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T helper subset differentiation in the absence of invariant chain.

Brown DR, Swier K, Moskowitz NH, Naujokas MF, Locksley RM, Reiner SL - J. Exp. Med. (1997)

Bottom Line: We wished to determine whether this unexpected efficiency of T helper subset induction despite inefficient T cell stimulation could be modeled in vitro.In the presence of rIL-12 or rIL-4 naive parasite-specific transgenic T cells could mature into IFN-gamma-or IL-4-secreting T helper cells, respectively, even when antigen presentation was suboptimal or antigen dose was submitogenic.These experiments demonstrate that activation of T helper cells to a threshold required for IL-2 production or proliferation is not required to achieve induction of disease-regulating T helper cell effector functions, and that pathogen-associated secondary activation signals may facilitate the full differentiation of T helper subsets during limiting presentation of antigenic peptides.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Gwen Knapp Center for Lupus & Immunology Research, University of Chicago, Illinois 60637, USA.

ABSTRACT
The outcome of murine infection with Leishmania major is regulated by major histocompatibility complex class II-restricted T helper cells. Invariant chain-deficient (Ii -/-) mice have impaired ability to present major histocompatibility complex class II-restricted antigens, and reduced numbers of CD4+ T cells. Despite these deficits, C57BL/6 Ii -/- mice controlled L. major infection comparably to wild-type mice. As assessed by mRNA analysis and in vitro antigen restimulation for IFN-gamma, Ii -/- mice had normal induction of Th1 subset differentiation even though antigen-dependent proliferation of their lymph node cells was substantially compromised. In addition, BALB/c Ii -/- mice exhibited a progressive course of infection and Th2 effector cell development that were comparable to that seen in wild-type BALB/c mice. We wished to determine whether this unexpected efficiency of T helper subset induction despite inefficient T cell stimulation could be modeled in vitro. In the presence of rIL-12 or rIL-4 naive parasite-specific transgenic T cells could mature into IFN-gamma-or IL-4-secreting T helper cells, respectively, even when antigen presentation was suboptimal or antigen dose was submitogenic. These experiments demonstrate that activation of T helper cells to a threshold required for IL-2 production or proliferation is not required to achieve induction of disease-regulating T helper cell effector functions, and that pathogen-associated secondary activation signals may facilitate the full differentiation of T helper subsets during limiting presentation of antigenic peptides.

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IL-12 can maintain IFN-γ production and implement Th1 development by T cells primed on Ii −/− APCs. (A) Irradiated spleen cells (105/ well) from Ii +− (filled symbols) or Ii −/− (open symbols) H-2d littermates were incubated with varying amounts of recombinant LACK Ag in the presence of 2 × 105 TCR transgenic lymph node cells and 2 ng/ml of rIL-12. After 48 h, supernatants were assayed for IL-2 and IFN-γ using ELISA. Values  represent means and standard deviations from triplicate determinations, and are representative of four separate experiments. (B) B220+ cell–depleted  TCR transgenic lymph node cells (2 × 105 cells/well) were stimulated in a 1 ml volume in 24-well plates with either Ii + or Ii −/− irradiated H-2d  spleen cells (1 × 106 cells/well), plus recombinant LACK antigen (2 μg/ml), with (+) or without (−) rIL-12 (0.5 ng/ml) as designated on the x-axis.  6 d later, viable cells were purified and washed, and 2 × 105 T cells were restimulated with the same Ii-genotype APCs (1.2 × 106/well) used in the primary stimulation in the presence of antigen (2 μg/ml), but without rIL-12. After 48 h, supernatants were collected and assayed for IL-2 and IFN-γ by  ELISA. Values represent means and standard deviations of triplicate determinations, and are representative of two separate experiments.
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Figure 8: IL-12 can maintain IFN-γ production and implement Th1 development by T cells primed on Ii −/− APCs. (A) Irradiated spleen cells (105/ well) from Ii +− (filled symbols) or Ii −/− (open symbols) H-2d littermates were incubated with varying amounts of recombinant LACK Ag in the presence of 2 × 105 TCR transgenic lymph node cells and 2 ng/ml of rIL-12. After 48 h, supernatants were assayed for IL-2 and IFN-γ using ELISA. Values represent means and standard deviations from triplicate determinations, and are representative of four separate experiments. (B) B220+ cell–depleted TCR transgenic lymph node cells (2 × 105 cells/well) were stimulated in a 1 ml volume in 24-well plates with either Ii + or Ii −/− irradiated H-2d spleen cells (1 × 106 cells/well), plus recombinant LACK antigen (2 μg/ml), with (+) or without (−) rIL-12 (0.5 ng/ml) as designated on the x-axis. 6 d later, viable cells were purified and washed, and 2 × 105 T cells were restimulated with the same Ii-genotype APCs (1.2 × 106/well) used in the primary stimulation in the presence of antigen (2 μg/ml), but without rIL-12. After 48 h, supernatants were collected and assayed for IL-2 and IFN-γ by ELISA. Values represent means and standard deviations of triplicate determinations, and are representative of two separate experiments.

Mentions: To confirm that rIL-12 could similarly sustain IFN-γ production from T cells primed on Ii −/− APCs, Ii +/− or −/− APCs were incubated with varying concentrations of the recombinant LACK antigen and transgenic T cells in the presence of rIL-12. After 48 h, supernatants were analyzed for IL-2 and IFN-γ (Fig. 8 A). Consistent with the earlier results using normal APCs under conditions of low antigenic stimulation, T cells primed on Ii −/− APCs generated IFN-γ that was comparable to that generated by T cells primed on Ii +/− APCs, although only the latter generated IL-2 that could be recovered in supernatants. Recovery of IFN-γ was dependent on both antigen and rIL-12; in the absence of either, no cytokine could be recovered from transgenic T cells primed on Ii −/− APCs. This experiment additionally confirms that the LACK protein antigen, in contrast to the immunogenic LACK peptide (Fig. 5), requires Ii for optimal presentation.


T helper subset differentiation in the absence of invariant chain.

Brown DR, Swier K, Moskowitz NH, Naujokas MF, Locksley RM, Reiner SL - J. Exp. Med. (1997)

IL-12 can maintain IFN-γ production and implement Th1 development by T cells primed on Ii −/− APCs. (A) Irradiated spleen cells (105/ well) from Ii +− (filled symbols) or Ii −/− (open symbols) H-2d littermates were incubated with varying amounts of recombinant LACK Ag in the presence of 2 × 105 TCR transgenic lymph node cells and 2 ng/ml of rIL-12. After 48 h, supernatants were assayed for IL-2 and IFN-γ using ELISA. Values  represent means and standard deviations from triplicate determinations, and are representative of four separate experiments. (B) B220+ cell–depleted  TCR transgenic lymph node cells (2 × 105 cells/well) were stimulated in a 1 ml volume in 24-well plates with either Ii + or Ii −/− irradiated H-2d  spleen cells (1 × 106 cells/well), plus recombinant LACK antigen (2 μg/ml), with (+) or without (−) rIL-12 (0.5 ng/ml) as designated on the x-axis.  6 d later, viable cells were purified and washed, and 2 × 105 T cells were restimulated with the same Ii-genotype APCs (1.2 × 106/well) used in the primary stimulation in the presence of antigen (2 μg/ml), but without rIL-12. After 48 h, supernatants were collected and assayed for IL-2 and IFN-γ by  ELISA. Values represent means and standard deviations of triplicate determinations, and are representative of two separate experiments.
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Figure 8: IL-12 can maintain IFN-γ production and implement Th1 development by T cells primed on Ii −/− APCs. (A) Irradiated spleen cells (105/ well) from Ii +− (filled symbols) or Ii −/− (open symbols) H-2d littermates were incubated with varying amounts of recombinant LACK Ag in the presence of 2 × 105 TCR transgenic lymph node cells and 2 ng/ml of rIL-12. After 48 h, supernatants were assayed for IL-2 and IFN-γ using ELISA. Values represent means and standard deviations from triplicate determinations, and are representative of four separate experiments. (B) B220+ cell–depleted TCR transgenic lymph node cells (2 × 105 cells/well) were stimulated in a 1 ml volume in 24-well plates with either Ii + or Ii −/− irradiated H-2d spleen cells (1 × 106 cells/well), plus recombinant LACK antigen (2 μg/ml), with (+) or without (−) rIL-12 (0.5 ng/ml) as designated on the x-axis. 6 d later, viable cells were purified and washed, and 2 × 105 T cells were restimulated with the same Ii-genotype APCs (1.2 × 106/well) used in the primary stimulation in the presence of antigen (2 μg/ml), but without rIL-12. After 48 h, supernatants were collected and assayed for IL-2 and IFN-γ by ELISA. Values represent means and standard deviations of triplicate determinations, and are representative of two separate experiments.
Mentions: To confirm that rIL-12 could similarly sustain IFN-γ production from T cells primed on Ii −/− APCs, Ii +/− or −/− APCs were incubated with varying concentrations of the recombinant LACK antigen and transgenic T cells in the presence of rIL-12. After 48 h, supernatants were analyzed for IL-2 and IFN-γ (Fig. 8 A). Consistent with the earlier results using normal APCs under conditions of low antigenic stimulation, T cells primed on Ii −/− APCs generated IFN-γ that was comparable to that generated by T cells primed on Ii +/− APCs, although only the latter generated IL-2 that could be recovered in supernatants. Recovery of IFN-γ was dependent on both antigen and rIL-12; in the absence of either, no cytokine could be recovered from transgenic T cells primed on Ii −/− APCs. This experiment additionally confirms that the LACK protein antigen, in contrast to the immunogenic LACK peptide (Fig. 5), requires Ii for optimal presentation.

Bottom Line: We wished to determine whether this unexpected efficiency of T helper subset induction despite inefficient T cell stimulation could be modeled in vitro.In the presence of rIL-12 or rIL-4 naive parasite-specific transgenic T cells could mature into IFN-gamma-or IL-4-secreting T helper cells, respectively, even when antigen presentation was suboptimal or antigen dose was submitogenic.These experiments demonstrate that activation of T helper cells to a threshold required for IL-2 production or proliferation is not required to achieve induction of disease-regulating T helper cell effector functions, and that pathogen-associated secondary activation signals may facilitate the full differentiation of T helper subsets during limiting presentation of antigenic peptides.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Gwen Knapp Center for Lupus & Immunology Research, University of Chicago, Illinois 60637, USA.

ABSTRACT
The outcome of murine infection with Leishmania major is regulated by major histocompatibility complex class II-restricted T helper cells. Invariant chain-deficient (Ii -/-) mice have impaired ability to present major histocompatibility complex class II-restricted antigens, and reduced numbers of CD4+ T cells. Despite these deficits, C57BL/6 Ii -/- mice controlled L. major infection comparably to wild-type mice. As assessed by mRNA analysis and in vitro antigen restimulation for IFN-gamma, Ii -/- mice had normal induction of Th1 subset differentiation even though antigen-dependent proliferation of their lymph node cells was substantially compromised. In addition, BALB/c Ii -/- mice exhibited a progressive course of infection and Th2 effector cell development that were comparable to that seen in wild-type BALB/c mice. We wished to determine whether this unexpected efficiency of T helper subset induction despite inefficient T cell stimulation could be modeled in vitro. In the presence of rIL-12 or rIL-4 naive parasite-specific transgenic T cells could mature into IFN-gamma-or IL-4-secreting T helper cells, respectively, even when antigen presentation was suboptimal or antigen dose was submitogenic. These experiments demonstrate that activation of T helper cells to a threshold required for IL-2 production or proliferation is not required to achieve induction of disease-regulating T helper cell effector functions, and that pathogen-associated secondary activation signals may facilitate the full differentiation of T helper subsets during limiting presentation of antigenic peptides.

Show MeSH
Related in: MedlinePlus