S100A9/CD163 Expression in Circulating Classical Monocytes in Chronic Obstructive Pulmonary Disease

Figures & data

Table 1. Patient characteristics.

	Nonsmokers with normal LFT (17)	Smokers with normal LFT (19)	COPD (32)
Age	58.3 ± 4.4^§	57.0 ± 3.0*	71.5 ± 1.7
Female (%)	62.5 ^¶	14.3	9.4
Smoker (%)	0	100	100
Current smoker (%)	0	100	18.8
FVC (L)	3.05 ± 0.34	3.75 ± 0.25	2.91 ± 0.21
%FVC	101.2 ± 5.4	101.6 ± 3.8	85.6 ± 5.9
FEV (L)	2.56 ± 0.34	2.90 ± 0.24	1.58 ± 0.11
%FEV	101.1 ± 2.7	90.1 ± 4.2	57.3 ± 4.4
FEV/FVC (%)	80.6 ± 2.7	78.4 ± 1.1	49.8 ± 2.4

COPD: chronic obstructive pulmonary disease; FVC: forced vital capacity; FEV_1.0: forced expiratory volume in 1 s; LFT: lung function tests.

^¶ p < 0.0001, ^§ p < 0.001, and *p < 0.01, analyzed by one-way ANOVA followed by Tukey test. Data are shown as mean ± SEM.

Figure 1. Representative flow cytometric analyses for peripheral blood circulating monocytes. Cells obtained from the peripheral blood monocytes (PBMCs) were used for analyses, and doublets and debris were excluded. Representative plots illustrate the gating strategy in samples obtained from COPD patients. (Top). CD14⁺CD16⁻—classical monocytes, CD14⁺CD16⁺—intermediate monocytes, and CD14^diminishCD16⁺—alternative monocytes were identified. (Bottom left and right). Dot plots represent isotype controls and primary antibodies against S100A9 and CD163 in classical monocytes.

Table 2. Impact of background factors on cell polarization of circulating classical monocytes.

		Patient (N)	G1	p value	G2	p value	G4	p value	Ratio	p value
Age	≤ 64 [24–64, Median 45]	20	4.1 ± 0.9	< 0.01	39.0 ± 5.9	n.s	39.6 ± 5.6	n.s	1.16 ± 0.85	n.s
	≥ 65 [65–80, Median 72]	16	9.9 ± 1.7		46.0 ± 8.0		27.1 ± 5.0		1.83 ± 0.69
Sex	Female	13	9.0 ± 2.0	n.s	35.8 ± 6.9	n.s	39.0 ± 5.0	n.s	0.58 ± 0.22	n.s
	Male	23	5.7 ± 1.3		47.6 ± 7.3		30.2 ± 5.5		2.08 ± 0.85
Cigarette	Never Smoker	17	6.6 ± 1.5	n.s	32.3 ± 5.9	n.s	45.9 ± 4.4	<0.01	0.25 ± 0.08	<0.05
	Current smoker	19	7.2 ± 1.6		53.2 ± 8.0		22.4 ± 5.3		2.64 ± 0.10

G1: S100A9⁺CD163⁻ monocytes (Gate 1); G2: S100A9⁺CD163⁺ monocytes (Gate 2); S100A9⁻CD163⁺ monocytes (Gate 4); Ratio: Gate1/Gate4. Data are shown as mean ± SEM.

Figure 2. Rates of S100A9⁺ and CD163⁺ circulating classical monocytes in three groups. (A). Rates of S100A9⁺ monocytes (Gate 1 + 2). (B). Rates of S100A9⁺CD163⁻ monocytes (Gate 1). (C). Rates of S100A9⁺CD163⁺ monocytes (Gate 2). (D). Rates of CD163⁺ monocytes (Gate 2 + 4). (E). Rates of S100A9⁻CD163⁺ monocytes (Gate 4). (F). Ratios of S100A9⁺/CD163⁺ monocytes (Gate 1/Gate 4). LFT: lung function tests. ^∗ p < 0.05, ^∗∗ p < 0.01, and ^∗∗∗ p < 0.001.

Figure 3. Receiver operating characteristic (ROC) curve analysis to assess the diagnostic validity of the ratio of S100A9⁺CD163⁻/S100A9⁻CD163⁺ cells in circulating classical monocytes in discriminating COPD patients from current smokers with normal lung functions. The area under the curve (AUC) was 0.719 (p < 0.01, 95% confidence interval [CI] = 0.567–0.871) for the diagnoses of COPD.

Table 3. Univariate and multivariate logistic regression analyses for COPD diagnoses discriminating from smokers with normal lung functions.

	Exponent	95% CI		p value	Exponent	95% CI		p value
Age	1.0912	1.0316	1.1543	0.0023**	1.107	1.0394	1.179	0.0016**
Sex	1.2083	0.1825	8.0019	0.8445	–	–	–	–
S100A9^+	1.0249	1.0040	1.0462	0.0194*	1.0336	1.0063	1.0617	0.0154*
S100A9^+CD163^−	1.0790	0.9907	1.1752	0.0809	–	–	–	–
S100A9^−CD163^+	0.9629	0.9302	0.9968	0.0324*	–	–	–	–
S100A9^+CD163^−/S100A9^−CD163^+	1.0730	0.9601	1.1992	0.2139	–	–	–	–

Abbreviation: CI; confidence interval.

**p < 0.01 and *p < 0.05.

Table 4. Specificity, sensitivity, and threshold values of each parameter with area under the receiver operating characteristic.

Parameters	Sens	Spec	Threshold	OR	95% CI	p value	AUC	95% CI	p value
S100A9 (%)	93.8	36.8	42.7	6.3	1.06-36.8	0.027*	0.607	0.430-0.784	0.236
S100A9^+CD163^- (%)	70.0	57.9	5.1	3.9	1.20-13.0	0.021*	0.691	0.528-0.854	0.021*
S100A9^-CD163^+(%)	90.6	68.4	33.4	4.5	0.96-20.7	0.044*	0.663	0.505-0.821	0.043*
S100A9^+CD163^- /S100A9^-CD163^+	90.6	47.4	0.16	8.7	1.96-38.7	0.002**	0.719	0.567-0.871	0.0048**

Abbreviations: AUC = area under the receiver operating characteristic curve; CI = confidence interval; OR = odds ratio; Sens = sensitivity; Spec = specificity.

**p < 0.01 and *p < 0.05.