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Telomere erosion in memory T cells induced by telomerase inhibition at the site of antigenic challenge in vivo.

Reed JR, Vukmanovic-Stejic M, Fletcher JM, Soares MV, Cook JE, Orteu CH, Jackson SE, Birch KE, Foster GR, Salmon M, Beverley PC, Rustin MH, Akbar AN - J. Exp. Med. (2004)

Bottom Line: Furthermore, significant telomere erosion occurred in specific T cells that respond in the skin, but not in those that are found in the blood from the same individuals.Antibody inhibition studies indicated that type I interferon (IFN), which was identified at high levels in the tissue fluid and by immunohistology, was responsible in part for the telomerase inhibition.Furthermore, the addition of IFN-alpha to PPD-stimulated CD4+ T cells directly inhibited telomerase activity in vitro.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Immunology and Molecular Pathology, Div. of Infection and Immunity, University College London, 46 Cleveland St., London W1T 4JF, England, UK.

ABSTRACT
The extent of human memory T cell proliferation, differentiation, and telomere erosion that occurs after a single episode of immune challenge in vivo is unclear. To investigate this, we injected tuberculin purified protein derivative (PPD) into the skin of immune individuals and isolated responsive T cells from the site of antigenic challenge at different times. PPD-specific CD4+ T cells proliferated and differentiated extensively in the skin during this secondary response. Furthermore, significant telomere erosion occurred in specific T cells that respond in the skin, but not in those that are found in the blood from the same individuals. Tissue fluid obtained from the site of PPD challenge in the skin inhibited the induction of the enzyme telomerase in T cells in vitro. Antibody inhibition studies indicated that type I interferon (IFN), which was identified at high levels in the tissue fluid and by immunohistology, was responsible in part for the telomerase inhibition. Furthermore, the addition of IFN-alpha to PPD-stimulated CD4+ T cells directly inhibited telomerase activity in vitro. Therefore, these results suggest that the rate of telomere erosion in proliferating, antigen-specific CD4+ T cells may be accelerated by type I IFN during a secondary response in vivo.

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Type I IFN in blister fluid inhibits telomerase activity in PPD-stimulated T cells. IFN-α–producing cells present in day 3 MT skin sections were identified using indirect alkaline phosphatase staining (a, top). The bottom panel shows staining with an isotype control (magnification, 20). Keratinocytes (K) and isolated cells within the perivascular infiltrates (PV; indicated by arrows) expressed IFN-α. (b) The presence of type I IFN in blister fluid collected at different times after MT induction. The data represent the mean ± SEM of type I IFN in blister samples from five different individuals assayed per time point. Type I IFN mediates the suppression of telomerase activity induced by day 3 blister fluid (c). PBMCs were stimulated with PPD in vitro in the presence of autologous day 3 blister fluid (+MT BF) or autologous serum (+serum). Blocking antibody reactive with type I IFN receptor 2 (CD118; +anti-IFN R) or isotype control antibody (+cnt Ab) were added to cultures stimulated in the presence of MT blister fluid. Samples were collected 3 d after stimulation, and samples were adjusted to 500 proliferating Ki67+ T cells per reaction for telomerase analysis (one out of three representative experiments is shown). Direct inhibition of telomerase activity by type I IFN (d). A PPD-specific CD4+ T cell line was restimulated with PPD in the presence of recombinant IFN-α (black bars) or Roferon (white bars), a clinical preparation of IFN-α for 4 d. Samples were adjusted so that telomerase activity was measured in an equivalent number of cycling (Ki67+) T cells (one of three representative experiments is shown). Blister-derived CD4+ T cells can expand in vitro (e). Cells recovered from a day-19 blister were stimulated in vitro with PPD-pulsed irradiated autologous PBMCs as APCs. Arrows indicate each point of restimulation in vitro. The asterisks indicate points when samples were collected to measure telomerase activity. Telomerase activity of CD4+ T cells isolated from day 19 blisters decreases with repeated stimulation in vitro (f). Samples were collected on day 4 after each point of PPD restimulation. Telomerase activity per 500 Ki67+ T cells was determined as before. Stim 4, Stim 8, and Stim 9 indicate the 4th, 8th, and 9th stimulations with PPD and irradiated APCs in vitro.
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fig6: Type I IFN in blister fluid inhibits telomerase activity in PPD-stimulated T cells. IFN-α–producing cells present in day 3 MT skin sections were identified using indirect alkaline phosphatase staining (a, top). The bottom panel shows staining with an isotype control (magnification, 20). Keratinocytes (K) and isolated cells within the perivascular infiltrates (PV; indicated by arrows) expressed IFN-α. (b) The presence of type I IFN in blister fluid collected at different times after MT induction. The data represent the mean ± SEM of type I IFN in blister samples from five different individuals assayed per time point. Type I IFN mediates the suppression of telomerase activity induced by day 3 blister fluid (c). PBMCs were stimulated with PPD in vitro in the presence of autologous day 3 blister fluid (+MT BF) or autologous serum (+serum). Blocking antibody reactive with type I IFN receptor 2 (CD118; +anti-IFN R) or isotype control antibody (+cnt Ab) were added to cultures stimulated in the presence of MT blister fluid. Samples were collected 3 d after stimulation, and samples were adjusted to 500 proliferating Ki67+ T cells per reaction for telomerase analysis (one out of three representative experiments is shown). Direct inhibition of telomerase activity by type I IFN (d). A PPD-specific CD4+ T cell line was restimulated with PPD in the presence of recombinant IFN-α (black bars) or Roferon (white bars), a clinical preparation of IFN-α for 4 d. Samples were adjusted so that telomerase activity was measured in an equivalent number of cycling (Ki67+) T cells (one of three representative experiments is shown). Blister-derived CD4+ T cells can expand in vitro (e). Cells recovered from a day-19 blister were stimulated in vitro with PPD-pulsed irradiated autologous PBMCs as APCs. Arrows indicate each point of restimulation in vitro. The asterisks indicate points when samples were collected to measure telomerase activity. Telomerase activity of CD4+ T cells isolated from day 19 blisters decreases with repeated stimulation in vitro (f). Samples were collected on day 4 after each point of PPD restimulation. Telomerase activity per 500 Ki67+ T cells was determined as before. Stim 4, Stim 8, and Stim 9 indicate the 4th, 8th, and 9th stimulations with PPD and irradiated APCs in vitro.

Mentions: Our previous studies indicate that type I IFN (IFN-α and IFN-β) can regulate the rate of differentiation of human T cells (32). Together with reports that IFN-α can inhibit the activity of telomerase in tumor cell lines in vitro (33), this prompted us to determine whether this group of cytokines was responsible for the inhibition of telomerase activity in PPD-specific CD4+ T cells in the skin in situ. By immunohistological analysis, we identified cells that expressed high levels of IFN-α in the perivascular infiltrates of skin sections 3 d after PPD challenge (Fig. 6 a). High uniform levels of expression were also detected in keratinocytes (Fig. 6 a). This was confirmed by measurement of peak concentrations of type I IFN in SB fluid that was obtained at day 3 after PPD injection (Fig. 6 b). This coincided with the time at which telomerase was greatest in CD4+ T cells that were PPD-stimulated in vitro, but absent from cycling CD4+ T cells in the skin (Fig. 5 b).


Telomere erosion in memory T cells induced by telomerase inhibition at the site of antigenic challenge in vivo.

Reed JR, Vukmanovic-Stejic M, Fletcher JM, Soares MV, Cook JE, Orteu CH, Jackson SE, Birch KE, Foster GR, Salmon M, Beverley PC, Rustin MH, Akbar AN - J. Exp. Med. (2004)

Type I IFN in blister fluid inhibits telomerase activity in PPD-stimulated T cells. IFN-α–producing cells present in day 3 MT skin sections were identified using indirect alkaline phosphatase staining (a, top). The bottom panel shows staining with an isotype control (magnification, 20). Keratinocytes (K) and isolated cells within the perivascular infiltrates (PV; indicated by arrows) expressed IFN-α. (b) The presence of type I IFN in blister fluid collected at different times after MT induction. The data represent the mean ± SEM of type I IFN in blister samples from five different individuals assayed per time point. Type I IFN mediates the suppression of telomerase activity induced by day 3 blister fluid (c). PBMCs were stimulated with PPD in vitro in the presence of autologous day 3 blister fluid (+MT BF) or autologous serum (+serum). Blocking antibody reactive with type I IFN receptor 2 (CD118; +anti-IFN R) or isotype control antibody (+cnt Ab) were added to cultures stimulated in the presence of MT blister fluid. Samples were collected 3 d after stimulation, and samples were adjusted to 500 proliferating Ki67+ T cells per reaction for telomerase analysis (one out of three representative experiments is shown). Direct inhibition of telomerase activity by type I IFN (d). A PPD-specific CD4+ T cell line was restimulated with PPD in the presence of recombinant IFN-α (black bars) or Roferon (white bars), a clinical preparation of IFN-α for 4 d. Samples were adjusted so that telomerase activity was measured in an equivalent number of cycling (Ki67+) T cells (one of three representative experiments is shown). Blister-derived CD4+ T cells can expand in vitro (e). Cells recovered from a day-19 blister were stimulated in vitro with PPD-pulsed irradiated autologous PBMCs as APCs. Arrows indicate each point of restimulation in vitro. The asterisks indicate points when samples were collected to measure telomerase activity. Telomerase activity of CD4+ T cells isolated from day 19 blisters decreases with repeated stimulation in vitro (f). Samples were collected on day 4 after each point of PPD restimulation. Telomerase activity per 500 Ki67+ T cells was determined as before. Stim 4, Stim 8, and Stim 9 indicate the 4th, 8th, and 9th stimulations with PPD and irradiated APCs in vitro.
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Related In: Results  -  Collection

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fig6: Type I IFN in blister fluid inhibits telomerase activity in PPD-stimulated T cells. IFN-α–producing cells present in day 3 MT skin sections were identified using indirect alkaline phosphatase staining (a, top). The bottom panel shows staining with an isotype control (magnification, 20). Keratinocytes (K) and isolated cells within the perivascular infiltrates (PV; indicated by arrows) expressed IFN-α. (b) The presence of type I IFN in blister fluid collected at different times after MT induction. The data represent the mean ± SEM of type I IFN in blister samples from five different individuals assayed per time point. Type I IFN mediates the suppression of telomerase activity induced by day 3 blister fluid (c). PBMCs were stimulated with PPD in vitro in the presence of autologous day 3 blister fluid (+MT BF) or autologous serum (+serum). Blocking antibody reactive with type I IFN receptor 2 (CD118; +anti-IFN R) or isotype control antibody (+cnt Ab) were added to cultures stimulated in the presence of MT blister fluid. Samples were collected 3 d after stimulation, and samples were adjusted to 500 proliferating Ki67+ T cells per reaction for telomerase analysis (one out of three representative experiments is shown). Direct inhibition of telomerase activity by type I IFN (d). A PPD-specific CD4+ T cell line was restimulated with PPD in the presence of recombinant IFN-α (black bars) or Roferon (white bars), a clinical preparation of IFN-α for 4 d. Samples were adjusted so that telomerase activity was measured in an equivalent number of cycling (Ki67+) T cells (one of three representative experiments is shown). Blister-derived CD4+ T cells can expand in vitro (e). Cells recovered from a day-19 blister were stimulated in vitro with PPD-pulsed irradiated autologous PBMCs as APCs. Arrows indicate each point of restimulation in vitro. The asterisks indicate points when samples were collected to measure telomerase activity. Telomerase activity of CD4+ T cells isolated from day 19 blisters decreases with repeated stimulation in vitro (f). Samples were collected on day 4 after each point of PPD restimulation. Telomerase activity per 500 Ki67+ T cells was determined as before. Stim 4, Stim 8, and Stim 9 indicate the 4th, 8th, and 9th stimulations with PPD and irradiated APCs in vitro.
Mentions: Our previous studies indicate that type I IFN (IFN-α and IFN-β) can regulate the rate of differentiation of human T cells (32). Together with reports that IFN-α can inhibit the activity of telomerase in tumor cell lines in vitro (33), this prompted us to determine whether this group of cytokines was responsible for the inhibition of telomerase activity in PPD-specific CD4+ T cells in the skin in situ. By immunohistological analysis, we identified cells that expressed high levels of IFN-α in the perivascular infiltrates of skin sections 3 d after PPD challenge (Fig. 6 a). High uniform levels of expression were also detected in keratinocytes (Fig. 6 a). This was confirmed by measurement of peak concentrations of type I IFN in SB fluid that was obtained at day 3 after PPD injection (Fig. 6 b). This coincided with the time at which telomerase was greatest in CD4+ T cells that were PPD-stimulated in vitro, but absent from cycling CD4+ T cells in the skin (Fig. 5 b).

Bottom Line: Furthermore, significant telomere erosion occurred in specific T cells that respond in the skin, but not in those that are found in the blood from the same individuals.Antibody inhibition studies indicated that type I interferon (IFN), which was identified at high levels in the tissue fluid and by immunohistology, was responsible in part for the telomerase inhibition.Furthermore, the addition of IFN-alpha to PPD-stimulated CD4+ T cells directly inhibited telomerase activity in vitro.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Immunology and Molecular Pathology, Div. of Infection and Immunity, University College London, 46 Cleveland St., London W1T 4JF, England, UK.

ABSTRACT
The extent of human memory T cell proliferation, differentiation, and telomere erosion that occurs after a single episode of immune challenge in vivo is unclear. To investigate this, we injected tuberculin purified protein derivative (PPD) into the skin of immune individuals and isolated responsive T cells from the site of antigenic challenge at different times. PPD-specific CD4+ T cells proliferated and differentiated extensively in the skin during this secondary response. Furthermore, significant telomere erosion occurred in specific T cells that respond in the skin, but not in those that are found in the blood from the same individuals. Tissue fluid obtained from the site of PPD challenge in the skin inhibited the induction of the enzyme telomerase in T cells in vitro. Antibody inhibition studies indicated that type I interferon (IFN), which was identified at high levels in the tissue fluid and by immunohistology, was responsible in part for the telomerase inhibition. Furthermore, the addition of IFN-alpha to PPD-stimulated CD4+ T cells directly inhibited telomerase activity in vitro. Therefore, these results suggest that the rate of telomere erosion in proliferating, antigen-specific CD4+ T cells may be accelerated by type I IFN during a secondary response in vivo.

Show MeSH
Related in: MedlinePlus