Figure 2.
Different developmental dynamics of BM CD34+ cells from CVID patients distinguish intrinsic defects from nonpermissive BM environment. (A) The table indicates the distribution of early B-cell stages in BM aspirates, analyzed by flow cytometry in the cohort of CVID patients and in patients with defined mutations in BTK (BTK1, BTK2) and IKZF1 gene (Ikaros). Pro-B-cells are defined as CD22+ CD79a+ CD34+ CD22+, pre-BI cells as CD19+, cytoplasmic IgM−, cytoplasmic CD179a+, pre-BII cells as CD19+, cytoplasmic IgM−, IgM−. Also, peripheral B-cell counts per microliter and the percentages (%) of transitional B cells in the CD19 B-cell population are indicated. In gray shade is the reference range, high values are indicated in red, and low values in blue. Pro-B: pro-B-cells; pre-BI and pre-BII: pre-BI and pre-BII cells. *Lymphoid precursors 0.7% (reduced, normal range 2% to 22%). Below det.: below detection level; Not ass.: not assessed. (B) Distribution of developing B-cell subpopulations over time in culture starting from CVID CD34+ cells. Live CD10+ cell counts (top) and proportion of immature B cells (ImmB, %, bottom) within the CD10+ gate at days 14, 21, and 49 in HD and CVID patients. CD34+ cells from HD (yellow), from CVID patients with normal BM analysis ex vivo (group 1, blue), from CVID patients with altered early B-cell development ex vivo (group 2, red: P2, P6, and P11; group 3, purple: P5, P7, P8, P14, and P15; group 4, green: P3 and P4). Each experiment was performed with 3 to 10 replicates. Each symbol represents 1 patient. Mean and standard error of mean are represented. Filled symbols represent NFkB1-deficient (P8 in purple and P13 in blue) patients. (C) Quantitative polymerase chain reaction analysis of expression levels relative to CD79a of transcription factors driving B-cell specification (E2A) and commitment (PAX5) in cultivated B cells. Mean (line) and standard error of mean (shade) are shown and evaluated on 7 HD, 3 CVID patients in group 1 (blue), 2 patients from group 2 (red), 3 patients from group 3 (purple), and 1 patient from group 4 (green).

Different developmental dynamics of BM CD34+ cells from CVID patients distinguish intrinsic defects from nonpermissive BM environment. (A) The table indicates the distribution of early B-cell stages in BM aspirates, analyzed by flow cytometry in the cohort of CVID patients and in patients with defined mutations in BTK (BTK1, BTK2) and IKZF1 gene (Ikaros). Pro-B-cells are defined as CD22+ CD79a+ CD34+ CD22+, pre-BI cells as CD19+, cytoplasmic IgM−, cytoplasmic CD179a+, pre-BII cells as CD19+, cytoplasmic IgM−, IgM−. Also, peripheral B-cell counts per microliter and the percentages (%) of transitional B cells in the CD19 B-cell population are indicated. In gray shade is the reference range, high values are indicated in red, and low values in blue. Pro-B: pro-B-cells; pre-BI and pre-BII: pre-BI and pre-BII cells. *Lymphoid precursors 0.7% (reduced, normal range 2% to 22%). Below det.: below detection level; Not ass.: not assessed. (B) Distribution of developing B-cell subpopulations over time in culture starting from CVID CD34+ cells. Live CD10+ cell counts (top) and proportion of immature B cells (ImmB, %, bottom) within the CD10+ gate at days 14, 21, and 49 in HD and CVID patients. CD34+ cells from HD (yellow), from CVID patients with normal BM analysis ex vivo (group 1, blue), from CVID patients with altered early B-cell development ex vivo (group 2, red: P2, P6, and P11; group 3, purple: P5, P7, P8, P14, and P15; group 4, green: P3 and P4). Each experiment was performed with 3 to 10 replicates. Each symbol represents 1 patient. Mean and standard error of mean are represented. Filled symbols represent NFkB1-deficient (P8 in purple and P13 in blue) patients. (C) Quantitative polymerase chain reaction analysis of expression levels relative to CD79a of transcription factors driving B-cell specification (E2A) and commitment (PAX5) in cultivated B cells. Mean (line) and standard error of mean (shade) are shown and evaluated on 7 HD, 3 CVID patients in group 1 (blue), 2 patients from group 2 (red), 3 patients from group 3 (purple), and 1 patient from group 4 (green).

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