The role of nuclear factor-kappaB essential modulator (NEMO) in B cell development and survival - PubMed (original) (raw)

The role of nuclear factor-kappaB essential modulator (NEMO) in B cell development and survival

Shinyop Kim et al. Proc Natl Acad Sci U S A. 2003.

Abstract

The transcription factor nuclear factor-kappaB (NF-kappaB) is essential for immune and inflammatory responses. NF-kappaB essential modulator (NEMO) is a scaffolding component of the IkappaB kinase complex required for NF-kappaB activation in vitro. Because NF-kappaB activation is involved in B cell development and function, we set out to determine whether NEMO is required for these processes. NEMO(-/-) mice die very early during embryogenesis, and fetal livers from NEMO(-/-) embryos can not reconstitute either B or T lymphopoiesis in irradiated host mice. We therefore used NEMO(-/-) embryonic stem cells and the OP9 in vitro differentiation system to demonstrate that NEMO is not required for B cell development but plays an important role in B cell survival.

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Figures

Figure 1

Figure 1

Early stages of the in vitro differentiation of WT and NEMO(−/−) ES cells cocultured with OP9 bone marrow cells. Surface expression levels of CD45, Flk-1, and CD31 on days 5 and 8 of coculture were analyzed by immunostaining and flow cytometric analysis. Numbers in quadrants represent percentage of positively staining cells. For all figures, results shown are representative of two independent experiments involving two cocultures per genotype.

Figure 2

Figure 2

Time course of hematopoietic differentiation in WT and NEMO(−/−) ES/OP9 cocultures. CD45, Mac-1, and Ter-119 surface expression on days 5, 8, 12, and 14 were analyzed as in Fig. 1.

Figure 3

Figure 3

B cell development in WT and NEMO(−/−) ES/OP9 cocultures. Surface expression of B220, CD19, and IgM on days 14, 16, and 19 was analyzed as in Fig. 1.

Figure 4

Figure 4

Impaired survival and mitogenic response of IgM+/B220+ cells from stimulated NEMO(−/−) cocultures. (A) On day 19, WT and NEMO(−/−) ES/OP9 cocultures were either mock-stimulated (−LPS) or treated with LPS (10 μg/ml) for 3 days. Each coculture was analyzed by flow cytometry to determine the relative percentage of IgM+/B220+ cells. (B) Surface expression of the activation marker CD80. On day 19, NEMO(−/−) and WT cocultures were either mock-stimulated or treated with LPS (10 μg/ml) for 3 days. The percentage of CD19+/CD80+ cells was determined by flow cytometry. (C) The number of viable IgM+ B cells in cocultures of A as calculated by immunostaining and flow cytometry.

Figure 5

Figure 5

Impaired survival of isolated NEMO-deficient immature B cells. (A) Surface expression of the activation marker CD80. On day 19, CD19+/IgM+ cells were isolated from NEMO(−/−) and WT cocultures by cell sorting. Isolated CD19+/IgM+ B cells were either mock-stimulated or treated with LPS (10 μg/ml) for 2 days. The percentage of IgM+/CD80+ cells was determined by immunostaining and flow cytometry. (B) Surface expression of IgM. On day 19, CD19+/IgM− cells were isolated from NEMO(−/−) and WT cocultures by cell sorting. Isolated CD19+/IgM− B cells were stimulated as in A. The percentage of IgM+/CD19+ cells was determined by immunostaining and flow cytometry.

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References

    1. Baeuerle P A, Baltimore D. Cell. 1996;87:13–20. - PubMed
    1. Barnes P J, Karin M. N Engl J Med. 1997;336:1066–1071. - PubMed
    1. Sen R, Baltimore D. Cell. 1986;46:705–716. - PubMed
    1. Sen R, Baltimore D. Cell. 1986;47:921–928. - PubMed
    1. Brown K, Gerstberger S, Carlson L, Franzoso G, Siebenlist U. Science. 1995;267:1485–1488. - PubMed

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