The prevalence of chronic obstructive pulmonary disease (COPD) is increasing dramatically in women. In fact, COPD now kills more women than breast and lung cancer combined [
Human immunodeficiency virus (HIV) has emerged as an independent risk factor for COPD in smokers, as data in the pre- and postantiretroviral (ART) era demonstrate increased susceptibility to cigarette smoke and a high percentage develop abnormalities in lung function, including loss of diffusing capacity and irreversible air-flow obstruction [
The purpose of the current study was to compare respiratory symptoms, lung function, and results of high resolution computed tomography (HRCT) scanning among HIV-infected women and men. While previous studies have defined phenotypic differences between men and women in advanced COPD in the general population [
This was a cross-sectional, baseline analysis of a prospective study performed between 2006 and 2010. The study involved the longitudinal assessment of the pulmonary status of HIV-infected subjects (
All subjects answered questions related to the presence or absence of respiratory symptoms, specifically, shortness of breath, cough, phlegm production, and wheezing.
All participants underwent complete pulmonary function testing, including spirometry, as well as measurement of lung volumes and carbon monoxide diffusing capacity according to American Thoracic Society guidelines. Predicted equations for spirometry were those of Goldman [
All subjects underwent HRCT (high resolution computed tomography) of the chest. Scans were performed on a Siemens multislice CT scanner (16-slice, 20-slice open CT, or 64-slice), without IV contrast. Inspiratory and expiratory images were performed. All scans were read by an experienced chest radiologist. The presence or absence of emphysema (bullae, thin-walled cystic spaces, or abnormal decreases in attenuation accompanied by vascular disruption) was recorded, as was the presence of bronchial dilatation, bronchial wall thickening, and air trapping as previously described [
To examine alveolar macrophage proteomics, we matched 6 female subjects with 6 male subjects of similar age, smoking history, and use of ART. Briefly, a bronchoalveolar lavage (BAL) in the right middle lobe was performed to obtain a lavage sample of approximately 50 mL for the isolation of AMs [
For first dimension electrophoresis, 100
After 2D electrophoresis, the gels were fixed and stained with SyproRuby fluorescence dye according to manufacturer’s protocol. Gel images were captured on a Typhoon 9200 laser scanner (Amersham) that offers high resolution and quantification of protein spots. Protein quantification on all 2D gels was performed using ImageMaster 2D software (Nonlinear Dynamics) [
The prevalence of respiratory symptoms, findings on high resolution chest CT, and pulmonary function testing were analyzed using linear/logistic models with sex, age, pack-years smoking history, viral load, BMI, and IV drug use as covariates. Because of skewness in the data, log transformation was performed on pack-years smoking history, residual volume, BMI, and viral load before model fitting.
Table
Baseline demographics of the study participants.
Demographics | All subjects |
---|---|
Subject number | 243 |
Male number (%) | 213 (87.6%) |
Female number (%) | 30 (12.3%) |
Age, mean (std dev) | 44.3 (8.4) |
BMI, mean (std dev) | 26.98 (8.35) |
Race | |
Caucasian (%) | 40.7 |
African American (%) | 57.6 |
Other (%) | 1.7 |
IV drug use number (%) | 43 (17.7%) |
Current smokers number (%) | 157 (64.6%) |
Pack years, mean (std dev) |
|
CD4 count, mean (std dev) |
|
Viral load, mean (std dev) |
|
Table
Comparison of sex differences in respiratory symptoms, chest CT findings, and pulmonary function testing.
Male | Female | Odds ratio/regression coefficient |
|
|
---|---|---|---|---|
Symptoms | ||||
Cough (%) | 72.8 | 51.7 | 4.33 | 0.0015 |
Phlegm (%) | 71.2 | 51.7 | 2.68 | 0.025 |
Shortness of breath (%) | 32.4 | 44.8 | 1.08 | 0.87 |
Wheeze (%) | 49.8 | 55.2 | 1.06 | 0.89 |
CT chest findings | ||||
Emphysema (%) | 45.8 | 33.3 | 1.51 | 0.39 |
Bronchial dilatation (%) | 22.2 | 16.6 | 1.21 | 0.73 |
Air trapping (%) | 29.1 | 22.2 | 2.62 | 0.083 |
Bronchial wall thickening (%) | 16.5 | 13.3 | 1.11 | 0.86 |
Pulmonary function testing | ||||
FEV1/FVC | 76.3 | 78.3 | −1.48 | 0.40 |
FEV1% predicted | 91.4 | 97.3 | −8.35 | 0.0086 |
RV% predicted | 115.1 | 99.4 | 0.116* | 0.0496 |
TLC% predicted | 103.5 | 99.8 | 2.57 | 0.42 |
DLCO% predicted | 79.9 | 75.5 | 4.35 | 0.19 |
A comparison of the HRCT findings did not reveal any differences regarding the prevalence of emphysema, bronchial dilatation, or bronchial wall thickening. Although there was no statistically significant difference in the prevalence of air trapping, there was a trend towards significance (29.1% males versus 22.2% females (
Based on pulmonary function testing, males had a lower percent predicted forced expiratory volume in one second (FEV1). The average FEV1 percent predicted was 91.4% in males versus 97.3% in females (
Table
Clinical characteristics of subjects undergoing bronchoalveolar lavage with alveolar macrophage proteomic analysis.
Gender | HAART | Current smoker | Pack years | Age | Race |
---|---|---|---|---|---|
F | Y | N | 1 | 37 | W |
F | Y | Y | 31 | 47 | W |
F | Y | Y | 16 | 48 | B |
F | Y | Y | 6 | 47 | B |
F | Y | Y | 11 | 33 | W |
F | Y | Y | 28 | 52 | W |
M | Y | N | 1 | 50 | W |
M | Y | Y | 36 | 42 | W |
M | Y | Y | 11 | 44 | B |
M | Y | Y | 11 | 51 | B |
M | Y | Y | 14 | 34 | W |
M | Y | Y | 29 | 45 | W |
Proteins that had at least a 2-fold higher expression in alveolar macrophages from men.
Spot |
Mean of normalized |
Mean of normalized |
Fold changes |
Accession number |
Protein ID | Mowse |
Observed pI/Mr |
Number of peptides |
Total seq. |
---|---|---|---|---|---|---|---|---|---|
867 | 0.04867 | 0.02083 | 2.34 | NP_006079 | Tubulin, beta, 2 | 2103 | 5.0/53.3 | 8 | 62 |
1009 | 0.05650 | 0.02167 | 2.61 | BAD96752 | Beta-actin variant | 471 | 7.5/45.1 | 8 | 31 |
1318 | 0.12600 | 0.02983 | 4.22 | P11177 | Pyruvate dehydrogenase E1 component subunit beta, mitochondrial | 427 | 6.2/39.6 | 4 | 39 |
1773 | 0.22083 | 0.06217 | 3.55 | NP_001900 | Cathepsin D preproprotein | 898 | 5.2/27.7 | 5 | 33 |
1802 | 0.06200 | 0.03067 | 2.02 | P30041 | Peroxiredoxin 6 | 364 | 6.0/27.4 | 7 | 76 |
1872 | 0.05017 | 0.01600 | 3.14 | P25787 | Proteasome subunit alpha type 2 | 265 | 6.6/25.7 | 6 | 76 |
3074 | 0.17217 | 0.07350 | 2.34 | 1IVY_A | Chain A, Physiological Dimer Hpp Precursor | 230 | 4.5/29.4 | 3 | 9 |
3157 | 0.06600 | 0.03250 | 2.03 | P05092 | Cyclophilin A | 206 | 6.4/16.4 | 3 | 52 |
3235 | 0.11967 | 0.01050 | 11.40 | P11021 | 78 kDa glucose-regulated protein | 2193 | 5.3/74.9 | 24 | 50 |
4880 | 0.09550 | 0.00567 | 16.85 | CAA68491 | Glutathione peroxidase | 1143 | 5.5/23.6 | 4 | 89 |
4925 | 0.12867 | 0.00100 | 128.67 | P26447 | Protein S100-A4 | 208 | 5.9/11.9 | 5 | 45 |
4990 | 0.11867 | 0.01933 | 6.14 | P00558 | Phosphoglycerate kinase 1 | 2011 | 7.9/44.9 | 13 | 72 |
5173 | 0.11133 | 0.03450 | 3.23 | P30048 | Thioredoxin-dependent peroxide reductase, mitochondrial | 204 | 7.7/28.0 | 3 | 16 |
5180 | 0.10050 | 0.03300 | 3.05 | Q06830 | Peroxiredoxin-1 | 636 | 7.3/22.2 | 7 | 47 |
8849 | 0.06850 | 0.00100 | 68.50 | P54727 | UV excision repair protein RAD23 homolog B | 643 | 4.8/43.2 | 8 | 41 |
8872 | 0.05933 | 0.02533 | 2.34 | P06576 | ATP synthase subunit beta, mitochondrial | 513 | 5.3/56.5 | 8 | 34 |
8985 | 0.09233 | 0.01900 | 4.86 | P06733 | Enolase 1 | 638 | 5.7/42.3 | 8 | 28 |
9003 | 0.15800 | 0.03033 | 5.21 | AAA51580 | Gamma-actin | 620 | 4.6/43.3 | 3 | 19 |
9023 | 0.63483 | 0.28000 | 2.27 | Q3ZCM7 | Tubulin beta-8 chain | 344 | 4.8/49.7 | 4 | 18 |
9103 | 0.05133 | 0.01850 | 2.77 | P06396 | Gelsolin | 1241 | 5.9/85.6 | 4 | 25 |
9126 | 0.06717 | 0.01100 | 6.11 | O95336 | 6-phosphogluconolactonase | 331 | 5.7/27.8 | 4 | 39 |
9141 | 0.10867 | 0.03400 | 3.20 | 1HJO_A | Chain A, Heat-Shock 70 kd Protein 42 kd Atpase N-Terminal Domain | 1329 | 7.4/28.6 | 5 | 45 |
9152 | 0.08583 | 0.00217 | 39.62 | P09668 | Procathepsin H | 517 | 8.4/37.4 | 3 | 29 |
9153 | 0.13433 | 0.01400 | 9.60 | P52566 | Rho GDP-dissociation inhibitor 2 | 616 | 5.1/23.0 | 10 | 75 |
9169 | 0.09733 | 0.04633 | 2.10 | AAA35607 | IgE-binding protein | 257 | 7.4/26.1 | 2 | 22 |
9337 | 0.10317 | 0.04183 | 2.47 | 1GLO_A | Chain A, Crystal Structure Of Cys25ser Mutant Of Human Cathepsin S | 193 | 6.6/16.5 | 5 | 32 |
9408 | 0.29833 | 0.12683 | 2.35 | 1AWI_A | Chain A, Human Platelet Profilin complexed with The L-Pro10 Peptide | 988 | 9.1/14.5 | 2 | 93 |
9504 | 2.49950 | 1.10067 | 2.27 | P08670 | Vimentin | 2541 | 5.1/53.6 | 12 | 63 |
9734 | 0.11817 | 0.00100 | 118.17 | P10809 | 60 kDa heat shock protein, mitochondrial | 2283 | 5.7/61.0 | 13 | 56 |
Only identified proteins are shown. Accession number: international database of the name and ID number of protein. Fold change: average density of the specific protein in males compared to females. Mowse score: measures reliability of the protein identification; greater than 100 ensures higher reliability that the protein identified is correct. Observed pI/Mr: isoelectric point and molecular weight of the protein. Ms/Ms: number of peptides sequenced after digestion. Total sequence coverage: percentage of the peptide sequenced.
Proteins that had at least a 2-fold higher expression in alveolar macrophages from women.
Spot |
Mean of normalized |
Mean of normalized |
Fold changes |
Accession number |
Protein ID | Mowse |
Observed pI/Mr |
Number of peptides |
Total seq. |
---|---|---|---|---|---|---|---|---|---|
778 | 0.0317 | 0.1257 | 3.97 | P19971 | Thymidine phosphorylase | 326 | 5.2/55.0 | 8 | 31 |
947 | 0.4037 | 0.9677 | 2.40 | O14773 | Tripeptidyl-peptidase I | 427 | 5.7/47.6 | 3 | 19 |
1526 | 0.0402 | 0.0855 | 2.13 | P07355 | Annexin A2 | 422 | 7.6/38.6 | 6 | 36 |
2342 | 0.0182 | 0.0443 | 2.44 | Q16781 | Ubiquitin-conjugating enzyme E2 N | 134 | 5.7/15.7 | 2 | 22 |
2536 | 0.0575 | 0.1312 | 2.28 | P01884 | Beta-2-microglobulin | 107 | 6.0/12.0 | 2 | 34 |
4917 | 0.2527 | 1.0122 | 4.01 | CAA23759 | Unnamed protein product | 1004 | 7.0/13.6 | 2 | 95 |
4947 | 0.0657 | 0.1440 | 2.19 | NP_004842 | Napsin A preproprotein | 1678 | 5.3/35.0 | 3 | 22 |
8844 | 0.0028 | 0.0513 | 18.12 | Q02818 | Nucleobindin-1 | 282 | 5.2/53.8 | 11 | 29 |
9158 | 0.0893 | 0.1852 | 2.07 | 1JHW_A | Chain A, Ca2+-binding mimicry in the crystal structure of the Eu3+-bound mutant human macrophage capping protein cap G | 1063 | 4.9/26.3 | 5 | 34 |
9207 | 0.0112 | 0.0510 | 4.57 | P07203 | Glutathione peroxidase | 320 | 5.5/24.2 | 4 | 54 |
9501 | 0.0447 | 0.0922 | 2.06 | P43686-2 | Proteasome 26S ATPase subunit 4 isoform 2 | 192 | 5.0/62.7 | 3 | 18 |
9629 | 0.0725 | 0.2178 | 3.00 | P07237 | Protein disulfide-isomerase | 937 | 4.8/57.1 | 8 | 34 |
9658 | 0.0245 | 0.1052 | 4.29 | P26038 | Moesin | 539 | 6.1/67.9 | 6 | 13 |
Only identified proteins are shown. Accession number: international database of the name and ID number of protein. Fold change: average density of the specific protein in males compared to females. Mowse score: measures reliability of the protein identification; greater than 100 ensures higher reliability that the protein identified is correct. Observed pI/Mr: isoelectric point and molecular weight of the protein. Ms/Ms: number of peptides sequenced after digestion. Total sequence coverage: percentage of the peptide sequenced.
The current study indicates that in a population of HIV-infected smokers there are gender differences in pulmonary function and respiratory symptoms. Males have an increased prevalence of cough and phlegm production, as well as lower % of predicted FEV1 and a higher % of predicted RV. HRCT scanning demonstrated a trend to increased air trapping. Of note, the degree of diffusion impairment was similar between the two sexes as was the degree of emphysema on chest CT. The overall findings suggest among the HIV-infected population, male smokers may be more likely to develop early airways dysfunction than female smokers.
Studies examining gender differences among individuals with COPD in the general population suggest that women may be more susceptible to the effects of cigarette smoke. In a study by Silverman and colleagues [
Our study is somewhat different from other studies that have examined established, advanced COPD; instead, we studied a group of at risk smokers finding that airway symptoms and loss of FEV1 appear to be more common in men. While airways disease seemed to be less affected in our population of women, diffusion impairment was prominent. Interestingly other studies on the non-HIV infected population have suggested that early in the presentation of COPD, women may have less airway disease, but greater diffusion impairment [
Our results differ from those of Gingo and colleagues who have recently reported lung function and respiratory symptoms among a cohort of HIV-infected subjects [
A limitation of our study was the disproportionate number of males compared to females, a reflection of the HIV-population in Central Ohio. Future studies, such as the ongoing multicenter Lung HIV study, are prospectively investigating lung function, and respiratory symptoms in over 4000 subjects [
In addition to symptoms, PFTs, and radiographic difference, we performed alveolar macrophage proteomics in a subgroup of matched females and males to explore potential mechanisms underlying the phenotypic differences in this population. A number of proteins were significantly different between the two sexes. This hypothesis generating approach involves the investigation of the protein content of a biological system [
A recent study analyzed gender differences in the proteome of BAL cells in healthy smokers and subjects with COPD. They compared healthy nonsmokers, healthy smokers, and subjects with COPD, while we compared HIV-infected smokers. However, there are some similarities with both studies finding dysregulation of lysosomal proteins in females. In their cohort there was downregulation of cathepsin B in female subjects with COPD as compared with healthy smokers [
In conclusion, among a group of relatively young HIV-infected smokers there appear to be differences in the manifestation of respiratory abnormalities, as men seem to develop airway symptoms earlier than women, while both groups have prominent abnormalities in gas exchange. Alveolar macrophage proteomics demonstrate differences in protein expression between the two groups which may provide mechanistic insight. Future study into this phenomenon is warranted as this process may provide insight into COPD pathogenesis in the general population.
Chronic obstructive pulmonary disease
Antiretroviral
High resolution computed tomography
Alveolar macrophage
Bronchoalveolar lavage
Forced expiratory volume in one second
Residual volume
Diffusing capacity for carbon monoxide
Human immunodeficiency virus.
The work was supported in part by the NIH and therefore in accordance with their public access policy is required to be placed in PubMed Central upon acceptance.
All of the authors report no financial conflict of interests in relation to the completion of this research.
The authors would like to thank Janice Drake and Karen Martin for clinical study coordination and technical support. This work was supported by the following Grants from the NIH: HL083478 and HL090313 and NCRR CTSA award UL1RR025755.