Circulating human B and plasma cells. Age-associated changes in counts and detailed characterization of circulating normal CD138- and CD138+ plasma cells

Abstract

Generation of B and plasma cells involves several organs with a necessary cell trafficking between them. A detailed phenotypic characterization of four circulating B-cell subsets (immature-, naïve-, memory- B-lymphocytes and plasma cells) of 106 healthy adults was realized by multiparametric flow cytometry. We show that CD10, CD27 and CD38 is the minimal combination of subsetting markers allowing unequivocal identification of immature (CD10(+)CD27(-)CD38(+), 6+/-6 cells/microL), naïve (CD10(-)CD27(-)CD38(-), 125+/-90 cells/microL), memory B lymphocytes (CD10(-)CD27(+)CD38(-), 58+/-42 cells/microL), and plasma cells (CD10(-)CD27(++)CD38(++), 2.1+/-2.1 cells/microL) within circulating CD19(+) cells. From these four subsets, only memory B lymphocytes and plasma cells decreased with age, both in relative and absolute counts. Circulating plasma cells split into CD138(-) (57+/-12%) and CD138(+) (43+/-12%) cells, the latter displaying a more mature phenotypic profile: absence of surface immunoglobulin, lower CD45 positivity and higher amounts of cytoplasmic immunoglobulin, CD38 and CD27. Unlike B lymphocytes, both populations of plasma cells are KI-67(+) and show weak CXCR4 expression.

Figure 1.

Phenotype of peripheral blood B-cell subsets, and focus on circulating plasma cells. (A) Distribution of immature (green), naïve (pink), memory B lymphocytes (orange), and plasma cells (red), according to the expression of CD10, CD27 and CD38. (B) Cell phenotype was analyzed by gating on CD19⁺CD20⁺CD38^−/+ - naïve and memory - B lymphocytes (green), and CD19⁺CD20⁻ CD38⁺⁺ plasma cells (blue). Histograms and dotplots show FACS labelings of cytoplasmic Ig (cyIg). The contribution of each Ig isotype is represented in the pie charts and numbers are the mean percentages ± one SD for 13 healthy individuals. (C) CD20⁻ CD38⁺⁺CD138⁺ (a) and CD20⁻ CD38⁺⁺CD138⁻ (b) peripheral blood plasma cells were FACS sorted and stained with May-Grünwald-Giemsa (×1000 magnification). (D) Open histograms show FACS labelings with indicated mAbs. Gray histograms display the corresponding negative control mAbs. Cell phenotype was analyzed by gating on CD19⁺CD20⁺ B lymphocytes, and both CD38⁺⁺CD138⁻ and CD38⁺⁺CD138⁺ plasma cells. Data from one representative experiment is shown. Numbers in panels indicate mean values of the staining indexes for each specific mAb used or the percentage of positive cells, determined on between 3 to 30 different healthy donors.

Figure 2.

Age-related changes in circulating B-cell subsets. (A) Data plotted in each diagram represent correlation between the age of each individual healthy donor and the percentage and absolute counts of total B cells (panels A and F, respectively), immature (panels B and G), naïve (panels C and H), and memory (panels D and I) B lymphocytes, as well as plasma cells (panels E and J).