Limits...
Caspase-8 inactivation in T cells increases necroptosis and suppresses autoimmunity in Bim-/- mice.

Bohgaki T, Mozo J, Salmena L, Matysiak-Zablocki E, Bohgaki M, Sanchez O, Strasser A, Hakem A, Hakem R - J. Cell Biol. (2011)

Bottom Line: Dysregulation of either the extrinsic or intrinsic apoptotic pathway can lead to various diseases including immune disorders and cancer.In addition to its role in the extrinsic apoptotic pathway, caspase-8 plays nonapoptotic functions and is essential for T cell homeostasis.We show that, similar to caspase-8(-/-) T cells, Bim(-/-) T cells that also lack caspase-8 displayed elevated levels of necroptosis and that inhibition of this cell death process fully rescued the survival and proliferation of these cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G 2M9, Canada.

ABSTRACT
Dysregulation of either the extrinsic or intrinsic apoptotic pathway can lead to various diseases including immune disorders and cancer. In addition to its role in the extrinsic apoptotic pathway, caspase-8 plays nonapoptotic functions and is essential for T cell homeostasis. The pro-apoptotic BH3-only Bcl-2 family member Bim is important for the intrinsic apoptotic pathway and its inactivation leads to autoimmunity that is further exacerbated by loss of function of the death receptor Fas. We report that inactivation of caspase-8 in T cells of Bim(-/-) mice restrained their autoimmunity and extended their life span. We show that, similar to caspase-8(-/-) T cells, Bim(-/-) T cells that also lack caspase-8 displayed elevated levels of necroptosis and that inhibition of this cell death process fully rescued the survival and proliferation of these cells. Collectively, our data demonstrate that inactivation of caspase-8 suppresses the survival and proliferative capacity of Bim(-/-) T cells and restrains autoimmunity in Bim(-/-) mice.

Show MeSH

Related in: MedlinePlus

Impaired proliferation and increased necroptosis in tcasp8−/−Bim−/− T cells. (A) [3H]thymidine incorporation of purified T cells from mice of the indicated genotypes after 24, 48, or 72 h of stimulation with anti-CD3 antibody with or without costimulation with anti-CD28 antibody or IL-2. Representative data are shown from three independent experiments of young mice. (B) Representative TEM images of T cells activated for 24 h with anti-CD3/anti-CD28 antibody, showing increased frequency of necroptotic cells in cultures of anti-CD3/anti-CD28 antibody stimulated tcasp8−/− and tcasp8−/−Bim−/− T cells. Bar, 10 µm. (C) The percentages of necroptotic primary and anti-CD3 antibody–activated T cells identified by electron microscopy. 125–209 untreated or activated T cells were scored for each genotype. *, P < 0.05 compared with WT and Bim−/− control mice. (D) Representative immunoblots of the levels of LC3-I and LC3-II in freshly isolated T cells (0 h) and T cells cultured for 24 h in the absence or presence of anti-CD3/anti-CD28 antibody. (E) FACS analysis of CFSE dilution profiles. Proliferation of CFSE-labeled T cells was examined after a 3-d activation with anti-CD3/anti-CD28 antibodies with or without 10 µM Nec1. Histograms represent the mean ± SEM (error bars).
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC3198166&req=5

fig5: Impaired proliferation and increased necroptosis in tcasp8−/−Bim−/− T cells. (A) [3H]thymidine incorporation of purified T cells from mice of the indicated genotypes after 24, 48, or 72 h of stimulation with anti-CD3 antibody with or without costimulation with anti-CD28 antibody or IL-2. Representative data are shown from three independent experiments of young mice. (B) Representative TEM images of T cells activated for 24 h with anti-CD3/anti-CD28 antibody, showing increased frequency of necroptotic cells in cultures of anti-CD3/anti-CD28 antibody stimulated tcasp8−/− and tcasp8−/−Bim−/− T cells. Bar, 10 µm. (C) The percentages of necroptotic primary and anti-CD3 antibody–activated T cells identified by electron microscopy. 125–209 untreated or activated T cells were scored for each genotype. *, P < 0.05 compared with WT and Bim−/− control mice. (D) Representative immunoblots of the levels of LC3-I and LC3-II in freshly isolated T cells (0 h) and T cells cultured for 24 h in the absence or presence of anti-CD3/anti-CD28 antibody. (E) FACS analysis of CFSE dilution profiles. Proliferation of CFSE-labeled T cells was examined after a 3-d activation with anti-CD3/anti-CD28 antibodies with or without 10 µM Nec1. Histograms represent the mean ± SEM (error bars).

Mentions: Loss of caspase-8 or its adaptor Fadd impairs proliferation of T cells in response to antigen or mitogen stimulation (Newton et al., 1998; Zhang et al., 1998; Salmena et al., 2003). Because loss of caspase-8 also increases the death of T cells (Salmena et al., 2003) and loss of Bim inhibits T cell apoptosis (Bouillet et al., 1999), we investigated whether loss of Bim could rescue the proliferative defects of tcasp8−/− T cells. In response to anti-CD3 antibody alone or together with anti-CD28 antibody or exogenous IL-2, purified tcasp8−/−Bim−/− T cells, similar to tcasp8−/− T cells, displayed decreased levels of [3H]thymidine incorporation compared with Bim−/− and WT T cells (Fig. 5 A). As previously reported (Salmena et al., 2003), sub-G1 population increased in anti-CD3/anti-CD28–stimulated tcasp8−/− T cells compared with WT controls, which indicates increased cell death of T cells deficient for caspase-8 (Fig. S4 A). Sub-G1 population was also increased in anti-CD3/anti-CD28–stimulated tcasp8−/−Bim−/− T cells compared with WT and Bim−/− controls (Fig. S4 A). These results demonstrate that loss of Bim is not sufficient to rescue T cell proliferative defects associated with caspase-8 deficiency.


Caspase-8 inactivation in T cells increases necroptosis and suppresses autoimmunity in Bim-/- mice.

Bohgaki T, Mozo J, Salmena L, Matysiak-Zablocki E, Bohgaki M, Sanchez O, Strasser A, Hakem A, Hakem R - J. Cell Biol. (2011)

Impaired proliferation and increased necroptosis in tcasp8−/−Bim−/− T cells. (A) [3H]thymidine incorporation of purified T cells from mice of the indicated genotypes after 24, 48, or 72 h of stimulation with anti-CD3 antibody with or without costimulation with anti-CD28 antibody or IL-2. Representative data are shown from three independent experiments of young mice. (B) Representative TEM images of T cells activated for 24 h with anti-CD3/anti-CD28 antibody, showing increased frequency of necroptotic cells in cultures of anti-CD3/anti-CD28 antibody stimulated tcasp8−/− and tcasp8−/−Bim−/− T cells. Bar, 10 µm. (C) The percentages of necroptotic primary and anti-CD3 antibody–activated T cells identified by electron microscopy. 125–209 untreated or activated T cells were scored for each genotype. *, P < 0.05 compared with WT and Bim−/− control mice. (D) Representative immunoblots of the levels of LC3-I and LC3-II in freshly isolated T cells (0 h) and T cells cultured for 24 h in the absence or presence of anti-CD3/anti-CD28 antibody. (E) FACS analysis of CFSE dilution profiles. Proliferation of CFSE-labeled T cells was examined after a 3-d activation with anti-CD3/anti-CD28 antibodies with or without 10 µM Nec1. Histograms represent the mean ± SEM (error bars).
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3198166&req=5

fig5: Impaired proliferation and increased necroptosis in tcasp8−/−Bim−/− T cells. (A) [3H]thymidine incorporation of purified T cells from mice of the indicated genotypes after 24, 48, or 72 h of stimulation with anti-CD3 antibody with or without costimulation with anti-CD28 antibody or IL-2. Representative data are shown from three independent experiments of young mice. (B) Representative TEM images of T cells activated for 24 h with anti-CD3/anti-CD28 antibody, showing increased frequency of necroptotic cells in cultures of anti-CD3/anti-CD28 antibody stimulated tcasp8−/− and tcasp8−/−Bim−/− T cells. Bar, 10 µm. (C) The percentages of necroptotic primary and anti-CD3 antibody–activated T cells identified by electron microscopy. 125–209 untreated or activated T cells were scored for each genotype. *, P < 0.05 compared with WT and Bim−/− control mice. (D) Representative immunoblots of the levels of LC3-I and LC3-II in freshly isolated T cells (0 h) and T cells cultured for 24 h in the absence or presence of anti-CD3/anti-CD28 antibody. (E) FACS analysis of CFSE dilution profiles. Proliferation of CFSE-labeled T cells was examined after a 3-d activation with anti-CD3/anti-CD28 antibodies with or without 10 µM Nec1. Histograms represent the mean ± SEM (error bars).
Mentions: Loss of caspase-8 or its adaptor Fadd impairs proliferation of T cells in response to antigen or mitogen stimulation (Newton et al., 1998; Zhang et al., 1998; Salmena et al., 2003). Because loss of caspase-8 also increases the death of T cells (Salmena et al., 2003) and loss of Bim inhibits T cell apoptosis (Bouillet et al., 1999), we investigated whether loss of Bim could rescue the proliferative defects of tcasp8−/− T cells. In response to anti-CD3 antibody alone or together with anti-CD28 antibody or exogenous IL-2, purified tcasp8−/−Bim−/− T cells, similar to tcasp8−/− T cells, displayed decreased levels of [3H]thymidine incorporation compared with Bim−/− and WT T cells (Fig. 5 A). As previously reported (Salmena et al., 2003), sub-G1 population increased in anti-CD3/anti-CD28–stimulated tcasp8−/− T cells compared with WT controls, which indicates increased cell death of T cells deficient for caspase-8 (Fig. S4 A). Sub-G1 population was also increased in anti-CD3/anti-CD28–stimulated tcasp8−/−Bim−/− T cells compared with WT and Bim−/− controls (Fig. S4 A). These results demonstrate that loss of Bim is not sufficient to rescue T cell proliferative defects associated with caspase-8 deficiency.

Bottom Line: Dysregulation of either the extrinsic or intrinsic apoptotic pathway can lead to various diseases including immune disorders and cancer.In addition to its role in the extrinsic apoptotic pathway, caspase-8 plays nonapoptotic functions and is essential for T cell homeostasis.We show that, similar to caspase-8(-/-) T cells, Bim(-/-) T cells that also lack caspase-8 displayed elevated levels of necroptosis and that inhibition of this cell death process fully rescued the survival and proliferation of these cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G 2M9, Canada.

ABSTRACT
Dysregulation of either the extrinsic or intrinsic apoptotic pathway can lead to various diseases including immune disorders and cancer. In addition to its role in the extrinsic apoptotic pathway, caspase-8 plays nonapoptotic functions and is essential for T cell homeostasis. The pro-apoptotic BH3-only Bcl-2 family member Bim is important for the intrinsic apoptotic pathway and its inactivation leads to autoimmunity that is further exacerbated by loss of function of the death receptor Fas. We report that inactivation of caspase-8 in T cells of Bim(-/-) mice restrained their autoimmunity and extended their life span. We show that, similar to caspase-8(-/-) T cells, Bim(-/-) T cells that also lack caspase-8 displayed elevated levels of necroptosis and that inhibition of this cell death process fully rescued the survival and proliferation of these cells. Collectively, our data demonstrate that inactivation of caspase-8 suppresses the survival and proliferative capacity of Bim(-/-) T cells and restrains autoimmunity in Bim(-/-) mice.

Show MeSH
Related in: MedlinePlus