Figures
- FIGURE 1
Patients with sarcoidosis have increased nonclassical monocytes and reduced classical monocytes compared with controls. Flow cytometric analysis of peripheral blood mononuclear cells (PBMCs) was used to define monocyte subsets based on expression of CD14 and CD16. Ungated PBMCs are shown here for illustrative purposes. a) Monocytes were defined as classical (CD14++/CD16–/low: “C”), intermediate (CD14++/CD16+: “I”) or nonclassical (CD14+/CD16++: “NC”). b–d) Proportions of b) classical, c) intermediate and d) nonclassical monocyte subsets within total blood monocytes for patients with sarcoidosis (n=25) and healthy controls (n=12). Individual data points are presented along with median and interquartile range. **: p<0.01 (Mann–Whitney U-test).
- FIGURE 2
Nonclassical monocytes express low levels of CD200R and expression of CD200R is lower on all monocyte subsets in patients with sarcoidosis compared with healthy controls. GMFI: geometric mean fluorescence intensity. a) Representative flow cytometry histograms from a healthy control subject comparing CD200R expression on classical, intermediate and nonclassical monocytes compared with isotype control antibody. b, c) Representative heat-map dot plots from b) a healthy control subject demonstrating low CD200R on nonclassical monocytes and c) a patient with sarcoidosis showing reduced CD200R expression on sarcoidosis monocyte subsets compared with the healthy control. On these pseudo-colour plots, orange indicates higher CD200R expression and green indicates lower CD200R expression. d–f) Expression of CD200R on d) classical, e) intermediate and f) nonclassical monocyte subsets in sarcoidosis patients and controls. Note the small y-axis scale for nonclassical monocytes. Individual data points are presented along with median and interquartile range. *: p<0.05; **: p<0.01 (Mann–Whitney U-test).
- FIGURE 3
CD200R expression on sarcoidosis classical and intermediate monocytes is dichotomous and consistent over time. GMFI: geometric mean fluorescence intensity. Patients with sarcoidosis in a longitudinal cohort (n=19) underwent serial measurements of CD200R expression on a) classical, b) intermediate and c) nonclassical monocyte subsets. Each patient is presented by a different coloured line.
- FIGURE 4
Oral corticosteroid therapy for sarcoidosis normalises monocyte subsets but has no effect on monocyte CD200R expression. GMFI: geometric mean fluorescence intensity. Pre- and 2-month post-steroid treatment data for three sarcoidosis patients who were treated with oral prednisolone 20 mg·day−1. a, c, e) Percentages of classical, intermediate and nonclassical monocytes, respectively. b, d, f) CD200R expression on the respective monocyte subsets.
- FIGURE 5
Monocyte subsets have distinct expression profiles of signal regulatory protein-α (SIRP-α), CD47 and interleukin (IL)-10R in sarcoidosis and healthy controls. Expression of a–c) SIRP-α, d–f) CD47 and g–i) IL-10R on a, d, g) classical, b, e, h) intermediate and c, f, i) nonclassical monocyte subsets in patients with sarcoidosis (n=25) and healthy controls (n=7). Individual data points are presented along with median and interquartile range. **: p<0.01; ***: p<0.001; ****: p<0.0001 (Mann–Whitney U-test).
- FIGURE 6
Distribution of the regulatory receptor CD200R and its ligand CD200L in sarcoidosis granulomas: transbronchial lung biopsy samples from two patients with sarcoidosis stained for a) CD200L and b) CD200R, with respective isotype control antibody staining in c) and d). f: fibroblasts; h: histiocytes (macrophages). Original magnification ×100.
Tables
- TABLE 1
Demographic and clinical profiles of patients with sarcoidosis and healthy controls
Sarcoidosis Controls Patients 25 12 Age years 53 (38–73) 51 (30–67) Sex Male 15 7 Female 10 5 Time since diagnosis months 22 (0–193) Scadding chest radiograph stage 0 3 1 5 2 12 3 5 Angiotensin converting enzyme U·L−1# 94 (9–440) Blood lymphocyte count ×109 L−1¶ 0.86 (0.29–2.03) FEV1 % pred 93 (54–126) FVC % pred 95 (64–129) FEV1/FVC 0.81 (0.49–0.91) Data are presented as n or median (range). FEV1: forced expiratory volume in 1 s; FVC: forced vital capacity. #: reference range 8–65 U·L−1; ¶: reference range 0.8–3.4×109 L−1.
- TABLE 2
Characteristics of sarcoidosis patients stratified by monocyte CD200R phenotype
CD200Rlow CD200Rhigh p-value Patients 14 11 Scadding chest radiograph stage 0.265 0 1 2 1 4 1 2 5 7 4 4 1 Angiotensin converting enzyme U·L−1 92 (52–136) 104 (74–155) 0.695 C-reactive protein mg·L−1 2.0 (1.4–5.5) 2.8 (0.6–9.6) 0.217 Age years# 53.5 (40–73) 50.0 (38–67) 1.00 FEV1 % pred 99 (69–110) 92 (82–113) 0.428 FVC % pred 103 (76–115) 90 (82–114) 0.428 FEV1/FVC 0.79 (0.75–0.84) 0.83 (0.76–0.86) 0.238 Time since diagnosis months 20 (9–51) 31 (10–73) 0.695 Sex 1.00 Male 8 7 Female 6 4 White blood cell count ×109 L−1 5.4 (4.15–6.20) 4.3 (3.50–5.60) 0.196 Lymphocyte count ×109 L−1 0.93 (0.74–1.41) 0.74 (0.62–1.02) 0.279 Monocyte count ×109 L−1 0.55 (0.44–0.72) 0.42 (0.24–0.70) 0.252 Neutrophil count ×109 L−1 3.42 (2.70–3.98) 2.70 (2.33–3.97) 0.336 Eosinophil count ×109 L−1 0.15 (0.13–0.23) 0.20 (0.08–0.21) 0.797 T-lymphocyte count ×109 L−1 0.61 (0.36–0.91) 0.48 (0.33–0.63) 0.403 CD4+ T-lymphocyte count ×109 L−1 0.36 (0.22–0.70) 0.30 (0.16–0.45) 0.596 CD8+ T-lymphocyte count ×109 L−1 0.20 (0.11–0.40) 0.17 (0.09–0.21) 0.290 B-lymphocyte count ×109 L−1 0.16 (0.07–0.21) 0.14 (0.08–0.23) 0.499 Natural killer cell count ×109 L−1 0.17 (0.12–0.22) 0.15 (0.09–0.26) 0.450 Data are presented as n or median (interquartile range), unless otherwise stated. FEV1: forced expiratory volume in 1 s; FVC: forced vital capacity. #: median (range). Statistical comparisons by nonparametric independent samples median test or Pearson's Chi-squared test.
Supplementary Material
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Supplementary material 00804-2020.SUPPLEMENT
