Expression of CD117 (c-Kit) on Circulating B Cells in Pediatric Schistosomiasis

Abstract

Few B cells express CD27, the primary marker for memory B cells, in pediatric schistosomiasis, suggesting B cell malfunction. This study further demonstrates unexpected high expression of CD117 on circulating B cells in children highly exposed to Schistosoma mansoni infectious larvae. CD117 is expressed by immature or lymphoma B cells, but not by mature, circulating cells. We therefore sought to define the significance of CD117 on blood B cells. We found that CD117-positive (CD117⁺) B cells increased with the intensity of schistosome infection. In addition, CD117 expression was reduced on CD23⁺ B cells previously shown to correlate with resistance to infection. Stimulation with a panel of cytokines demonstrated that CD117 levels were upregulated in response to a combination of interleukin 4 (IL-4) and stem cell factor (SCF), the ligand for CD117, whereas IL-2 led to a reduction. In addition, stimulation with SCF generally reduced B cell activation levels. Upon further investigation, it was established that multiple circulating cells expressed increased levels of CD117, including monocytes, neutrophils, and eosinophils, and expression levels correlated with that of B cells. Finally, we identified a population of large circulating cells with features of reticulocytes. Overall, our results suggest that hyperexposure to intravascular parasitic worms elicits immature cells from the bone marrow. Levels of SCF were shown to reduce as children began to transition through puberty. The study results pose an explanation for the inability of children to develop significant immunity to infection until after puberty.

Keywords: B cells; CD117; CD23; pediatric schistosomiasis; schistosomiasis.

FIG 1

Increased percentages of circulating CD117⁺ B cells in human schistosomiasis. (A) Flow cytometry analysis of whole blood cells showing the forward/side scatterplot separating lymphocytes, monocytes, and mast cell precursors; large cells; neutrophils; and eosinophils used for all subsequent analyses. (B, C, and E) Cells stained with anti-CD19 (C) plus anti-CD27 or anti-CD117 (E) were used to define B cell subsets, respectively, using a threshold based on fluorescence minus one (FMO) for CD19 with no other stain. (D) Mean percentages of CD27⁺ B cells in the infected children with light (n = 37), moderate (n = 21), and heavy (n = 17) S. mansoni infection intensities. (F) Mean percentages of CD117⁺ B cells in the infected children with light (n = 48), moderate (n = 34), and heavy (n = 27) S. mansoni infection intensities. Statistically significant differences in means are indicated. Bars on dot plots represent medians. Sample sizes differ for different measures due to the unavailability of certain patient samples or reagents at the time of blood draw.

FIG 2

Demographics of CD117 expression in B cells. (A) Percentages of B cells that express CD117 by infected (n = 109) exposed, but no eggs detected (n = 16), and unexposed and uninfected samples purchased from the United States (n = 8). (B) Percentages of blood eosinophils (see Fig. 1A) by infected (n = 109), exposed but no parasites detected (n = 16), and unexposed and uninfected (n = 8). The P value between exposed and infected versus exposed and uninfected is 0.64. (C) CD117⁺ B cell levels in children infected with S. mansoni alone (n = 95), children with a coinfection of S. mansoni and malaria (n = 7), or children with a coinfection of S. mansoni and soil-transmitted helminths (STH; n = 7). (D) CD117⁺ B cell levels based on sex, female (n = 65) versus male (n = 33). (E) Percentages of B cells expressing CD117 based on age, 10 years (n = 9) 11 years (n = 33), and 12 years (n = 67). (F) CD117⁺ B cells were plotted against the infected children’s HB concentration as assessed using a portable B-hemoglobin photometer (n = 121). HB values lower than 11.5 indicate anemia. The bars in dot plots in panels A and B represent the medians. The error bars in panels D, E, and F represent standard deviation. P values demonstrate statistical differences in means.

FIG 3

Differential expression of surface markers on CD117⁺ B cells. (A) Mean ex vivo percentages of CD23⁺, CXCR5⁺, CXCR4⁺, and CD36⁺ CD117⁺ versus CD117-negative CD117⁻ B cells are shown. (B) Percentages of CD117⁺ B cells plotted against CD23⁺ B cells (n = 38). (C) Percentages of CD117⁺ B cells plotted against CD27⁺ B cells (n = 21). Data are from children infected with schistosomes. The statistically significant differences in means and correlation coefficient are indicated.

FIG 4

Effect of cytokines on CD117 expression on B cells. PBMCs were cultured in the presence of various cytokines indicated in figures for 72 h. (A and B) Effect of cytokines on the expression of CD117 by B cells cultured within PBMC [percentages (left) and mean fluorescence intensity [MFI] right]. (C) Effect of cytokines on CD117 expression on B cells from uninfected/unexposed (n = 4). (D and E) For additional control, the effect of the cytokines on CD23 levels was assessed on percentages (D) and MFI (E). Bars on dot plots represent medians. (F and G) Effect of SCF (F) or IL-4 (G) on CD69 expression on B cells. Plots demonstrate individual changes in CD69 following culture. The number of samples for blood from infected children (A, B, and D to G) ranges from 5 to 40 depending upon availability of sufficient cells or reagents at the time of bleed.

FIG 5

Association between plasma levels of SCF and CD117 expression by circulating B cells. (A and B) Plasma SCF levels were plotted against the infected children’s circulating percentage of CD117⁺ B cells (n = 53) or MFI of CD117 (n = 53). (C) SCF levels in plasma based on the age of children infected with S. mansoni; 10 (n = 21), 11 (n = 22), and 12 (n = 63) years. The mean SCF levels for each age and statistically significant differences are shown. Bars represent medians.

FIG 6

CD117 is expressed by other circulating cells in children with schistosomiasis. (A) Shown is an example of CD117 expression on monocytes. Gray fill, isotype control; black line, anti-CD117. CD117 levels on B cells were then plotted against levels measured on populations of CD14⁺ monocytes (B), neutrophils (C), eosinophils (D), or mast cell precursors (E) (CD117 low, CD14 negative, CD36 negative) (E). N = 54 for all dot plots. The correlation coefficients and P values are indicated.

FIG 7

Accumulation of reticulocyte-like cells in the blood of children. (A and B) Expression of CD117 and CD36, known to be expressed by reticulocytes, in the large cell population as shown in (Fig. 1A). Gray fill, isotype control; black line, anti-CD117 (A) or anti-CD36 (B). (C) Mean percentages of CD36^high large cells in children infected with S. mansoni (n = 93), exposed children with undetected infection (n = 13), and children uninfected and unexposed (n = 8). Bars on dot plot represent medians.