The principal finding of this study was that the serum concentrations of SAA, SP-D and IL-4 were greater in COPD patients who experienced 2 or more exacerbations in the preceding year compared with those who did not. In addition, SAA, but not the other inflammatory biomarkers, was significantly associated with the frequent exacerbator phenotype after adjusting for potential covariates and was independent of spirometric measures of pulmonary impairment. We also found that the risk of frequent exacerbator status in the highest SAA quartile was 18 times of those in the lowest quartile, suggesting that SAA might be a good indicator of the frequent exacerbator phenotype in COPD. SAA = 131.7 ng/ml was optimal to separate infrequent from frequent exacerbators, to our knowledge using ROC analysis. Although the inflammatory biomarker SAA is known to be raised during an acute COPD exacerbation14, this is the first study to demonstrate an association of SAA with the exacerbation phenotype in stable phase of COPD. SAA may therefore be a useful biomarker to identify patients at risk of frequent exacerbations, to inform progression and to guide management of COPD.
Investigators have long sought a circulating biomarker that separates non-exacerbators from exacerbators. In the ECLIPSE trial, 2,138 COPD patients were followed for three years to determine predictors of exacerbations. None of the measured circulating biomarkers were independent predictors of exacerbation frequency. More recently, Gulati and colleagues reported a greater concentration of fibroblast growth factor 23 (FGF23) in frequent exacerbators compared with patients without frequent exacerbations21. In addition, these investigators showed that FGF23 is independently associated with frequent exacerbations, after adjusting for age, lung function, smoking and oxygen use21.
Previous studies showed that SAA is dramatically increased during acute phase of a COPD exacerbation, and that SAA is a sensitive biomarker for exacerbation severity in COPD patients14. Other studies showed increased circulating SAA concentration in COPD and other pulmonary diseases such as idiopathic pulmonary fibrosis (IPF)22–24. Formiga et al. reported that inspiratory muscle performance was considerably lower in COPD patients with greater SAA concentration23. In addition, SAA was greater in patients with IPF than in healthy controls22. Furthermore, serum levels of SAA and CRP together with other inflammatory markers (e.g. IL-6,IL-8, TNF-a, IP-10) were significantly greater in COPD patients experiencing an exacerbation than during remission and in healthy control subjects24.
Elevated SAA was reported in patients with cardiovascular disease where COPD was a comorbidity25. Since half of the deaths in COPD are associated with cardiovascular events, and the incidence of these events increases dramatically after each exacerbation26,27, and also because SAA is a good predictor of coronary artery disease as well as future cardiovascular events, SAA might be considered as a potential indicator of frequent exacerbation phenotype as well as an indicator of comorbid cardiovascular disease.
As well as acting as a biomarker for the frequent exacerbator phenotype during the stable phase of COPD, evidence suggests that SAA may mediate an acute COPD exacerbation14. SAA induces lung inflammation in COPD by promoting lung neutrophilia, which is activated by increasing expression of IL-17a in γδT cells28. In addition, SAA activates the NLRP3 inflammasome and elicits robust TLR2-, MyD88-, and IL-1-dependent pulmonary inflammation29. Consistent with this, SAA is found in the lungs and bronchoalveolar lavage fluid of smoke exposed COPD patients and endotoxin-challenged mice30,31.
In stable COPD patients we also showed that the circulating concentration of SP-D was greater in frequent exacerbators compared to infrequent exacerbators, although SP-D was not independently associated with exacerbation. Although no previous study compared the serum concentration of SP-D between frequent and infrequent exacerbators, the ECLIPSE study showed that SP-D significantly, although weakly, predicted exacerbation during the first year of follow-up17. SP-D is mainly produced by type II alveolar cells in the lung32, and is involved in regulating pulmonary surfactants, lipid homeostasis and innate immunity in order to protect the lungs from pathogens33–35. SP-D has many protective properties including anti-inflammatory and anti-oxidant functions34 that associate with protection against the development of COPD36. SP-D knockout mice have large lungs with enlarged airspaces and activated macrophages; they develop progressive pulmonary emphysema and subpleural fibrosis in association with chronic inflammation37.
A major role of SP-D is to regulate antibody-independent immune responses against invading microorganisms38. Indeed, SP-D deficient mice are more susceptible to respiratory infections with Pneumocystis carinii, influenza, respiratory syncytial virus, and bacteria39–42. This is consistent with the observation that COPD patients with serum SP-D concentration greater than the 95th percentile of nonsmokers had an increased risk of exacerbation over the following 12 months43.
Our results showed no significant correlation between serum SP-D and spirometric lung function (FEV1%pred, FVC%pred, FEV1/FVC), which is consistent with several other studies43. The interpretation of association of serum SP-D and lung function is complicated in that serum SP-D is affected not only by lung synthesis, but also by increased leakage of SP-D from the COPD lung into the systemic circulation36. Nevertheless, in our study, SP-D was not as strongly (or independently) associated with the frequent exacerbator phenotype as SAA.
We also found serum IL-4 was significantly greater in frequent exacerbators, but regression analysis showed no significant independent correlation between IL-4 and exacerbator phenotype in our stable COPD patients. IL-4 is secreted by Th-2 lymphocytes which stimulate the proliferation of B lymphocytes to produce IgG and IgE, and to mediate humoral immunization44. To our knowledge this is the first study showing greater concentration of IL-4 in frequent than infrequent exacerbators. Previous studies showed that serum IL-4 is greater in COPD patients during an acute exacerbation compared with stable COPD and control groups45. Additionally, IL-4 concentration was dramatically increased in Mycoplasma pneumoniae-induced airway diseases, including those occurring in COPD patients46. These data suggest that Th2 secreted cytokines (e.g., IL-4) might play an important role in reducing inflammation. Based on this rationale, greater IL-4 in frequent exacerbators compared with infrequent exacerbators might indicate a protective role of Th2 lymphocytes and their secreted cytokines in balancing immune system function during acute infection and exacerbation. On the other hand, increased concentration of IL-4 due to hyper-activated Th2 cells might increase the concentration of IgE immunoglobulin which leads to airway hyper-responsiveness in exacerbating COPD patients45. Despite this proposal, we did not find a correlation between IL-4 concentration and spirometric pulmonary function.
This study has limitations. We enrolled fewer frequent exacerbators than infrequent exacerbators. This might affect our ability to detect differences between the two groups. In addition, our study suffers from gender imbalance. The majority of our subjects were men (96%), limiting our ability to assert whether our findings also apply to women with COPD.