Comparative proteome analysis of peripheral neutrophils from sulfur mustard-exposed and COPD patients

Figures & data

Table 1. Clinical characteristics, spirometric, hematologic, andserologic findings in study subjects.

Variables	Healthy	SM-exposed	COPD
Age (year)	54.4 ± 2.9	52.5 ± 3.62	60.08 ± 2.34
Lung function tests
FEV1 (%)	92.3 ± 12.5	60.3 ± 17.6*	58.9 ± 21*
FEV1/FVC	95.6 ± 6.4	68.2 ± 3.5*	60.2 ± 4.8*
Cell differential count (blood)
Polymorphonuclear (%)	57.2 ± 5.9	58.0 ± 9.1	56.2 ± 15.4
Lymphocyte (%)	34.9 ± 3.7	33.9 ± 7.0	34.8 ± 10
Monocyte (%)	5.3 ± 4.5	5.8 ± 2.8	6.4 ± 3.8
Eosinophil (%)	0.3 ± 0.1	0.4 ± 0.0	0.3 ± 0.1
Serology and enzymology
CRP (mg/L)	2.8 ± 0.1	3.0 ± 0.3	3.5 ± 1.2
Elastase (U/mg protein)	0.3 ± 0.1	1.1 ± 0.2^#	0.9 ± 0.1^#

Data shown are mean ± SEM (n = 10/subject group).

*p < 0.05, #p < 0.01 versus healthy control value (Tukey’s post-hoc test).

Figure 1. Representative 2-D protein patterns of PMN from study subjects. (A) Healthy control. (B) COPD patient. (C) SM-exposed subject. Sample preparation and 2-D analysis was performed as described in Materials and methods. Proteins (500 µg/subject) were separated first using linear IPG strips (pH 4–7) followed by 12% SDS-PAGE. Protein spots were then visualized by colloidal CBB staining.

Table 2. Sequence coverage (in %) and MASCOT scores of proteins spots analyzed by MS/Ms.

Spot No.^a	Protein name	Database ID^b	MW (kDa)/pI	Score value^c	Coverage (%)^d
1	S100-A9	gi4506773	13.2/5.71	308	64%
2	S100-A12	gi5032059	10.5/5.83	387	48%
3	S100-A12	gi5032059	10.5/5.83	295	48%
4	S100-A8	gi21614544	10.8/6.5	292	47%
5	GAPDH	gi31645	36.2/8.26	102	22%
6	SOD [Cu-Zn]	gi4507149	16.1/5.7	399	83%
7	Coactosin-like protein	gi21624607	16.0/5.54	267	87%
8	PDI(Chain A)	gi159162689	13.3/5.94	531	60%
9	Rho GDPDI	gi36038	23.2/5.02	372	44%
10	Rho GDPDI 2	gi56676393	23.0/5.1	301	53%
11	GST	gi2204207	23.5/5.43	403	49%
12	GST P	gi4504183	23.5/5.43	344	57%
13	GST Omega 1	gi4758484	27.8/6.23	374	37%
14	Actin isoforms	gi15277503	40.5/5.55	448	48%
15	Actin isoforms	gi15277503	40.5/5.55	447	55%
16	Actin isoforms	gi15277503	40.5/5.55	605	65%
17	Actin isoforms	gi15277503	40.5/5.55	496	53%
18	Grp78(Chain A)	gi320089786	42.2/6	481	53%
19	ER-protease 60	gi1208427	57.1/5.98	717	47%
20	SerpinB1	gi13489087	42.8/5.9	1000	54%
21	Coronin-1A	gi5902134	51.6/6.25	530	53%

^aSpot number related to Figure 1 annotation. Mass quality determined using BSA and transferrin (Tf) as internal standards. Score/coverage: BSA = 1060/52 and Tf = 254/25.

^bNRDB1 (6655203 protein sequence).

^cMASCOT score value indicates the quality of database search results.

^dSequence coverage refers to the observed sequence coverage of the assigned protein.

Table 3. Up-/down-regulated proteins in PMN of SM-exposed and COPD patients versus PMN of healthy controls.

Spot No.#	Protein	Healthy	SM-exposed	COPD
1	S100-A9	3.8 ± 0.3	3.6 ± 0.2 ↔	8.3 ± 0.3* ↑
2	S100-A12	4.3 ± 0.1	6.8 ± 0.1* ↑	9.2 ± 0.2* ↑
3	S100-A12	1.4 ± 0.0	3.2 ± 0.3* ↑	4.8 ± 1.8* ↑
4	S100-A8	1.1 ± 0.0	1.8 ± 0.2* ↑	2.7 ± 0.8* ↑
5	GAPDH	5.5 ± 0.4	8.8 ± 0.8* ↑	10.2 ± 0.1* ↑
6	SOD	1.2 ± 0.1	2.4 ± 0.1* ↑	2.1 ± 0.2* ↑
7	Coactosin-like protein	3.3 ± 0.3	1.1 ± 0.4* ↓	1.4 ± 0.5* ↓
8	PDI	1.2 ± 0.0	2.0 ± 0.1* ↑	1.9 ± 0.1* ↑
9	Rho GDPDI	2.7 ± 0.1	1.6 ± 0.1* ↓	1.5 ± 0.0* ↓
10	Rho GDPDI 2	4.2 ± 0.4	1.8 ± 0.2* ↓	1.9 ± 0.3* ↓
12	GST P	8.1 ± 1.2	7.1 ± 0.7 ↔	4.6 ± 0.2* ↓
13	GST Omega 1	1.0 ± 0.0	1.3 ± 0.1 ↔	1.8 ± 0.2* ↑
14	Actin isoforms	8.6 ± 1.6	4.2 ± 0.1* ↓	4.6 ± 1.3* ↓
15	Actin isoforms	11.8 ± 1.3	4.4 ± 1.0* ↓	4.8 ± 1.4* ↓
16	Actin isoforms	17.6 ± 3.6	6.0 ± 1.4* ↓	7.2 ± 2.9* ↓
17	Actin isoforms	5.8 ± 0.6	2.2 ± 0.3* ↓	1.9 ± 0.2* ↓
19	ER-protease 60	1.9 ± 0.1	3.4 ± 0.2* ↑	1.1 ± 0.0* ↓
20	SerpinB1	3.0 ± 0.5	Not detectable	Not detectable
21	Coronin-1A	3.5 ± 0.7	Not detectable	Not detectable

Data shown are mean (±SEM; n = 10/subject group) of % volume of protein spots in 2-D gels.

*p < 0.05 versus healthy control value (Tukey’s post-hoc test). Significantly increased (↑), decreased (↓) or no differences (↔).

#Spot numbers corresponding to those in Figure 1.

Figure 2. Representative 3-D images of differentially-expressed proteins in PMN. Differentially-expressed proteins spots in lysate from PMN of the same control, SM-exposed, and COPD subject shown in Figure 1. (A) PDI (Spot 8) and SOD (Spot 6). (B) SerpinB1 (Spot 20) and S100A12 (Spot 2). Data were analyzed using Image master 2-D platinum software.

Figure 3. Differenially-expressed proteins in PMN from SM-exposed and COPD patients. Protein spots expressed differently in the PMN of the same SM-exposed and COPD patients shown in Figure 1. (A) ER-proteses 60 (Spot 19) and S100A9 (spot 1). (B) GSTp (Spot 12) and GST omega1 (Spot 13). The 3-D images of differential spots between the patients were analyzed using ImageMaster 2D Platinum software.