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Membrane Fas ligand kills human peripheral blood T lymphocytes, and soluble Fas ligand blocks the killing.

Suda T, Hashimoto H, Tanaka M, Ochi T, Nagata S - J. Exp. Med. (1997)

Bottom Line: In contrast, soluble FasL kills only the latter.Soluble FasL inhibited the killing of fresh PBT by membrane FasL.These results indicate that the shedding of FasL from the membrane is a mechanism for downregulating at least part of its killing activity.

View Article: PubMed Central - PubMed

Affiliation: Osaka Bioscience Institute, Department of Molecular Biology, Suita, Osaka 565, Japan. sudat@obi.or.jp

ABSTRACT
It has been believed that the Fas expressed on human peripheral blood T cells (PBT) is nonfunctional, because these cells are insensitive to agonistic anti-Fas/Apo-1 mAbs that efficiently kill in vitro-activated T cells and many Fas-expressing cell lines. Here, we demonstrate that membrane-bound Fas ligand (FasL) kills both fresh and in vitro-activated PBT, indicating that the Fas expressed on fresh PBT is functional. In contrast, soluble FasL kills only the latter. Naive T cells in umbilical cord blood do not express Fas, but can be induced to express Fas by IFN-gamma or by a combination of IL-2 and anti-CD28 mAb, after which they acquire sensitivity to membrane but not to soluble FasL. Soluble FasL inhibited the killing of fresh PBT by membrane FasL. These results indicate that the shedding of FasL from the membrane is a mechanism for downregulating at least part of its killing activity.

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Function of Fas on CBT induced by IFN-γ or a combination of IL-2 and immobilized anti-CD28 mAb. (a–h) Cord blood lymphocytes were cultured for 20 h with medium only (dotted line in each  panel), IFN–γ (10 ng/ml, a and e), IL-2 (10 ng/ml, b and f), immobilized  anti-CD28 mAb (coated on plate at 10 μg/ml, c and g), or IL-2 plus immobilized anti-CD28 mAb (d and h), respectively. Cells were then stained  with FITC–anti-Fas mAb, PE-conjugated mAb against either CD4 or  CD8, and PI. FITC–anti-Fas mAb staining profiles of viable (PI−) CD4+  (a–d) or CD8+ cells (e–h) are shown. (i and j) CBT treated with IFN-γ or  with IL-2 plus anti-CD28 mAb as described above were cultured with  CH11, WX1, AL-1, or 1A12.
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Figure 3: Function of Fas on CBT induced by IFN-γ or a combination of IL-2 and immobilized anti-CD28 mAb. (a–h) Cord blood lymphocytes were cultured for 20 h with medium only (dotted line in each panel), IFN–γ (10 ng/ml, a and e), IL-2 (10 ng/ml, b and f), immobilized anti-CD28 mAb (coated on plate at 10 μg/ml, c and g), or IL-2 plus immobilized anti-CD28 mAb (d and h), respectively. Cells were then stained with FITC–anti-Fas mAb, PE-conjugated mAb against either CD4 or CD8, and PI. FITC–anti-Fas mAb staining profiles of viable (PI−) CD4+ (a–d) or CD8+ cells (e–h) are shown. (i and j) CBT treated with IFN-γ or with IL-2 plus anti-CD28 mAb as described above were cultured with CH11, WX1, AL-1, or 1A12.

Mentions: Essentially all cord blood T cells (CBT) are CD45RO− naive T cells and do not express Fas at detectable levels. Accordingly, they are not killed by any form of FasL or anti-Fas mAb (data not shown). Previously, it was reported that CBT cultured with phytohemagglutinin for 5 d express Fas (20). Here, we found that IFN-γ induces Fas expression on both CD4+ and CD8+ CBT within 20 h (Fig. 3, a and e). IL-2 alone induced FasL weakly, while other stimuli including IL-1, IL-4, IL-6, TNF-α, and immobilized anti-CD28 mAb did not, when added separately (Fig. 3, b, c, f, and g, and data not shown). However, a combination of IL-2 and anti-CD28 mAb showed Fas-inducing activity comparable to IFN-γ (Fig. 3, d and h). The levels of Fas expression induced by IFN-γ plus anti-CD28 mAb were not much different from those induced by IFN-γ alone (data not shown).


Membrane Fas ligand kills human peripheral blood T lymphocytes, and soluble Fas ligand blocks the killing.

Suda T, Hashimoto H, Tanaka M, Ochi T, Nagata S - J. Exp. Med. (1997)

Function of Fas on CBT induced by IFN-γ or a combination of IL-2 and immobilized anti-CD28 mAb. (a–h) Cord blood lymphocytes were cultured for 20 h with medium only (dotted line in each  panel), IFN–γ (10 ng/ml, a and e), IL-2 (10 ng/ml, b and f), immobilized  anti-CD28 mAb (coated on plate at 10 μg/ml, c and g), or IL-2 plus immobilized anti-CD28 mAb (d and h), respectively. Cells were then stained  with FITC–anti-Fas mAb, PE-conjugated mAb against either CD4 or  CD8, and PI. FITC–anti-Fas mAb staining profiles of viable (PI−) CD4+  (a–d) or CD8+ cells (e–h) are shown. (i and j) CBT treated with IFN-γ or  with IL-2 plus anti-CD28 mAb as described above were cultured with  CH11, WX1, AL-1, or 1A12.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2199173&req=5

Figure 3: Function of Fas on CBT induced by IFN-γ or a combination of IL-2 and immobilized anti-CD28 mAb. (a–h) Cord blood lymphocytes were cultured for 20 h with medium only (dotted line in each panel), IFN–γ (10 ng/ml, a and e), IL-2 (10 ng/ml, b and f), immobilized anti-CD28 mAb (coated on plate at 10 μg/ml, c and g), or IL-2 plus immobilized anti-CD28 mAb (d and h), respectively. Cells were then stained with FITC–anti-Fas mAb, PE-conjugated mAb against either CD4 or CD8, and PI. FITC–anti-Fas mAb staining profiles of viable (PI−) CD4+ (a–d) or CD8+ cells (e–h) are shown. (i and j) CBT treated with IFN-γ or with IL-2 plus anti-CD28 mAb as described above were cultured with CH11, WX1, AL-1, or 1A12.
Mentions: Essentially all cord blood T cells (CBT) are CD45RO− naive T cells and do not express Fas at detectable levels. Accordingly, they are not killed by any form of FasL or anti-Fas mAb (data not shown). Previously, it was reported that CBT cultured with phytohemagglutinin for 5 d express Fas (20). Here, we found that IFN-γ induces Fas expression on both CD4+ and CD8+ CBT within 20 h (Fig. 3, a and e). IL-2 alone induced FasL weakly, while other stimuli including IL-1, IL-4, IL-6, TNF-α, and immobilized anti-CD28 mAb did not, when added separately (Fig. 3, b, c, f, and g, and data not shown). However, a combination of IL-2 and anti-CD28 mAb showed Fas-inducing activity comparable to IFN-γ (Fig. 3, d and h). The levels of Fas expression induced by IFN-γ plus anti-CD28 mAb were not much different from those induced by IFN-γ alone (data not shown).

Bottom Line: In contrast, soluble FasL kills only the latter.Soluble FasL inhibited the killing of fresh PBT by membrane FasL.These results indicate that the shedding of FasL from the membrane is a mechanism for downregulating at least part of its killing activity.

View Article: PubMed Central - PubMed

Affiliation: Osaka Bioscience Institute, Department of Molecular Biology, Suita, Osaka 565, Japan. sudat@obi.or.jp

ABSTRACT
It has been believed that the Fas expressed on human peripheral blood T cells (PBT) is nonfunctional, because these cells are insensitive to agonistic anti-Fas/Apo-1 mAbs that efficiently kill in vitro-activated T cells and many Fas-expressing cell lines. Here, we demonstrate that membrane-bound Fas ligand (FasL) kills both fresh and in vitro-activated PBT, indicating that the Fas expressed on fresh PBT is functional. In contrast, soluble FasL kills only the latter. Naive T cells in umbilical cord blood do not express Fas, but can be induced to express Fas by IFN-gamma or by a combination of IL-2 and anti-CD28 mAb, after which they acquire sensitivity to membrane but not to soluble FasL. Soluble FasL inhibited the killing of fresh PBT by membrane FasL. These results indicate that the shedding of FasL from the membrane is a mechanism for downregulating at least part of its killing activity.

Show MeSH
Related in: MedlinePlus