The goal of this study was to investigate local and systemic neutrophilic inflammation in patients with lung cancer and COPD considering the common inflammatory signaling pathway in both diseases. It is known that chronic inflammation plays a role in pathogenesis of both diseases there are limited integrated data comparing chronic inflammatory processes when there is coexistence of lung cancer and COPD. It is known that COPD is a major independent risk factor for lung cancer among smokers and about 50-90% of patients with lung cancer also have COPD. A mechanism, explaining why smokers with COPD have an increased risk for lung cancer when compared to smokers without COPD, is still not clear. Furthermore, it is still not known why some patients with COPD get lung cancer and some patients don't. In order to understand the inflammatory mechanisms and associations between lung cancer and COPD we aimed to analyze the patterns of local (BAL fluid) and systemic (peripheral blood) chronic neutrophilic inflammation in lung cancer and COPD.
Our results are consistent with the findings of previous reviews, showing that the count of neutrophils in peripherial blood was higher in lung cancer patients as well as in patients with COPD when compared to healthy individuals. To our knowledge such data comparing neutrophil cell count and N/L ratio in lung cancer patients, patients with COPD and both diseases in coexistence are presented for the first time. Recent studies have shown that the N/L ratio has a significant prognostic value for chronic conditions such as hypertension, diabetes mellitus, many cancers as well as lung cancer[21–24]. We demonstrated an increased neutrophil cells count and N/L ratio in peripheral blood in patients with lung cancer compared with COPD patients, but did not find significant difference between the groups of lung cancer patients with COPD and without COPD. Furthermore, N/L ratio was significantly higher in patients with COPD than in healthy subjects. The development of chronic systemic neutrophilia in cancer, as well as in COPD, has not been fully understood. There are some hypotheses explaining pathogenetic mechanisms, which are responsible for the increased neutrophil count in the peripheral blood. One of them, states that increased neutrophil count in peripheral blood in case of COPD can be influenced by the airway inflammation. This enhanced neutrophil influx to the airways may stimulate the activation of inflammatory markers, which on the other hand activate the neutrophils, cause an increase in their vitality and migration to the lung tissue due to chemoattractants. The other hypothesis, suggests that an enhanced expression of granulocyte-macrophage colony-stimulating factor and IL-6 in patients with COPD[19, 26] plays and important role, as these mediators stimulate neutrophil release from the bone marrow, causing the increased neutrophil count in the peripheral blood. The same mechanisms can be responsible for the increased numbers of neutrophils and N/L ratio in lung cancer as well. However, there is additional mechanism that could contribute to the increase in neutrophil count. Cancer cells produce granulocyte colony-stimulating factor, which directly stimulates bone marrow to release neutrophils. Because of these mechanisms, lung cancer patients have higher neutrophil count and more pronounced systemic chronic inflammation than patients with COPD.
There are limited data comparing local inflammation in lung cancer patients and COPD. Analysis of BAL fluid cells and extracellular components allows to describe immune processes in the airway microenvironment. In agreement with the data from other studies, our results revealed that percentage and cell count of neutrophils were significantly higher in lung cancer patients[28, 29] and COPD patients compared with healthy subjects. In contrast to neutrophilia in peripheral blood, percentage of neutrophils in BAL fluid was significantly lower and percentage of macrophages was significantly higher in lung cancer patients compared with COPD patients. Furthermore, we observed that in lung cancer patients, when compared with COPD patients, not only the count of macrophages in BAL fluid was increased, percentage and cell count of monocytes in peripheral blood were also higher. Lung cancer cells generate chemotactic factors and chemokine ligand for circulating monocytes, which are recruited into the lungs, where they differentiate into the macrophages. It is known that immune cells, including macrophages, infiltrate the tumor stroma[2, 31]. It is well know that monocyte-lineage cells including macrophages also play an important role in the pathogenesis of COPD. A plausible explanation for the increased count of macrophages in BAL fluid of patients with lung cancer is that tumor promotes migration of monocytes and their differentiation into the macrophages. This confirms the fact, that macrophages play central role in the immune response to lung cancer.
We observed that mean serum and BAL fluid levels of NE and MPO were significantly higher in patients with lung cancer (with or without COPD) compared with COPD patients and healthy subjects. Neutrophil elastase and MPO are important markers of neutrophil activation and neutrophilic inflammation[9, 10]. Some studies have identified increased NE and MPO levels in patients with COPD and healthy smokers. The role of NE and MPO in chronic lung inflammation and association with lung cancer development has been characterized in cell line and animal models[11–13, 32]. Nevertheless, there are no clear data comparing the importance of NE and MPO in lung cancer and COPD. Although we demonstrated that neutrophil cell count and percentage in BAL fluid were lower in lung cancer groups compared with COPD patients, NE and MPO levels in BAL fluid were significantly higher in lung cancer groups than in COPD group. Thus, our findings let us hypothesize that in patients with lung cancer neutrophils appear in a more activated state through the production of NE, MPO than in patients with COPD. Furthermore, there are some data showing that NE and MPO is expressed not only in neutrophils but also in monocyte-derived macrophages.
It is generally accepted that oxidative stress, which is promoted by cigarette smoke, may be involved in cancer development[2, 5]. The literature is full of data on the increased oxidative stress in patients with COPD[3, 7]. However it is still little known about ROS production in lung cancer patients with COPD and data comparing lung cancer with COPD are lacking. In the present study we demonstrated that spontaneous ROS production in peripheral blood neutrophils was higher in both lung cancer groups (lung cancer and lung cancer with COPD) than in patients with COPD, but there were no differences between lung cancer patients with COPD or without COPD. Heijink and colleagues suggested that chemical factor like cigarette smoke may influence a more intensive ROS production in COPD and healthy individuals. As a chemical factor which causes neutrophil activation we used a PMA. Our study showed that PMA induced intensive ROS production not only in patients with COPD and healthy individuals but in lung cancer patient groups (with and without COPD) as well. Our results revealed that activated neutrophils after stimulation with PMA produced more ROS in lung cancer patients (with and without COPD) compared with COPD patients. It means that chemical factors cause enhanced inflammation in lung cancer. In addition, we provided evidence that intracellular ROS was increased in peripheral blood neutrophils of lung cancer patients which positively correlated with levels of both inflammatory markers (NE, MPO). Such correlation supports the hypothesis that intensive chronic neutrophilic inflammation promotes more intensive ROS production in lung cancer compared to COPD.
We also investigated levels of NE, MPO and ROS production in the patients with different stage of lung cancer (early vs. advanced). Our data demonstrated that patients with advanced lung cancer had significantly higher serum NE, MPO levels and more intensive ROS production in peripheral neutrophils than patients with early stage lung cancer. These results are consistent with the results reported in other studies, presenting additional evidence of the importance of chronic neutrophilic inflammation in lung cancer progression. Furthermore, our findings of higher systemical NE, MPO and ROS production in patients with early lung cancer compared with COPD patients further support the fact, that chronic inflammation can be more pronounced in lung cancer than COPD.
Additional studies have demonstrated that smoking stimulates not only local but also systemic inflammation[6, 16]. However, the data on the influence of smoking itself on chronic neutrophilic inflammation in lung cancer patients with coexisting COPD are scarce. The data are contradictory, some researches have shown that cigarette smoke is strongly associated with the increased inflammation of airways, proved by the exhaled breath condensate in lung cancer patients. Other study failed to prove these associations in lung cancer patients. There is evidence that cigarette smoking plays an important role as the starting point of chronic inflammation but has little influence in promoting inflammation in lung cancer. However, our results indicate that in patients with lung cancer local and systemic inflammation was increased independent of smoking and not only cigarette smoking but also other factors play a significant role in promotion of chronic inflammation in lung cancer.