SUBPOPULATIONS OF ALVEOLAR MACROPHAGES IN SMOKERS AND NONSMOKERS - RELATION TO THE EXPRESSION OF CD11/CD18 MOLECULES AND SUPEROXIDE ANION PRODUCTION

We were previously able to show that the number of alveolar macrophages (AM) expressing CD11/CD18 molecules is increased in smokers compared with nonsmokers and related to the superoxide anion (O-2(-)) production of these cells. Since it has been demonstrated that AM are a heterogeneous cell population that can be separated by density, we performed this study to investigate the expression of CD11/CD18 molecules and O-2(-) production in relation to cell density of AM from smokers and nonsmokers. AM were obtained by bronchoalveolar lavage (BAL) from smokers (n = 32) and nonsmokers (n = 20). Subpopulations were isolated using discontinuous Percoll(TM) density-gradient centrifugation with four densities (fraction 1: 1.030; fraction 2: 1.040; fraction 3: 1.050; and fraction 4: 1.070 g/ml). Expression of CD11/CD18 on freshly isolated cells and on AM before and after density centrifugation was studied using peroxidase-antiperoxidase staining. The contribution of AM subpopulations to O-2(-) production in smokers was determined by monitoring the reduction of ferricytochrome C to ferrocytochrome C. We obtained 0.99 +/- 0.1 x 10(5) AM/ml BAL in nonsmokers and 2.4 +/- 0.3 x 10(5) AM/ml in smokers. Recovery after density centrifugation was greater than or equal to 72%. The absolute number of AM in smokers was significantly increased in fractions 3 and 4 (median 4.37 x 10(6) and 2.05 x 10(6), respectively) compared with nonsmokers (median 1.26 x 10(6) and 0.7 x 10(6), respectively) (p < 0.05). In both smokers and nonsmokers, fractions 3 acid 4 showed a comparable increase in the percentage of CD11/CD18-positive AM compared with fractions 1 and 2. The absolute increase in CD11/CD18-positive AM in smokers was mainly due to the increased absolute number of positive AM with higher density from fractions 3 and 4 compared with nonsmokers (p < 0.05 to < 0.01). Analysis of O-2(-) production by smoker AM with lower and higher density showed that AM with higher density (> 1.040 g/ml) produced significantly more O-2(-) (median 26.3 nmol/10(6) cells/120 min) than AM with lower density (less than or equal to 1.040 g/ml) (median 12.7 nmol/10(6) cells/120 min) (p < 0.05). Our data indicate that AM with higher density are increased in smokers compared with nonsmokers and are responsible for the increased number of CD11/CD18-positive AM in smokers. These high-density AM of smokers are mainly responsible for the O-2(-) production of these cells.