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Rat spleen dendritic cells express natural killer cell receptor protein 1 (NKR-P1) and have cytotoxic activity to select targets via a Ca2+-dependent mechanism.

Josien R, Heslan M, Soulillou JP, Cuturi MC - J. Exp. Med. (1997)

Bottom Line: We show that both spleen and thymus DC express the natural killer cell receptor protein 1 (NKR-P1) as a disulfide linked homodimer of 60 kD.Freshly isolated DC express a low level of NKR-P1, which is strongly upregulated after overnight culture.Spleen, but not thymus DC, were able to kill the NK-sensitive YAC-1 cell line in vitro, and since this killing was Ca2+ dependent, a Fas ligand-Fas interaction was probably not involved.

View Article: PubMed Central - PubMed

Affiliation: Institut National de la Sante et de la Recherche Medicale U437, Institut de Transplantation et de Recherche en Transplantation, Immeuble Jean Monnet, Nantes, France. josienr@rockvax.rockefeller.edu

ABSTRACT
Dendritic cells (DC) are a subset of leukocytes whose major function is antigen presentation. We investigated the phenotype and function of enriched (95-98.5%) rat DC. We show that both spleen and thymus DC express the natural killer cell receptor protein 1 (NKR-P1) as a disulfide linked homodimer of 60 kD. Freshly isolated DC express a low level of NKR-P1, which is strongly upregulated after overnight culture. Spleen, but not thymus DC, were able to kill the NK-sensitive YAC-1 cell line in vitro, and since this killing was Ca2+ dependent, a Fas ligand-Fas interaction was probably not involved. Besides their potent antigen-presenting function, DC can thus be cytotoxic for some tumor targets.

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Related in: MedlinePlus

Spleen DC kill the  YAC-1 target cells using a Ca2+-dependent mechanism. 51Cr-labeled YAC-1 target cells (2 ×  103) were incubated with purified spleen DC (95–98.5%) in  triplicate for 6 h at 37°C in the  presence (open square) or absence (closed square) of 2 mM  EGTA and 2 mM MgCl2. 51Cr  release was then assessed in the  supernatant. Results of one experiment representative of eight  are shown.
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Figure 5: Spleen DC kill the YAC-1 target cells using a Ca2+-dependent mechanism. 51Cr-labeled YAC-1 target cells (2 × 103) were incubated with purified spleen DC (95–98.5%) in triplicate for 6 h at 37°C in the presence (open square) or absence (closed square) of 2 mM EGTA and 2 mM MgCl2. 51Cr release was then assessed in the supernatant. Results of one experiment representative of eight are shown.

Mentions: Two arguments indicated that rat spleen DC probably do not kill YAC-1 target cells through the FasL/Fas pathway. First, we could not detect membrane FasL expression on spleen or thymus DC, as determined by FACS® using a Fas-Ig chimeric molecule (data not shown). Second, the in vitro cytolytic activity of splenic DC was abrogated upon addition of EGTA (2 mM) to the culture, together with 2 mM MgCl2 to ensure excess Mg2+ concentrations (Fig. 5). The viability of spleen DC during the assay was not modified by the presence of EGTA (data not shown). These results indicated that the mechanism of spleen DC killing was Ca2+ dependent, and thus unlikely to be related to FasL/Fas interaction (24). This contrasts with the recent report of Süss and Shortman showing that a subset of spleen DC in mice express a functional Fas ligand molecule (15). It is unlikely that the complete inhibition of splenic DC killing by the chelating agent EGTA was related to the inhibition of the ligand binding capacity of NKR-P1 molecule because EGTA concentration used in our assays (2 mM) is not sufficient to inhibit its carbohydrate-binding capacity (25). The dependence on Ca2+ of rat splenic DC killing rather suggests a mechanism of granule exocytosis as described for CTL and NK cells. Whether rat spleen DC contain perforin-containing granules is under investigation.


Rat spleen dendritic cells express natural killer cell receptor protein 1 (NKR-P1) and have cytotoxic activity to select targets via a Ca2+-dependent mechanism.

Josien R, Heslan M, Soulillou JP, Cuturi MC - J. Exp. Med. (1997)

Spleen DC kill the  YAC-1 target cells using a Ca2+-dependent mechanism. 51Cr-labeled YAC-1 target cells (2 ×  103) were incubated with purified spleen DC (95–98.5%) in  triplicate for 6 h at 37°C in the  presence (open square) or absence (closed square) of 2 mM  EGTA and 2 mM MgCl2. 51Cr  release was then assessed in the  supernatant. Results of one experiment representative of eight  are shown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2198993&req=5

Figure 5: Spleen DC kill the YAC-1 target cells using a Ca2+-dependent mechanism. 51Cr-labeled YAC-1 target cells (2 × 103) were incubated with purified spleen DC (95–98.5%) in triplicate for 6 h at 37°C in the presence (open square) or absence (closed square) of 2 mM EGTA and 2 mM MgCl2. 51Cr release was then assessed in the supernatant. Results of one experiment representative of eight are shown.
Mentions: Two arguments indicated that rat spleen DC probably do not kill YAC-1 target cells through the FasL/Fas pathway. First, we could not detect membrane FasL expression on spleen or thymus DC, as determined by FACS® using a Fas-Ig chimeric molecule (data not shown). Second, the in vitro cytolytic activity of splenic DC was abrogated upon addition of EGTA (2 mM) to the culture, together with 2 mM MgCl2 to ensure excess Mg2+ concentrations (Fig. 5). The viability of spleen DC during the assay was not modified by the presence of EGTA (data not shown). These results indicated that the mechanism of spleen DC killing was Ca2+ dependent, and thus unlikely to be related to FasL/Fas interaction (24). This contrasts with the recent report of Süss and Shortman showing that a subset of spleen DC in mice express a functional Fas ligand molecule (15). It is unlikely that the complete inhibition of splenic DC killing by the chelating agent EGTA was related to the inhibition of the ligand binding capacity of NKR-P1 molecule because EGTA concentration used in our assays (2 mM) is not sufficient to inhibit its carbohydrate-binding capacity (25). The dependence on Ca2+ of rat splenic DC killing rather suggests a mechanism of granule exocytosis as described for CTL and NK cells. Whether rat spleen DC contain perforin-containing granules is under investigation.

Bottom Line: We show that both spleen and thymus DC express the natural killer cell receptor protein 1 (NKR-P1) as a disulfide linked homodimer of 60 kD.Freshly isolated DC express a low level of NKR-P1, which is strongly upregulated after overnight culture.Spleen, but not thymus DC, were able to kill the NK-sensitive YAC-1 cell line in vitro, and since this killing was Ca2+ dependent, a Fas ligand-Fas interaction was probably not involved.

View Article: PubMed Central - PubMed

Affiliation: Institut National de la Sante et de la Recherche Medicale U437, Institut de Transplantation et de Recherche en Transplantation, Immeuble Jean Monnet, Nantes, France. josienr@rockvax.rockefeller.edu

ABSTRACT
Dendritic cells (DC) are a subset of leukocytes whose major function is antigen presentation. We investigated the phenotype and function of enriched (95-98.5%) rat DC. We show that both spleen and thymus DC express the natural killer cell receptor protein 1 (NKR-P1) as a disulfide linked homodimer of 60 kD. Freshly isolated DC express a low level of NKR-P1, which is strongly upregulated after overnight culture. Spleen, but not thymus DC, were able to kill the NK-sensitive YAC-1 cell line in vitro, and since this killing was Ca2+ dependent, a Fas ligand-Fas interaction was probably not involved. Besides their potent antigen-presenting function, DC can thus be cytotoxic for some tumor targets.

Show MeSH
Related in: MedlinePlus