Figure 3
Figure 3. miR-142 functions in a cell-autonomous manner to control the development of B and T lymphocytes. (A) Diagram of mixed BM chimera generation. WT or miR-142−/− (CD45.2+) BM cells were mixed at 1:1 ratio with helper WT (CD45.1+) BM cells and transplanted into lethally irradiated WT (CD45.1+) hosts. Resulting chimeric mice were analyzed 12-17 weeks posttransplantation. (B) Splenomegaly in WT mice receiving miR-142−/− BM. Spleen weight in mice reconstituted with a mixed WT BM (WT/WT→WT) or a mixture of WT and KO BM cells (KO/WT→WT). (C-D) B1 B cell differentiation defect in mixed miR-142−/− BM chimeras. (C) FACS analysis of lymphocytes in peritoneal cavity of mixed BM chimeric mice with anti-B220 and anti-CD19 specific antibodies. B1 (CD19+B220lo) and B2 (CD19+B220hi) cell populations are gated and numbers indicate the percentage of cells in the gate. Cells were first gated on the basis of CD45.1 (left panel) or CD45.2 (right panel) expression. (D) Percentage of B1 B cells in peritoneal cavity of WT/WT→WT (n = 4) and KO/WT→WT (n = 4) mixed BM chimeric mice. (E-G) Excessive production of MZ and NFB B cells in mixed KO/WT→WT BM chimeras. (E) FACS analysis of B220+ B lymphocytes in the spleen of mixed BM chimeric mice with anti-CD21 and anti-CD23 specific antibodies. FO (CD21intCD23+), MZ (CD21hiCD23lo), and NFB (CD21intCD23lo) B-cell subsets are gated and numbers indicate the percentage of cells in the gate. Cells were first gated on the basis of CD45.1 (left panel) or CD45.2 (right panel) expression. (F) Percentage of MZ B cells in the spleens of WT/WT→WT (n = 4) and KO/WT→WT (n = 4) mixed BM chimeric mice. (G) Percentage of NFB cells in spleens of WT/WT→WT (n = 4) and KO/WT→WT (n = 4) mixed BM chimeric mice. (H-I) Decrease in the number of peripheral T cells in the mixed miR-142−/− BM chimeras. (H) FACS analysis of lymphocytes in the spleens of mixed BM chimeric mice with anti-CD3ε specific antibodies. T cells (CD3+) are gated and numbers indicate the percentage of cells in the gate. Cells were first gated on the basis of CD45.1 (left panel) or CD45.2 (right panel) expression. (I) Percentage of CD3+ T cells in the spleens of WT/WT→WT (n = 3) and KO/WT→WT (n = 3) mixed BM chimeric mice. Results are shown as means ± SD and are representative of 2 independent experiments. P values were calculated using either uncorrected Fisher’s least significant difference test or Student t test. ns, not significant; **P ≤ .01; ***P ≤ .001; ****P ≤ .0001. NF, newly formed.

miR-142 functions in a cell-autonomous manner to control the development of B and T lymphocytes. (A) Diagram of mixed BM chimera generation. WT or miR-142−/− (CD45.2+) BM cells were mixed at 1:1 ratio with helper WT (CD45.1+) BM cells and transplanted into lethally irradiated WT (CD45.1+) hosts. Resulting chimeric mice were analyzed 12-17 weeks posttransplantation. (B) Splenomegaly in WT mice receiving miR-142−/− BM. Spleen weight in mice reconstituted with a mixed WT BM (WT/WT→WT) or a mixture of WT and KO BM cells (KO/WT→WT). (C-D) B1 B cell differentiation defect in mixed miR-142−/− BM chimeras. (C) FACS analysis of lymphocytes in peritoneal cavity of mixed BM chimeric mice with anti-B220 and anti-CD19 specific antibodies. B1 (CD19+B220lo) and B2 (CD19+B220hi) cell populations are gated and numbers indicate the percentage of cells in the gate. Cells were first gated on the basis of CD45.1 (left panel) or CD45.2 (right panel) expression. (D) Percentage of B1 B cells in peritoneal cavity of WT/WT→WT (n = 4) and KO/WT→WT (n = 4) mixed BM chimeric mice. (E-G) Excessive production of MZ and NFB B cells in mixed KO/WT→WT BM chimeras. (E) FACS analysis of B220+ B lymphocytes in the spleen of mixed BM chimeric mice with anti-CD21 and anti-CD23 specific antibodies. FO (CD21intCD23+), MZ (CD21hiCD23lo), and NFB (CD21intCD23lo) B-cell subsets are gated and numbers indicate the percentage of cells in the gate. Cells were first gated on the basis of CD45.1 (left panel) or CD45.2 (right panel) expression. (F) Percentage of MZ B cells in the spleens of WT/WT→WT (n = 4) and KO/WT→WT (n = 4) mixed BM chimeric mice. (G) Percentage of NFB cells in spleens of WT/WT→WT (n = 4) and KO/WT→WT (n = 4) mixed BM chimeric mice. (H-I) Decrease in the number of peripheral T cells in the mixed miR-142−/− BM chimeras. (H) FACS analysis of lymphocytes in the spleens of mixed BM chimeric mice with anti-CD3ε specific antibodies. T cells (CD3+) are gated and numbers indicate the percentage of cells in the gate. Cells were first gated on the basis of CD45.1 (left panel) or CD45.2 (right panel) expression. (I) Percentage of CD3+ T cells in the spleens of WT/WT→WT (n = 3) and KO/WT→WT (n = 3) mixed BM chimeric mice. Results are shown as means ± SD and are representative of 2 independent experiments. P values were calculated using either uncorrected Fisher’s least significant difference test or Student t test. ns, not significant; **P ≤ .01; ***P ≤ .001; ****P ≤ .0001. NF, newly formed.

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