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Generation of a novel regulatory NK cell subset from peripheral blood CD34+ progenitors promoted by membrane-bound IL-15.

Giuliani M, Giron-Michel J, Negrini S, Vacca P, Durali D, Caignard A, Le Bousse-Kerdiles C, Chouaib S, Devocelle A, Bahri R, Durrbach A, Taoufik Y, Ferrini S, Croce M, Mingari MC, Moretta L, Azzarone B - PLoS ONE (2008)

Bottom Line: Through these factors, NK-ireg cells act as effectors of the down-regulation of the immune response: reconverting mature myeloid DC (mDC) into immature/tolerogenic DC, blocking cytolytic functions on conventional NK cells and inducing HLA-G membrane expression on PB-derived monocytes.In conclusion, NK-ireg cells represent a novel truly differentiated non-cytolytic NK subset with a self-sustainable phenotype (CD56(+) CD16(+) NKp30(+) NKp44(+) NKp46(+) CD94(+) CD69(+) CCR7(+)) generated from specific pSTAT6(+) GATA3(+) precursors.In addition, NK-ireg cells can be easily derived from peripheral blood of the patients and could constitute an autologous biotherapic tool to be used combined or in alternative to other immuno-regulatory cells.

View Article: PubMed Central - PubMed

Affiliation: INSERM, UMR 542, Université de Paris XI, Hôpital Paul Brousse, Villejuif, France.

ABSTRACT

Background: NK cells have been long time considered as cytotoxic lymphocytes competent in killing virus-infected cells and tumors. However, NK cells may also play essential immuno-regulatory functions. In this context, the real existence of a defined NK subset with negative regulatory properties has been hypothesized but never clearly demonstrated.

Methodology/principal findings: Herein, we show the in vitro generation from human peripheral blood haematopoietic progenitors (PB-HP), of a novel subset of non-cytolytic NK cells displaying a mature phenotype and remarkable immuno-regulatory functions (NK-ireg). The main functional hallmark of these NK-ireg cells is represented by the surface expression/release of HLA-G, a major immunosuppressive molecule. In addition, NK-ireg cells secrete two powerful immuno-regulatory factors: IL-10 and IL-21. Through these factors, NK-ireg cells act as effectors of the down-regulation of the immune response: reconverting mature myeloid DC (mDC) into immature/tolerogenic DC, blocking cytolytic functions on conventional NK cells and inducing HLA-G membrane expression on PB-derived monocytes. The generation of "NK-ireg" cells is obtained, by default, in culture conditions favouring cell-to-cell contacts, and it is strictly dependent on reciprocal trans-presentation of membrane-bound IL-15 forms constitutively and selectively expressed by human CD34(+) PB-HP. Finally, a small subset of NKp46(+) HLA-G(+) IL-10(+) is detected within freshly isolated decidual NK cells, suggesting that these cells could represent an in vivo counterpart of the NK-ireg cells.

Conclusions/significance: In conclusion, NK-ireg cells represent a novel truly differentiated non-cytolytic NK subset with a self-sustainable phenotype (CD56(+) CD16(+) NKp30(+) NKp44(+) NKp46(+) CD94(+) CD69(+) CCR7(+)) generated from specific pSTAT6(+) GATA3(+) precursors. NK-ireg cells could be employed to develop new immuno-suppressive strategies in autoimmune diseases, transplant rejection or graft versus host diseases. In addition, NK-ireg cells can be easily derived from peripheral blood of the patients and could constitute an autologous biotherapic tool to be used combined or in alternative to other immuno-regulatory cells.

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Autonomous NK commitment of PB-HP.Commitment into the NK pathway of PB-HP was evaluated by flow cytometry at different times in culture, analyzing CD34, CD38 and the NK markers CD56, CD16, CD161, CD94, NKp30, NKp44, NKp46, NKG2D, CD69 and CCR7. Shadowed peaks represent staining with isotype control Abs. Continuous lines represent staining with the indicated Abs. The frequency of PB-HP represents about 0.1% of mononuclear cells of the peripheral blood. The data are representative of more than 30 separate experiments.
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pone-0002241-g002: Autonomous NK commitment of PB-HP.Commitment into the NK pathway of PB-HP was evaluated by flow cytometry at different times in culture, analyzing CD34, CD38 and the NK markers CD56, CD16, CD161, CD94, NKp30, NKp44, NKp46, NKG2D, CD69 and CCR7. Shadowed peaks represent staining with isotype control Abs. Continuous lines represent staining with the indicated Abs. The frequency of PB-HP represents about 0.1% of mononuclear cells of the peripheral blood. The data are representative of more than 30 separate experiments.

Mentions: We next investigated whether the human CD34+ PB-HP expressing mb-IL-15 could trans-present IL-15 each other mimicking the in vivo effect of BM myeloid accessory cells on bystander BM-HP and trigger their commitment towards the NK cell lineage in the absence of exogenous lymphokines. Freshly isolated PB-HP expresses the CD34+ CD38+ CD56dull CD16− CD161− CD14− surface phenotype (Fig. 2, D0). When these cells are maintained at high cell density that favors cell to cell contact (5×105 cells/ml), there was a progressive expansion of uni-lineage homogeneous committed progenitors that, growing in suspension, progressively acquired the phenotype of mature NK cells. On day 7, these progenitors acquired markers typical of immature NK cells. However, they still maintained low expression of CD34 (CD34low CD56+ CD16low CD161+ CD94−/+) (Fig. 2, D7). At this stage, we frequently observed that a small subset of CD1a adherent cells stems from the cells growing in suspension. This small subset represents less then 3% of the total cell population, but could somehow influence the maturation process of these PB-NKP [16].


Generation of a novel regulatory NK cell subset from peripheral blood CD34+ progenitors promoted by membrane-bound IL-15.

Giuliani M, Giron-Michel J, Negrini S, Vacca P, Durali D, Caignard A, Le Bousse-Kerdiles C, Chouaib S, Devocelle A, Bahri R, Durrbach A, Taoufik Y, Ferrini S, Croce M, Mingari MC, Moretta L, Azzarone B - PLoS ONE (2008)

Autonomous NK commitment of PB-HP.Commitment into the NK pathway of PB-HP was evaluated by flow cytometry at different times in culture, analyzing CD34, CD38 and the NK markers CD56, CD16, CD161, CD94, NKp30, NKp44, NKp46, NKG2D, CD69 and CCR7. Shadowed peaks represent staining with isotype control Abs. Continuous lines represent staining with the indicated Abs. The frequency of PB-HP represents about 0.1% of mononuclear cells of the peripheral blood. The data are representative of more than 30 separate experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2376096&req=5

pone-0002241-g002: Autonomous NK commitment of PB-HP.Commitment into the NK pathway of PB-HP was evaluated by flow cytometry at different times in culture, analyzing CD34, CD38 and the NK markers CD56, CD16, CD161, CD94, NKp30, NKp44, NKp46, NKG2D, CD69 and CCR7. Shadowed peaks represent staining with isotype control Abs. Continuous lines represent staining with the indicated Abs. The frequency of PB-HP represents about 0.1% of mononuclear cells of the peripheral blood. The data are representative of more than 30 separate experiments.
Mentions: We next investigated whether the human CD34+ PB-HP expressing mb-IL-15 could trans-present IL-15 each other mimicking the in vivo effect of BM myeloid accessory cells on bystander BM-HP and trigger their commitment towards the NK cell lineage in the absence of exogenous lymphokines. Freshly isolated PB-HP expresses the CD34+ CD38+ CD56dull CD16− CD161− CD14− surface phenotype (Fig. 2, D0). When these cells are maintained at high cell density that favors cell to cell contact (5×105 cells/ml), there was a progressive expansion of uni-lineage homogeneous committed progenitors that, growing in suspension, progressively acquired the phenotype of mature NK cells. On day 7, these progenitors acquired markers typical of immature NK cells. However, they still maintained low expression of CD34 (CD34low CD56+ CD16low CD161+ CD94−/+) (Fig. 2, D7). At this stage, we frequently observed that a small subset of CD1a adherent cells stems from the cells growing in suspension. This small subset represents less then 3% of the total cell population, but could somehow influence the maturation process of these PB-NKP [16].

Bottom Line: Through these factors, NK-ireg cells act as effectors of the down-regulation of the immune response: reconverting mature myeloid DC (mDC) into immature/tolerogenic DC, blocking cytolytic functions on conventional NK cells and inducing HLA-G membrane expression on PB-derived monocytes.In conclusion, NK-ireg cells represent a novel truly differentiated non-cytolytic NK subset with a self-sustainable phenotype (CD56(+) CD16(+) NKp30(+) NKp44(+) NKp46(+) CD94(+) CD69(+) CCR7(+)) generated from specific pSTAT6(+) GATA3(+) precursors.In addition, NK-ireg cells can be easily derived from peripheral blood of the patients and could constitute an autologous biotherapic tool to be used combined or in alternative to other immuno-regulatory cells.

View Article: PubMed Central - PubMed

Affiliation: INSERM, UMR 542, Université de Paris XI, Hôpital Paul Brousse, Villejuif, France.

ABSTRACT

Background: NK cells have been long time considered as cytotoxic lymphocytes competent in killing virus-infected cells and tumors. However, NK cells may also play essential immuno-regulatory functions. In this context, the real existence of a defined NK subset with negative regulatory properties has been hypothesized but never clearly demonstrated.

Methodology/principal findings: Herein, we show the in vitro generation from human peripheral blood haematopoietic progenitors (PB-HP), of a novel subset of non-cytolytic NK cells displaying a mature phenotype and remarkable immuno-regulatory functions (NK-ireg). The main functional hallmark of these NK-ireg cells is represented by the surface expression/release of HLA-G, a major immunosuppressive molecule. In addition, NK-ireg cells secrete two powerful immuno-regulatory factors: IL-10 and IL-21. Through these factors, NK-ireg cells act as effectors of the down-regulation of the immune response: reconverting mature myeloid DC (mDC) into immature/tolerogenic DC, blocking cytolytic functions on conventional NK cells and inducing HLA-G membrane expression on PB-derived monocytes. The generation of "NK-ireg" cells is obtained, by default, in culture conditions favouring cell-to-cell contacts, and it is strictly dependent on reciprocal trans-presentation of membrane-bound IL-15 forms constitutively and selectively expressed by human CD34(+) PB-HP. Finally, a small subset of NKp46(+) HLA-G(+) IL-10(+) is detected within freshly isolated decidual NK cells, suggesting that these cells could represent an in vivo counterpart of the NK-ireg cells.

Conclusions/significance: In conclusion, NK-ireg cells represent a novel truly differentiated non-cytolytic NK subset with a self-sustainable phenotype (CD56(+) CD16(+) NKp30(+) NKp44(+) NKp46(+) CD94(+) CD69(+) CCR7(+)) generated from specific pSTAT6(+) GATA3(+) precursors. NK-ireg cells could be employed to develop new immuno-suppressive strategies in autoimmune diseases, transplant rejection or graft versus host diseases. In addition, NK-ireg cells can be easily derived from peripheral blood of the patients and could constitute an autologous biotherapic tool to be used combined or in alternative to other immuno-regulatory cells.

Show MeSH
Related in: MedlinePlus