levels of mRNAs produced was 20-30% lower than those of the bladders of non-treated mice. Nevertheless, these observations indicated that urothelial Ag-specific CD4+ T cells could function as direct effector cells to induce bladder autoimmune inflammation independent of CD8+ 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)

Pre-activated OVA-specific CD4+ T cells induce bladder inflammation independent of CD8+ T cells in URO-OVA mice. URO-OVA mice were injected i.p. with 2.43 antibody for CD8+ T cell depletion in 3-day intervals from day −3 to day 9. OT-II CD4+ T cells were prepared, activated with OVA323-339 peptide in vitro, and transferred i.v. into URO-OVA mice (1 × 107 cells/mouse) for cystitis induction at day 0. Mice were sacrificed for analysis at day 10. (a) Flow cytometric analysis of CD8+ T cell depletion in the spleen (left panel) and BLNs (right panel) of URO-OVA mice treated with 2.43 antibody. URO-OVA mice without 2.43 treatment were included for comparison. A gate was set on lymphocytes according to scatter criteria. The percent of both CD4+ and CD8+ T cell subsets in total lymphocytes is indicated. (b) Histological H&E staining of the bladders of URO-OVA mice treated with (right panel) or without (middle panel) 2.43 antibody at day 10. The normal bladders of URO-OVA mice (left panel) are included for comparison. Cellular infiltration and edema are indicated by green and red arrows, respectively. A representative of 3 bladders for each group is shown. (c) RT-PCR analysis of TNF-α, NGF and substance P precursor (pre-SP) mRNAs in the bladders of URO-OVA mice treated with or without 2.43 antibody at day 10. Specific band intensity is presented after normalization with GAPDH. 1 and 2: the normal bladders; 3 and 4: the inflamed bladders of mice with no 2.43 treatment; 5 and 6: the inflamed bladders of mice treated with 2.43 antibodies.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3118865&req=5

Figure 4: Pre-activated OVA-specific CD4+ T cells induce bladder inflammation independent of CD8+ T cells in URO-OVA mice. URO-OVA mice were injected i.p. with 2.43 antibody for CD8+ T cell depletion in 3-day intervals from day −3 to day 9. OT-II CD4+ T cells were prepared, activated with OVA323-339 peptide in vitro, and transferred i.v. into URO-OVA mice (1 × 107 cells/mouse) for cystitis induction at day 0. Mice were sacrificed for analysis at day 10. (a) Flow cytometric analysis of CD8+ T cell depletion in the spleen (left panel) and BLNs (right panel) of URO-OVA mice treated with 2.43 antibody. URO-OVA mice without 2.43 treatment were included for comparison. A gate was set on lymphocytes according to scatter criteria. The percent of both CD4+ and CD8+ T cell subsets in total lymphocytes is indicated. (b) Histological H&E staining of the bladders of URO-OVA mice treated with (right panel) or without (middle panel) 2.43 antibody at day 10. The normal bladders of URO-OVA mice (left panel) are included for comparison. Cellular infiltration and edema are indicated by green and red arrows, respectively. A representative of 3 bladders for each group is shown. (c) RT-PCR analysis of TNF-α, NGF and substance P precursor (pre-SP) mRNAs in the bladders of URO-OVA mice treated with or without 2.43 antibody at day 10. Specific band intensity is presented after normalization with GAPDH. 1 and 2: the normal bladders; 3 and 4: the inflamed bladders of mice with no 2.43 treatment; 5 and 6: the inflamed bladders of mice treated with 2.43 antibodies.
Mentions: We next tested whether OVA-specific CD4+ T cells upon activation could induce bladder inflammation in URO-OVA mice. To test this, OT-II CD4+ T cells were prepared, activated in vitro with OVA323-339 peptide, and transferred i.v. into URO-OVA mice for cystitis induction at day 0. Mice were sacrificed for analysis at day 10. The in vitro activation of OT-II CD4+ T cells increased the clonal phenotype Vβ5-positive CD4+ T cells from 38% to 92% (data not shown). To further test whether CD4+ T cells had an ability to induce bladder inflammation in the absence of CD8+ T cells, URO-OVA mice were injected intraperitoneally (i.p.) with a CD8+ T cell-depleting antibody (clone: 2.43) in 3-day intervals from days −3 to 9. Mice were transferred i.v. with pre-activated OT-II CD4+ T cells for cystitis induction at day 0 and sacrificed for analysis at day 10. Flow cytometry showed nearly complete CD8+ T cell depletion in both spleen (1% vs. 10%) and BLNs (2% vs. 25%) at day 10 (Figure 4a). However, despite this CD8+ T cell depletion, mice developed histological bladder inflammation (Figure 4b, right panel; score: 2+) although its severity was less than that of mice without CD8+ T cell depletion (Figure 4b, middle panel; score: 2-3+). Both inflamed bladders of 2.43-treated and non-treated mice showed clear edema and cellular infiltration by histological hematoxylin and eosin (H&E) staining. In addition, the inflamed bladders of 2.43-treated mice also produced increased neuroinflammatory factor mRNAs such as tumor necrosis factor (TNF)-α, nerve growth factor (NGF) and substance P precursor (Figure 4c), although the levels of mRNAs produced was 20-30% lower than those of the bladders of non-treated mice. Nevertheless, these observations indicated that urothelial Ag-specific CD4+ T cells could function as direct effector cells to induce bladder autoimmune inflammation independent of CD8+ 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