Figure 5
Figure 5. Impaired migration and adhesion of Rap1b-deficient B cells. (A) Chemokine-mediated migration of wild-type and Rap1b-deficient splenic B cells. Purified total splenic B cells from wild-type (+/+) or Rap1b-deficient (−/−) mice were subjected to in vitro transwell migration assays to BLC, SLC, SDF-1, and medium alone. Duplicates of input cells (from aliquots) and migrated cells were quantified by flow cytometry. The number of cells migrated is expressed as the percentage of total input cells. Data shown are from 3 independent experiments for BLC and SLC and 5 independent experiments for SDF-1. (B) Chemokine-mediated migration of wild-type and Rap1b-deficient MZ and FO B cells. Total splenocytes from wild-type or Rap1b-deficient mice were subjected to in vitro transwell migration assays to BLC, SLC, SDF-1, and medium alone. Duplicates of input cells (from aliquots) and migrated cells were stained with anti-B220, anti-CD21, anti-CD23, and anti-IgM antibodies. Input and migrated MZ and FO B cells were quantified by flow cytometry. The numbers of MZ and FO cells migrated are expressed as the percentages of input MZ and FO B cells, respectively. Data shown are from 3 independent experiments for BLC and SLC and 5 independent experiments for SDF-1. (C) Adhesion of wild-type and Rap1b-deficient splenic B cells to VCAM-1. Purified total splenic B cells from wild-type or Rap1b-deficient mice were allowed to adhere to VCAM-1–coated 96-well plates. Nonadherent cells were removed by washing. Triplicate of input and adherent cells were counted and analyzed by FACS. The adhesion of total B cells, MZ, and FO B cells was calculated as a percentage of total, MZ, or FO B cell input, respectively. Data shown are representative of 3 independent experiments. (D) The proportions of immature and mature B cells in the peripheral blood of wild-type and Rap1b-deficient mice. Red cell–depleted blood cells were stained with anti-B220, anti-IgM, and anti-IgD antibodies. Percentages indicate cells in the gated B220+ B cells. Data are representative of 6 mice per genotype.

Impaired migration and adhesion of Rap1b-deficient B cells. (A) Chemokine-mediated migration of wild-type and Rap1b-deficient splenic B cells. Purified total splenic B cells from wild-type (+/+) or Rap1b-deficient (−/−) mice were subjected to in vitro transwell migration assays to BLC, SLC, SDF-1, and medium alone. Duplicates of input cells (from aliquots) and migrated cells were quantified by flow cytometry. The number of cells migrated is expressed as the percentage of total input cells. Data shown are from 3 independent experiments for BLC and SLC and 5 independent experiments for SDF-1. (B) Chemokine-mediated migration of wild-type and Rap1b-deficient MZ and FO B cells. Total splenocytes from wild-type or Rap1b-deficient mice were subjected to in vitro transwell migration assays to BLC, SLC, SDF-1, and medium alone. Duplicates of input cells (from aliquots) and migrated cells were stained with anti-B220, anti-CD21, anti-CD23, and anti-IgM antibodies. Input and migrated MZ and FO B cells were quantified by flow cytometry. The numbers of MZ and FO cells migrated are expressed as the percentages of input MZ and FO B cells, respectively. Data shown are from 3 independent experiments for BLC and SLC and 5 independent experiments for SDF-1. (C) Adhesion of wild-type and Rap1b-deficient splenic B cells to VCAM-1. Purified total splenic B cells from wild-type or Rap1b-deficient mice were allowed to adhere to VCAM-1–coated 96-well plates. Nonadherent cells were removed by washing. Triplicate of input and adherent cells were counted and analyzed by FACS. The adhesion of total B cells, MZ, and FO B cells was calculated as a percentage of total, MZ, or FO B cell input, respectively. Data shown are representative of 3 independent experiments. (D) The proportions of immature and mature B cells in the peripheral blood of wild-type and Rap1b-deficient mice. Red cell–depleted blood cells were stained with anti-B220, anti-IgM, and anti-IgD antibodies. Percentages indicate cells in the gated B220+ B cells. Data are representative of 6 mice per genotype.

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