Differential serum protein markers and the clinical severity of asthma

Figures & data

Table 1 Characterization of asthma patients

Parameters	Cluster 1	Cluster 2	Unpaired t-test	Pearson’s chi-squared test	Number
Allergic asthma	70.0%	80.2%		0.122	191
Age (years)	54.4±12.0 (21–79)	48.1±15.9 (18–80)	0.003		191
Age of onset (years)	23.2±17.5 (0–52)	24.2±16.8 (0–65)	0.718		191
Sex (male)	43.3%	44.3%		0.903	191
Smoker	5.0%	3.1%		0.506	191
Pack years	4.2±6.8 (0–30)	4.0±7.3 (0–35)	0.871		191
BMI (kg/m^2)	24.8±4.1 (17.0–39.0)	25.7±3.8 (19.0–36.0)	0.129		191
ATS criteria	56.7%	18.3%		<0.001	191
Exacerbations during last year	5.4±3.5	3.8±3.0	0.004		191
Asthma control (%)					191
NC	71.7	32.1
PC	25.0	40.5
FC	3.3	27.5
Adherence to therapy	90.9%	93.4%		0.571	191
Serum IgE level (kU/l)	457.8±991.1 (0–5000)	620.8±2455.0 (0–26973)	0.516		190
Long-acting β2 agonist	100%	89.3%		0.009	191
Short-acting β2 agonist (puffs per day)	3.1±3.8 (0–20)	1.4±2.8 (0–20)	0.002		191
Systemic steroids	43.3%	19.1%		<0.001	191
Inhaled steroids	98.3%	93.1%		0.134	191
Theophylline	36.7%	15.3%		0.001	191

Notes: Asthma patients belonging to cluster 1 or 2 are characterized by indicated parameters. Percentages show the fraction of patients fulfilling indicated parameters/characteristics. Age, Age of onset, Pack years, BMI, serum IgE level and Short-acting β2 agonist are presented as mean ± SD (range). Asthma exacerbation is presented as mean ± SD.

Abbreviations: ATS, American Thoracic Society; BMI, body mass index; FC, fully controlled; IgE, immunoglobulin E; NC, not controlled; PC, partly controlled.

Figure 1 Classification of asthma patients according to airway inflammation.

Notes: 191 asthma patients were classified by hierarchical cluster analysis with indicated parameters. (A) Dendrogram. (B) At entry to the high-altitude clinic, FEV₁ values, ECP and blood eosinophil levels, and Junipers asthma symptom score were evaluated for cluster 1 and 2. (C) Changes in FEV₁, ECP, and blood eosinophils after 3 weeks of asthma therapy from cluster 1 and 2. C1: asthma patients in cluster 1; C2: asthma patients in cluster 2; *P<0.05, ***P<0.001; unpaired t-test was used.
Abbreviations: ECP, eosinophil cationic protein; FEV₁, forced expiratory volume in 1 second; n, number of asthma patients.

Figure 2 Asthma patients in cluster 1 respond better to asthma therapy.

Notes: (A) FEV₁, serum ECP levels, blood eosinophil levels and exhaled NO at the beginning of the high altitude stay (entry) and after 3 weeks (discharge). (B) Fraction of patients with systemic steroids and inhaled steroids, systemic steroid dose and frequency of inhaled short acting β2 agonist usage. The paired t-test was used to compare changes in indicated parameters between entry and discharge. **P<0.01; ***P<0.001.
Abbreviations: ECP, eosinophil cationic protein; FEV₁, forced expiratory volume in 1 second; n, number of asthma patients; NO, nitric oxide.

Figure 3 Upregulation of serum cytokines and chemokines in asthma patients belonging to uster 1.

Notes: Serum levels of IL-8, eotaxin, VEGF, CTACK, GROα, and HGF in patients belonging to cluster 1 or to cluster 2 are shown. Mann–Whitney U test was used. *P<0.05.
Abbreviations: CTACK, cutaneous T-cell-attracting chemokine; GRO, growth-related oncogene; HGF, hepatocyte growth factor; IL, interleukin; VEGF, vascular endothelial growth factor.

Figure 4 Prognostic value of serum cytokines to distinguish between asthma patients belonging to cluster 1 or 2.

Notes: ROC for the principal component of the serum cytokines IL-8, eotaxin, VEGF, CTACK, GROα, and HGF to distinguish between cluster 1 and 2 was analyzed. (A) Area under the curve produced a sensitivity of 55.9% and a specificity of 75.8% to discriminate between asthma patients from cluster 1 and cluster 2. (B) ROC analyses of the single cytokines. (C) There is a negative correlation between cytokine expression assessed by the principal component analyses of the serum cytokines upregulated in cluster 1 and FEV₁ values.
Abbreviations: CTACK, cutaneous T-cell-attracting chemokine; FEV₁, forced expiratory volume in 1 second; GRO, growth-related oncogene; HGF, hepatocyte growth factor; IL, interleukin; ROC, receiver operating characteristic; VEGF, vascular endothelial growth factor.

Table S1 Serum cytokine and chemokine levels in asthma patients belonging to cluster 1 or to cluster 2

	Cluster 1	Cluster 2	P-value
IL-1Ra	1,465.7±1,741.6	1,927.3±3,958.4	0.34
IL-2	62.8±97.0	90.4±334.3	0.34
IL-4	67.9±24.8	63.1±26.9	0.14
IL-9	864.1±2,783.0	631.1±3,446.8	0.23
IFN-γ	2,057.1±2,207.5	2,070.8±3,370.4	0.15
MIP-1β	510.5±509.2	1,228.1±858.3	0.19
TNF-α	237.4±217.1	262.8±348.5	0.31
IL-2Ra	1,016.5±1,140.8	676.4±574.2	0.06
IL-18	643.5±361.6	568.9±324.1	0.11
MIF	1,2541.2±7,713.4	10,153.0±5,276.7	0.10
MIG	13,156.3±19,172.1	9,572.8±19,840.7	0.10
SCF	894.2±418.2	804.4±326.6	0.21
SCGFβ	199,001.3±133,999.6	159,837.8±107,284.1	0.06
G-CSF	211.0±213.8	184.0±202.8	0.34
FGF	232.6±262.8	206.6±298.4	0.33
IL-6	62.7±70.9	62.5±142.5	0.08
IL-10	57.8±291.0	36.2±134.7	0.09
IL-15	41.1±62.8	26.9±57.1	0.07
MIP-1α	26.2±25.7	20.3±23.5	0.05
IFN-α	39.8±116.7	18.5±65.4	0.16
LIF	86.7±557.1	12.5±27.9	0.36
MCP-3	146.4±956.3	47.9±154.1	0.83
MCSF	27.6±61.0	22.0±58.3	0.32
SDF-1α	189.1±315.1	151.6±264.9	0.52
IL-13	43.9±87.9	36.2±75.5	0.06

Notes: Serum levels of cytokines and chemokines, which were not significantly different between cluster 1 and cluster 2 are shown. Mann–Whitney U test was used. Concentrations are shown in pg/mL ± SD.

Abbreviations: IL, interleukin; IFN-γ, gamma-interferon; MIP-1β, macrophage inflammatory protein 1 beta; TNF-α, tumor necrosis factor-alpha; MIF, macrophage migration inhibitory factor; MIG, monokine-induced by interferon-gamma; SCF, stem cell factor; SCGFβ, stem cell growth factor beta; G-CSF, granulocyte colony-stimulating factor; FGF, fibroblast growth factor; MIP-1α, macrophage inflammatory protein 1 alpha; IFN-α, alpha-interferon; LIF, leukemia inhibitory factor; MCP-3, monocyte chemotactic protein-3; MCSF, macrophage colony-stimulating factor; SDF-1α, stromal cell-derived factor-1 alpha; SD, standard deviation.