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Conditions that induce tolerance in mature CD4+ T cells.

Lanoue A, Bona C, von Boehmer H, Sarukhan A - J. Exp. Med. (1997)

Bottom Line: Mature CD4+6.5+ cells that were transferred into antigen-containing recipients went through an initial phase of expansion after which most cells were deleted and those remaining became unresponsive, as previously described for CD8+ cells.It was only after decreasing cell numbers by CD4 antibody treatment and by repeatedly reintroducing antigen thereafter that unresponsiveness of 6.5+ cells was achieved and maintained.In no case could we observe the appearance of antigen-specific T cells with a Th2 cytokine profile among the remaining cells and therefore conclude that deletion and anergy represent the major mechanisms of tolerance in our studies.

View Article: PubMed Central - PubMed

Affiliation: Unité Institut National de la Santé et de la Recherche Médicale 373, Institut Necker, Paris.

ABSTRACT
Establishment of antigen-specific tolerance among mature T cells has been a long debated, yet poorly understood issue. In this study we have used transgenic mice bearing a class II--restricted TCR specific for the hemmagglutinin of the influenza virus in order to test the behavior of CD4+ T cells upon exposure to antigen in different forms and doses. We first studied the fate of T cells expressing the transgenic TCR (6.5) in double transgenic mice where HA was expressed as a self antigen by hemapoietic cells. In these mice, we found some mature T cells in periphery that had escaped thymic deletion and that showed signs of activation but which were anergic. Mature CD4+6.5+ cells that were transferred into antigen-containing recipients went through an initial phase of expansion after which most cells were deleted and those remaining became unresponsive, as previously described for CD8+ cells. Inducing tolerance in CD4+6.5+ cells in situ in single transgenic mice proved a difficult task: classical protocols using single doses of soluble or deaggregated antigen as well as feeding antigen all failed to induce antigen-specific unresponsiveness. It was only after decreasing cell numbers by CD4 antibody treatment and by repeatedly reintroducing antigen thereafter that unresponsiveness of 6.5+ cells was achieved and maintained. In no case could we observe the appearance of antigen-specific T cells with a Th2 cytokine profile among the remaining cells and therefore conclude that deletion and anergy represent the major mechanisms of tolerance in our studies.

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Repeated antigen administration in mice previously  treated with anti-CD4 plus antigen. Mice having received 50 μg  of YTA3.1.2 plus 0.1 mg of HAIg 7 d earlier were repeatedly injected i.v. with 0.05, 0.1, or 0.5  mg of HA-Ig. For the first two  doses, four injections were  given every third day. For the  highest dose, two injections were  given, also every third day. Mice  were killed 7 d after the last injection. Control mice received  either PBS or anti-CD4 plus antigen only. Absolute numbers of  6.5+ cells in the thymus (A) and  in the periphery (B) are shown.  Proliferation values of sIg−  splenic cells upon in vitro antigenic stimulation with peptide  were calculated to correspond to  the response of 103 6.5+ cells (C).  Percentages of 6.5+ cells among  total T cells are also shown (D).
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Figure 6: Repeated antigen administration in mice previously treated with anti-CD4 plus antigen. Mice having received 50 μg of YTA3.1.2 plus 0.1 mg of HAIg 7 d earlier were repeatedly injected i.v. with 0.05, 0.1, or 0.5 mg of HA-Ig. For the first two doses, four injections were given every third day. For the highest dose, two injections were given, also every third day. Mice were killed 7 d after the last injection. Control mice received either PBS or anti-CD4 plus antigen only. Absolute numbers of 6.5+ cells in the thymus (A) and in the periphery (B) are shown. Proliferation values of sIg− splenic cells upon in vitro antigenic stimulation with peptide were calculated to correspond to the response of 103 6.5+ cells (C). Percentages of 6.5+ cells among total T cells are also shown (D).

Mentions: Since the above protocol was efficient in inducing unresponsiveness but not in maintaining it, we reasoned that administering antigen after such treatment in a manner that would favor its persistence could help to maintain the tolerant state and would render the treatment more antigen-specific. Therefore, mice treated 7 d earlier with anti-CD4 plus 0.1 mg of HA-Ig received further injections of antigen. Different doses of HA-Ig (ranging from 50 to 500 μg) were injected seven days after treatment with CD4 antibody and 0.1 mg HA-Ig in 3-d intervals. All mice were analyzed 7 d after the last injection of antigen. As show in Fig. 6 there was a further reduction in number of 6.5 cells in an antigen-dose dependent manner in mice that received antigen in addition to the initial CD4 treatment. Significantly lowering the numbers of CD4+ cells by the CD4 antibody avoided the splenomegaly that was otherwise seen with repeated doses of HA-Ig. Likewise, the proliferative capacity of CD4+ cells to HA was reduced in a dose-dependent manner and this reduction lasted for long periods of time (30 d) in thymectomized mice (data not shown). There was an increase in the proportion of 6.5+CD69+ and of 6.5+ CD62-L− cells that correlated with antigen doses (Fig. 7). The 6.5+ cells recovered from the mice that repeatedly received either 0.5 or 0.1 mg of HA-Ig secreted very low levels of IFN-γ (less than one-tenth of what was produced by the control mice) and produced no IL4 (data not shown). It is interesting to note that while IL4 production was detected after transfer of 6.5 cells into antigen-bearing recipients (Table 1), it could not be detected after any of the protocols aiming at tolerance induction in situ. This difference may be due to the fact that, upon transfer into antigen-containing recipients, cells were exposed to much larger doses of antigen and for a longer period of time.


Conditions that induce tolerance in mature CD4+ T cells.

Lanoue A, Bona C, von Boehmer H, Sarukhan A - J. Exp. Med. (1997)

Repeated antigen administration in mice previously  treated with anti-CD4 plus antigen. Mice having received 50 μg  of YTA3.1.2 plus 0.1 mg of HAIg 7 d earlier were repeatedly injected i.v. with 0.05, 0.1, or 0.5  mg of HA-Ig. For the first two  doses, four injections were  given every third day. For the  highest dose, two injections were  given, also every third day. Mice  were killed 7 d after the last injection. Control mice received  either PBS or anti-CD4 plus antigen only. Absolute numbers of  6.5+ cells in the thymus (A) and  in the periphery (B) are shown.  Proliferation values of sIg−  splenic cells upon in vitro antigenic stimulation with peptide  were calculated to correspond to  the response of 103 6.5+ cells (C).  Percentages of 6.5+ cells among  total T cells are also shown (D).
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Related In: Results  -  Collection

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Figure 6: Repeated antigen administration in mice previously treated with anti-CD4 plus antigen. Mice having received 50 μg of YTA3.1.2 plus 0.1 mg of HAIg 7 d earlier were repeatedly injected i.v. with 0.05, 0.1, or 0.5 mg of HA-Ig. For the first two doses, four injections were given every third day. For the highest dose, two injections were given, also every third day. Mice were killed 7 d after the last injection. Control mice received either PBS or anti-CD4 plus antigen only. Absolute numbers of 6.5+ cells in the thymus (A) and in the periphery (B) are shown. Proliferation values of sIg− splenic cells upon in vitro antigenic stimulation with peptide were calculated to correspond to the response of 103 6.5+ cells (C). Percentages of 6.5+ cells among total T cells are also shown (D).
Mentions: Since the above protocol was efficient in inducing unresponsiveness but not in maintaining it, we reasoned that administering antigen after such treatment in a manner that would favor its persistence could help to maintain the tolerant state and would render the treatment more antigen-specific. Therefore, mice treated 7 d earlier with anti-CD4 plus 0.1 mg of HA-Ig received further injections of antigen. Different doses of HA-Ig (ranging from 50 to 500 μg) were injected seven days after treatment with CD4 antibody and 0.1 mg HA-Ig in 3-d intervals. All mice were analyzed 7 d after the last injection of antigen. As show in Fig. 6 there was a further reduction in number of 6.5 cells in an antigen-dose dependent manner in mice that received antigen in addition to the initial CD4 treatment. Significantly lowering the numbers of CD4+ cells by the CD4 antibody avoided the splenomegaly that was otherwise seen with repeated doses of HA-Ig. Likewise, the proliferative capacity of CD4+ cells to HA was reduced in a dose-dependent manner and this reduction lasted for long periods of time (30 d) in thymectomized mice (data not shown). There was an increase in the proportion of 6.5+CD69+ and of 6.5+ CD62-L− cells that correlated with antigen doses (Fig. 7). The 6.5+ cells recovered from the mice that repeatedly received either 0.5 or 0.1 mg of HA-Ig secreted very low levels of IFN-γ (less than one-tenth of what was produced by the control mice) and produced no IL4 (data not shown). It is interesting to note that while IL4 production was detected after transfer of 6.5 cells into antigen-bearing recipients (Table 1), it could not be detected after any of the protocols aiming at tolerance induction in situ. This difference may be due to the fact that, upon transfer into antigen-containing recipients, cells were exposed to much larger doses of antigen and for a longer period of time.

Bottom Line: Mature CD4+6.5+ cells that were transferred into antigen-containing recipients went through an initial phase of expansion after which most cells were deleted and those remaining became unresponsive, as previously described for CD8+ cells.It was only after decreasing cell numbers by CD4 antibody treatment and by repeatedly reintroducing antigen thereafter that unresponsiveness of 6.5+ cells was achieved and maintained.In no case could we observe the appearance of antigen-specific T cells with a Th2 cytokine profile among the remaining cells and therefore conclude that deletion and anergy represent the major mechanisms of tolerance in our studies.

View Article: PubMed Central - PubMed

Affiliation: Unité Institut National de la Santé et de la Recherche Médicale 373, Institut Necker, Paris.

ABSTRACT
Establishment of antigen-specific tolerance among mature T cells has been a long debated, yet poorly understood issue. In this study we have used transgenic mice bearing a class II--restricted TCR specific for the hemmagglutinin of the influenza virus in order to test the behavior of CD4+ T cells upon exposure to antigen in different forms and doses. We first studied the fate of T cells expressing the transgenic TCR (6.5) in double transgenic mice where HA was expressed as a self antigen by hemapoietic cells. In these mice, we found some mature T cells in periphery that had escaped thymic deletion and that showed signs of activation but which were anergic. Mature CD4+6.5+ cells that were transferred into antigen-containing recipients went through an initial phase of expansion after which most cells were deleted and those remaining became unresponsive, as previously described for CD8+ cells. Inducing tolerance in CD4+6.5+ cells in situ in single transgenic mice proved a difficult task: classical protocols using single doses of soluble or deaggregated antigen as well as feeding antigen all failed to induce antigen-specific unresponsiveness. It was only after decreasing cell numbers by CD4 antibody treatment and by repeatedly reintroducing antigen thereafter that unresponsiveness of 6.5+ cells was achieved and maintained. In no case could we observe the appearance of antigen-specific T cells with a Th2 cytokine profile among the remaining cells and therefore conclude that deletion and anergy represent the major mechanisms of tolerance in our studies.

Show MeSH
Related in: MedlinePlus