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FcRγ controls the fas-dependent regulatory function of lymphoproliferative double negative T cells.

Juvet SC, Thomson CW, Kim EY, Han M, Zhang L - PLoS ONE (2013)

Bottom Line: We have shown previously that a significant proportion of DN T cells express FcRγ, and that this molecule is required for TCR transgenic DN T cells to suppress allogeneic immune responses.Furthermore, FcRγ acted in a cell-intrinsic fashion to limit DN T cell accumulation by increasing the rate of apoptosis in proliferated cells.These results indicate that FcRγ can confer Fas-dependent regulatory properties on LPR DN T cells, and suggest that FcRγ may be a novel marker for functional DN Tregs.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Science, University of Toronto, Toronto, Ontario, Canada.

ABSTRACT
Patients with autoimmune lymphoproliferative syndrome (ALPS) and lymphoproliferation (LPR) mice are deficient in Fas, and accumulate large numbers of αβ-TCR(+), CD4(-), CD8(-) double negative (DN) T cells. The function of these DN T cells remains largely unknown. The common γ subunit of the activating Fc receptors, FcRγ, plays an important role in mediating innate immune responses. We have shown previously that a significant proportion of DN T cells express FcRγ, and that this molecule is required for TCR transgenic DN T cells to suppress allogeneic immune responses. Whether FcRγ plays a critical role in LPR DN T cell-mediated suppression of immune responses to auto and allo-antigens is not known. Here, we demonstrated that FcRγ(+), but not FcRγ(-) LPR DN T cells could suppress Fas(+) CD4(+) and CD8(+) T cell proliferation in vitro and attenuated CD4(+) T cell-mediated graft-versus host disease. Although FcRγ expression did not allow LPR DN T cells to inhibit the expansion of Fas-deficient cells within the LPR context, adoptive transfer of FcRγ(+), but not FcRγ(-), DN T cells inhibited lymphoproliferation in generalized lymphoproliferative disease (GLD) mice. Furthermore, FcRγ acted in a cell-intrinsic fashion to limit DN T cell accumulation by increasing the rate of apoptosis in proliferated cells. These results indicate that FcRγ can confer Fas-dependent regulatory properties on LPR DN T cells, and suggest that FcRγ may be a novel marker for functional DN Tregs.

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Fc receptor γ-expressing DN T cells are lost with disease progression in LPR mice and have an increased rate of apoptosis ex vivo.A. Splenocytes of young (≤12 weeks, n = 4) and older (≥16 weeks, n = 4) female LPR mice were counted and stained for expression of CD4, CD8, NK1.1, TCRβ, and CD16 and then examined by flow cytometry. The percentage of DN T cells expressing CD16 was examined as a function of total spleen cell count; linear regression r2 = 0.91, p = 0.0002. B. LPR FcRγ+/+ (n = 5) and LPR FcRγ−/− mice (n = 5) aged 8 weeks were fed BrdU for 6 days. Their splenocytes were then stained for expression of CD4, CD8, NK1.1, TCRβ, annexin V and CD16, fixed and stained for BrdU incorporation. They were then examined by flow cytometry. Left panels show expression of CD16 versus side light scatter in LPR FcRγ+/+ (top) and LPR FcRγ−/− DN T cells (bottom); CD16hi and CD16lo gates are indicated, and the numbers above each gate indicate the percentage of DN T cells falling into the indicated gates. Right panels show representative BrdU and annexin V staining in LPR FcRγ−/− DN T cells (bottom) and in the CD16lo and CD16hi subsets of LPR FcRγ+/+ DN T cells (middle and top panels, respectively). Numbers inside contour plots reflect the percentage of gated cells falling into each quadrant. C. The percentages of DN T cells staining with annexin V in LPR FcRγ−/− mice and in the CD16hi and CD16lo subsets of LPR FcRγ+/+ mice are presented with respect to BrdU incorporation. Two-way ANOVA p<0.0001; **Bonferroni post tests p<0.01 compared with either CD16lo or LPR FcRγ−/− DN T cells amongst both BrdU+ and BrdU− DN T cells.
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pone-0065253-g003: Fc receptor γ-expressing DN T cells are lost with disease progression in LPR mice and have an increased rate of apoptosis ex vivo.A. Splenocytes of young (≤12 weeks, n = 4) and older (≥16 weeks, n = 4) female LPR mice were counted and stained for expression of CD4, CD8, NK1.1, TCRβ, and CD16 and then examined by flow cytometry. The percentage of DN T cells expressing CD16 was examined as a function of total spleen cell count; linear regression r2 = 0.91, p = 0.0002. B. LPR FcRγ+/+ (n = 5) and LPR FcRγ−/− mice (n = 5) aged 8 weeks were fed BrdU for 6 days. Their splenocytes were then stained for expression of CD4, CD8, NK1.1, TCRβ, annexin V and CD16, fixed and stained for BrdU incorporation. They were then examined by flow cytometry. Left panels show expression of CD16 versus side light scatter in LPR FcRγ+/+ (top) and LPR FcRγ−/− DN T cells (bottom); CD16hi and CD16lo gates are indicated, and the numbers above each gate indicate the percentage of DN T cells falling into the indicated gates. Right panels show representative BrdU and annexin V staining in LPR FcRγ−/− DN T cells (bottom) and in the CD16lo and CD16hi subsets of LPR FcRγ+/+ DN T cells (middle and top panels, respectively). Numbers inside contour plots reflect the percentage of gated cells falling into each quadrant. C. The percentages of DN T cells staining with annexin V in LPR FcRγ−/− mice and in the CD16hi and CD16lo subsets of LPR FcRγ+/+ mice are presented with respect to BrdU incorporation. Two-way ANOVA p<0.0001; **Bonferroni post tests p<0.01 compared with either CD16lo or LPR FcRγ−/− DN T cells amongst both BrdU+ and BrdU− DN T cells.

Mentions: In order to understand the relationship between FcRγ expression in DN T cells and the development of lymphoproliferative disease, we compared DN T cell FcRγ/CD16 expression in younger (∼12 weeks of age) and older (∼16 weeks of age) LPR mice and observed a loss of this subset as lymphocytes accumulated (Fig. 3A, r2 = 0.91). This finding suggested that FcRγ-expressing DN T cells might be proliferating more slowly and/or dying more rapidly than DN T cells not expressing this molecule in LPR mice.


FcRγ controls the fas-dependent regulatory function of lymphoproliferative double negative T cells.

Juvet SC, Thomson CW, Kim EY, Han M, Zhang L - PLoS ONE (2013)

Fc receptor γ-expressing DN T cells are lost with disease progression in LPR mice and have an increased rate of apoptosis ex vivo.A. Splenocytes of young (≤12 weeks, n = 4) and older (≥16 weeks, n = 4) female LPR mice were counted and stained for expression of CD4, CD8, NK1.1, TCRβ, and CD16 and then examined by flow cytometry. The percentage of DN T cells expressing CD16 was examined as a function of total spleen cell count; linear regression r2 = 0.91, p = 0.0002. B. LPR FcRγ+/+ (n = 5) and LPR FcRγ−/− mice (n = 5) aged 8 weeks were fed BrdU for 6 days. Their splenocytes were then stained for expression of CD4, CD8, NK1.1, TCRβ, annexin V and CD16, fixed and stained for BrdU incorporation. They were then examined by flow cytometry. Left panels show expression of CD16 versus side light scatter in LPR FcRγ+/+ (top) and LPR FcRγ−/− DN T cells (bottom); CD16hi and CD16lo gates are indicated, and the numbers above each gate indicate the percentage of DN T cells falling into the indicated gates. Right panels show representative BrdU and annexin V staining in LPR FcRγ−/− DN T cells (bottom) and in the CD16lo and CD16hi subsets of LPR FcRγ+/+ DN T cells (middle and top panels, respectively). Numbers inside contour plots reflect the percentage of gated cells falling into each quadrant. C. The percentages of DN T cells staining with annexin V in LPR FcRγ−/− mice and in the CD16hi and CD16lo subsets of LPR FcRγ+/+ mice are presented with respect to BrdU incorporation. Two-way ANOVA p<0.0001; **Bonferroni post tests p<0.01 compared with either CD16lo or LPR FcRγ−/− DN T cells amongst both BrdU+ and BrdU− DN T cells.
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pone-0065253-g003: Fc receptor γ-expressing DN T cells are lost with disease progression in LPR mice and have an increased rate of apoptosis ex vivo.A. Splenocytes of young (≤12 weeks, n = 4) and older (≥16 weeks, n = 4) female LPR mice were counted and stained for expression of CD4, CD8, NK1.1, TCRβ, and CD16 and then examined by flow cytometry. The percentage of DN T cells expressing CD16 was examined as a function of total spleen cell count; linear regression r2 = 0.91, p = 0.0002. B. LPR FcRγ+/+ (n = 5) and LPR FcRγ−/− mice (n = 5) aged 8 weeks were fed BrdU for 6 days. Their splenocytes were then stained for expression of CD4, CD8, NK1.1, TCRβ, annexin V and CD16, fixed and stained for BrdU incorporation. They were then examined by flow cytometry. Left panels show expression of CD16 versus side light scatter in LPR FcRγ+/+ (top) and LPR FcRγ−/− DN T cells (bottom); CD16hi and CD16lo gates are indicated, and the numbers above each gate indicate the percentage of DN T cells falling into the indicated gates. Right panels show representative BrdU and annexin V staining in LPR FcRγ−/− DN T cells (bottom) and in the CD16lo and CD16hi subsets of LPR FcRγ+/+ DN T cells (middle and top panels, respectively). Numbers inside contour plots reflect the percentage of gated cells falling into each quadrant. C. The percentages of DN T cells staining with annexin V in LPR FcRγ−/− mice and in the CD16hi and CD16lo subsets of LPR FcRγ+/+ mice are presented with respect to BrdU incorporation. Two-way ANOVA p<0.0001; **Bonferroni post tests p<0.01 compared with either CD16lo or LPR FcRγ−/− DN T cells amongst both BrdU+ and BrdU− DN T cells.
Mentions: In order to understand the relationship between FcRγ expression in DN T cells and the development of lymphoproliferative disease, we compared DN T cell FcRγ/CD16 expression in younger (∼12 weeks of age) and older (∼16 weeks of age) LPR mice and observed a loss of this subset as lymphocytes accumulated (Fig. 3A, r2 = 0.91). This finding suggested that FcRγ-expressing DN T cells might be proliferating more slowly and/or dying more rapidly than DN T cells not expressing this molecule in LPR mice.

Bottom Line: We have shown previously that a significant proportion of DN T cells express FcRγ, and that this molecule is required for TCR transgenic DN T cells to suppress allogeneic immune responses.Furthermore, FcRγ acted in a cell-intrinsic fashion to limit DN T cell accumulation by increasing the rate of apoptosis in proliferated cells.These results indicate that FcRγ can confer Fas-dependent regulatory properties on LPR DN T cells, and suggest that FcRγ may be a novel marker for functional DN Tregs.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Science, University of Toronto, Toronto, Ontario, Canada.

ABSTRACT
Patients with autoimmune lymphoproliferative syndrome (ALPS) and lymphoproliferation (LPR) mice are deficient in Fas, and accumulate large numbers of αβ-TCR(+), CD4(-), CD8(-) double negative (DN) T cells. The function of these DN T cells remains largely unknown. The common γ subunit of the activating Fc receptors, FcRγ, plays an important role in mediating innate immune responses. We have shown previously that a significant proportion of DN T cells express FcRγ, and that this molecule is required for TCR transgenic DN T cells to suppress allogeneic immune responses. Whether FcRγ plays a critical role in LPR DN T cell-mediated suppression of immune responses to auto and allo-antigens is not known. Here, we demonstrated that FcRγ(+), but not FcRγ(-) LPR DN T cells could suppress Fas(+) CD4(+) and CD8(+) T cell proliferation in vitro and attenuated CD4(+) T cell-mediated graft-versus host disease. Although FcRγ expression did not allow LPR DN T cells to inhibit the expansion of Fas-deficient cells within the LPR context, adoptive transfer of FcRγ(+), but not FcRγ(-), DN T cells inhibited lymphoproliferation in generalized lymphoproliferative disease (GLD) mice. Furthermore, FcRγ acted in a cell-intrinsic fashion to limit DN T cell accumulation by increasing the rate of apoptosis in proliferated cells. These results indicate that FcRγ can confer Fas-dependent regulatory properties on LPR DN T cells, and suggest that FcRγ may be a novel marker for functional DN Tregs.

Show MeSH
Related in: MedlinePlus