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Human BLyS facilitates engraftment of human PBL derived B cells in immunodeficient mice.

Schmidt MR, Appel MC, Giassi LJ, Greiner DL, Shultz LD, Woodland RT - PLoS ONE (2008)

Bottom Line: Although human B cells bound both human and murine BLyS, nuclear accumulation of NF-kappaB p52, an indication of the induction of a protective anti-apoptotic response, following stimulation with human BLyS was more robust than that induced with murine BLyS suggesting a fundamental disparity in BLyS receptor signaling.Human BLyS treated recipients had on average 40-fold higher levels of serum Ig than controls and mounted a de novo antibody response to the thymus-independent antigens in pneumovax vaccine.The data indicate that production of fully immunologically competent humanized mice from PBL can be markedly facilitated by providing human BLyS.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America. Madelyn.Schmidt@umassmed.edu

ABSTRACT
The production of fully immunologically competent humanized mice engrafted with peripheral lymphocyte populations provides a model for in vivo testing of new vaccines, the durability of immunological memory and cancer therapies. This approach is limited, however, by the failure to efficiently engraft human B lymphocytes in immunodeficient mice. We hypothesized that this deficiency was due to the failure of the murine microenvironment to support human B cell survival. We report that while the human B lymphocyte survival factor, B lymphocyte stimulator (BLyS/BAFF) enhances the survival of human B cells ex vivo, murine BLyS has no such protective effect. Although human B cells bound both human and murine BLyS, nuclear accumulation of NF-kappaB p52, an indication of the induction of a protective anti-apoptotic response, following stimulation with human BLyS was more robust than that induced with murine BLyS suggesting a fundamental disparity in BLyS receptor signaling. Efficient engraftment of both human B and T lymphocytes in NOD rag1(-/-) Prf1(-/-) immunodeficient mice treated with recombinant human BLyS is observed after adoptive transfer of human PBL relative to PBS treated controls. Human BLyS treated recipients had on average 40-fold higher levels of serum Ig than controls and mounted a de novo antibody response to the thymus-independent antigens in pneumovax vaccine. The data indicate that production of fully immunologically competent humanized mice from PBL can be markedly facilitated by providing human BLyS.

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FACS analysis of B cell cultures.B cell populations were FACS analyzed for surface markers associated with resting B cells (CD45, CD19), memory cells (CD27), plasma cells (CD38) and kappa and lambda light chains on the day of isolation and after 4 days of culture either unstimulated or stimulated with human or murine BLyS. All samples were initially gated for lymphocytes by forward and side scatter. Data representative of 4 experiments.
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pone-0003192-g002: FACS analysis of B cell cultures.B cell populations were FACS analyzed for surface markers associated with resting B cells (CD45, CD19), memory cells (CD27), plasma cells (CD38) and kappa and lambda light chains on the day of isolation and after 4 days of culture either unstimulated or stimulated with human or murine BLyS. All samples were initially gated for lymphocytes by forward and side scatter. Data representative of 4 experiments.

Mentions: To determine if huBLyS conferred a selective survival advantage to a particular subpopulation of PBL derived human B cells, input populations and cells remaining after 4 days of culture were compared by FACS using antibodies to CD19 (all B cells), CD27 (memory B), kappa and lambda light chains, CD10 (immature B), and CD38 (immature and plasma cells). Immature B cells (CD10+, CD27−, CD38+) represented 1–3% of input B cells from the various donors (our data and [37], [38]) and were undectable on day 4 analysis under all culture conditions. No other significant change in the character of the surviving B cell population relative to the input population as assessed by these markers was observed, Figure 2 data shown from CD19 and CD27 analysis.


Human BLyS facilitates engraftment of human PBL derived B cells in immunodeficient mice.

Schmidt MR, Appel MC, Giassi LJ, Greiner DL, Shultz LD, Woodland RT - PLoS ONE (2008)

FACS analysis of B cell cultures.B cell populations were FACS analyzed for surface markers associated with resting B cells (CD45, CD19), memory cells (CD27), plasma cells (CD38) and kappa and lambda light chains on the day of isolation and after 4 days of culture either unstimulated or stimulated with human or murine BLyS. All samples were initially gated for lymphocytes by forward and side scatter. Data representative of 4 experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2527131&req=5

pone-0003192-g002: FACS analysis of B cell cultures.B cell populations were FACS analyzed for surface markers associated with resting B cells (CD45, CD19), memory cells (CD27), plasma cells (CD38) and kappa and lambda light chains on the day of isolation and after 4 days of culture either unstimulated or stimulated with human or murine BLyS. All samples were initially gated for lymphocytes by forward and side scatter. Data representative of 4 experiments.
Mentions: To determine if huBLyS conferred a selective survival advantage to a particular subpopulation of PBL derived human B cells, input populations and cells remaining after 4 days of culture were compared by FACS using antibodies to CD19 (all B cells), CD27 (memory B), kappa and lambda light chains, CD10 (immature B), and CD38 (immature and plasma cells). Immature B cells (CD10+, CD27−, CD38+) represented 1–3% of input B cells from the various donors (our data and [37], [38]) and were undectable on day 4 analysis under all culture conditions. No other significant change in the character of the surviving B cell population relative to the input population as assessed by these markers was observed, Figure 2 data shown from CD19 and CD27 analysis.

Bottom Line: Although human B cells bound both human and murine BLyS, nuclear accumulation of NF-kappaB p52, an indication of the induction of a protective anti-apoptotic response, following stimulation with human BLyS was more robust than that induced with murine BLyS suggesting a fundamental disparity in BLyS receptor signaling.Human BLyS treated recipients had on average 40-fold higher levels of serum Ig than controls and mounted a de novo antibody response to the thymus-independent antigens in pneumovax vaccine.The data indicate that production of fully immunologically competent humanized mice from PBL can be markedly facilitated by providing human BLyS.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America. Madelyn.Schmidt@umassmed.edu

ABSTRACT
The production of fully immunologically competent humanized mice engrafted with peripheral lymphocyte populations provides a model for in vivo testing of new vaccines, the durability of immunological memory and cancer therapies. This approach is limited, however, by the failure to efficiently engraft human B lymphocytes in immunodeficient mice. We hypothesized that this deficiency was due to the failure of the murine microenvironment to support human B cell survival. We report that while the human B lymphocyte survival factor, B lymphocyte stimulator (BLyS/BAFF) enhances the survival of human B cells ex vivo, murine BLyS has no such protective effect. Although human B cells bound both human and murine BLyS, nuclear accumulation of NF-kappaB p52, an indication of the induction of a protective anti-apoptotic response, following stimulation with human BLyS was more robust than that induced with murine BLyS suggesting a fundamental disparity in BLyS receptor signaling. Efficient engraftment of both human B and T lymphocytes in NOD rag1(-/-) Prf1(-/-) immunodeficient mice treated with recombinant human BLyS is observed after adoptive transfer of human PBL relative to PBS treated controls. Human BLyS treated recipients had on average 40-fold higher levels of serum Ig than controls and mounted a de novo antibody response to the thymus-independent antigens in pneumovax vaccine. The data indicate that production of fully immunologically competent humanized mice from PBL can be markedly facilitated by providing human BLyS.

Show MeSH
Related in: MedlinePlus