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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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Low telomerase activity in CD4+ T cells in the skin after PPD stimulation in vivo. Telomerase activity in T cells isolated from SBs after PPD injection in two different individuals on days 3 and 7 after PPD injection (in vivo) and also from blood CD4+ T cells from one individual at different times after PPD stimulation in vitro (a). Telomerase activity (total product generated) per 500 Ki67+ proliferating T cells isolated from MTs (b, white bars) and PBMCs stimulated in vitro with PPD (black bars). The mean ± SEM of three experiments is shown. Skin CD4+ T cells can up-regulate telomerase activity when stimulated with PPD in vitro (c). T cells were isolated from day 3 MTs, and telomerase activity was determined in T cells immediately upon isolation (Blister), whereas the remaining cells were stimulated with PPD in vitro for 3 d (In vitro re-stim). Data are representative of four experiments performed. The negative control contains the PCR mix without cell extract, and the positive control contains an extract of a telomerase-positive tumor cell line. TSR8 denotes the internal quantitative control. Day 3 blister fluid suppresses telomerase activity (d). PBMCs were stimulated with PPD in vitro in the presence of autologous serum as a control or autologous blister fluid that was obtained from day 3 MT. Telomerase activity per 500 Ki67+ T cells was measured on day 3. The data are representative of three experiments performed.
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fig5: Low telomerase activity in CD4+ T cells in the skin after PPD stimulation in vivo. Telomerase activity in T cells isolated from SBs after PPD injection in two different individuals on days 3 and 7 after PPD injection (in vivo) and also from blood CD4+ T cells from one individual at different times after PPD stimulation in vitro (a). Telomerase activity (total product generated) per 500 Ki67+ proliferating T cells isolated from MTs (b, white bars) and PBMCs stimulated in vitro with PPD (black bars). The mean ± SEM of three experiments is shown. Skin CD4+ T cells can up-regulate telomerase activity when stimulated with PPD in vitro (c). T cells were isolated from day 3 MTs, and telomerase activity was determined in T cells immediately upon isolation (Blister), whereas the remaining cells were stimulated with PPD in vitro for 3 d (In vitro re-stim). Data are representative of four experiments performed. The negative control contains the PCR mix without cell extract, and the positive control contains an extract of a telomerase-positive tumor cell line. TSR8 denotes the internal quantitative control. Day 3 blister fluid suppresses telomerase activity (d). PBMCs were stimulated with PPD in vitro in the presence of autologous serum as a control or autologous blister fluid that was obtained from day 3 MT. Telomerase activity per 500 Ki67+ T cells was measured on day 3. The data are representative of three experiments performed.

Mentions: We investigated whether telomere erosion in CD4+ T cells after PPD stimulation in vivo resulted from lack of telomerase induction in these cells. At days 3 and 7 after PPD challenge, there was extensive CD4+ T cell cycling as 7.7 ± 2.0% and 17.8 ± 6.2% of these cells, respectively, were Ki67+. The extent of cell cycling was greater than that observed when CD4+ T cells from blood were stimulated by PPD in vitro for 3 and 7 d, respectively (2.4 ± 0.4% and 9.7 ± 4.0% Ki67+). Because telomerase is only up-regulated in the activated proliferating population of T cells (29, 30), we standardized telomerase assays to include an equivalent number of Ki67+ T cells in each sample. For the first time, this allowed a direct comparison of the activity of this enzyme in cycling CD4+ T cells from both blood and blister at different times after stimulation (30, 31). Telomerase activity was minimal in cycling CD4+ T cells that were isolated from the skin compared with CD4+ T cells from blood that were stimulated by PPD in vitro (Fig. 5, a and b). This indicated that telomere erosion in PPD-specific CD4+ T cells was linked to the lack of telomerase activity in these cells in vivo.


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)

Low telomerase activity in CD4+ T cells in the skin after PPD stimulation in vivo. Telomerase activity in T cells isolated from SBs after PPD injection in two different individuals on days 3 and 7 after PPD injection (in vivo) and also from blood CD4+ T cells from one individual at different times after PPD stimulation in vitro (a). Telomerase activity (total product generated) per 500 Ki67+ proliferating T cells isolated from MTs (b, white bars) and PBMCs stimulated in vitro with PPD (black bars). The mean ± SEM of three experiments is shown. Skin CD4+ T cells can up-regulate telomerase activity when stimulated with PPD in vitro (c). T cells were isolated from day 3 MTs, and telomerase activity was determined in T cells immediately upon isolation (Blister), whereas the remaining cells were stimulated with PPD in vitro for 3 d (In vitro re-stim). Data are representative of four experiments performed. The negative control contains the PCR mix without cell extract, and the positive control contains an extract of a telomerase-positive tumor cell line. TSR8 denotes the internal quantitative control. Day 3 blister fluid suppresses telomerase activity (d). PBMCs were stimulated with PPD in vitro in the presence of autologous serum as a control or autologous blister fluid that was obtained from day 3 MT. Telomerase activity per 500 Ki67+ T cells was measured on day 3. The data are representative of three experiments performed.
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Related In: Results  -  Collection

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fig5: Low telomerase activity in CD4+ T cells in the skin after PPD stimulation in vivo. Telomerase activity in T cells isolated from SBs after PPD injection in two different individuals on days 3 and 7 after PPD injection (in vivo) and also from blood CD4+ T cells from one individual at different times after PPD stimulation in vitro (a). Telomerase activity (total product generated) per 500 Ki67+ proliferating T cells isolated from MTs (b, white bars) and PBMCs stimulated in vitro with PPD (black bars). The mean ± SEM of three experiments is shown. Skin CD4+ T cells can up-regulate telomerase activity when stimulated with PPD in vitro (c). T cells were isolated from day 3 MTs, and telomerase activity was determined in T cells immediately upon isolation (Blister), whereas the remaining cells were stimulated with PPD in vitro for 3 d (In vitro re-stim). Data are representative of four experiments performed. The negative control contains the PCR mix without cell extract, and the positive control contains an extract of a telomerase-positive tumor cell line. TSR8 denotes the internal quantitative control. Day 3 blister fluid suppresses telomerase activity (d). PBMCs were stimulated with PPD in vitro in the presence of autologous serum as a control or autologous blister fluid that was obtained from day 3 MT. Telomerase activity per 500 Ki67+ T cells was measured on day 3. The data are representative of three experiments performed.
Mentions: We investigated whether telomere erosion in CD4+ T cells after PPD stimulation in vivo resulted from lack of telomerase induction in these cells. At days 3 and 7 after PPD challenge, there was extensive CD4+ T cell cycling as 7.7 ± 2.0% and 17.8 ± 6.2% of these cells, respectively, were Ki67+. The extent of cell cycling was greater than that observed when CD4+ T cells from blood were stimulated by PPD in vitro for 3 and 7 d, respectively (2.4 ± 0.4% and 9.7 ± 4.0% Ki67+). Because telomerase is only up-regulated in the activated proliferating population of T cells (29, 30), we standardized telomerase assays to include an equivalent number of Ki67+ T cells in each sample. For the first time, this allowed a direct comparison of the activity of this enzyme in cycling CD4+ T cells from both blood and blister at different times after stimulation (30, 31). Telomerase activity was minimal in cycling CD4+ T cells that were isolated from the skin compared with CD4+ T cells from blood that were stimulated by PPD in vitro (Fig. 5, a and b). This indicated that telomere erosion in PPD-specific CD4+ T cells was linked to the lack of telomerase activity in these cells in vivo.

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