aeruginosa. Two genes, SP-A1 and SP-A2, encode human SP-A. We hypothesized that genetically determined differences in the activity of SP-A1 and SP-A2 gene products exist. To test this, we studied association of a nonmucoid P. aeruginosa strain (ATCC 39018) with rat alveolar macrophages in the presence or absence of insect cellexpressed human SP-A variants. We used two trios, each consisting of SP-A1, SP-A2, and their coexpressed SP-A1/SP-A2 variants. We tested the 6A 2 and 6A 4 alleles (for SP-A1), the 1A 0 and 1A alleles (for SP-A2), and their respective coexpressed SP-A1/SP-A2 gene products. After incubation of alveolar macrophages with P. aeruginosa in the presence of the SP-A variants at 37°C for 1 h, the cell association of bacteria was assessed by light microscopy analysis. We found 1) depending on SP-A concentration and variant, SP-A2 variants significantly increased the cell association more than the SP-A1 variants (the phagocytic index for SP-A1 was ϳ52-95% of the SP-A2 activity); 2) coexpressed variants at certain concentrations were more active than single gene products; and 3) the phagocytic index for SP-A variants was ϳ18 -41% of the human SP-A from bronchoalveolar lavage. We conclude that human SP-A variants in vitro enhance association of P. aeruginosa with rat alveolar macrophages differentially and in a concentration-dependent manner, with SP-A2 variants having a higher activity compared with SP-A1 variants.
Surfactant protein A (SP-A) enhances phagocytosis of Pseudomonas aeruginosa. SP-A1 and SP-A2 encode human (h) SP-A; SP-A2 products enhance phagocytosis more than SP-A1. Oxidation can affect SP-A function. We hypothesized that in vivo and in vitro ozone-induced oxidation of SP-A (as assessed by its carbonylation level) negatively affects its function in phagocytosis (as assessed by bacteria cell association). To test this, we used P. aeruginosa, rat alveolar macrophages (AMs), hSP-As with varying levels of in vivo (natural) oxidation, and ozone-exposed SP-A2 (1A, 1A 0 ) and SP-A1 (6A 2 , 6A 4 ) variants. SP-A oxidation levels (carbonylation) were measured; AMs were incubated with bacteria in the presence of SP-A, and the phagocytic index was calculated. We found: 1) the phagocytic activity of hSP-A is reduced with increasing levels of in vivo SP-A carbonylation; 2) in vitro ozone exposure of hSP-A decreases its function in a dose-dependent manner as well as its ability to enhance phagocytosis of either gram-negative or gram-positive bacteria; 3) the activity of both SP-A1 and SP-A2 decreases in response to in vitro ozone exposure of proteins with SP-A2 being affected more than SP-A1. We conclude that both in vivo and in vitro oxidative modifications of SP-A by carbonylation reduce its ability to enhance phagocytosis of bacteria and that the activity of SP-A2 is affected more by in vitro ozone-induced oxidation. We speculate that functional differences between SP-A1 and SP-A2 exist in vivo and that the redox status of the lung microenvironment differentially affects function of SP-A1 and SP-
Surfactant protein A (SP-A) plays a role in innate host defense. Human SP-A is encoded by two functional genes (SP-A1 and SP-A2), and several alleles have been characterized for each gene. We assessed the effect of in vitro expressed human SP-A genetic variants, on TNF-α and IL-8 production by THP-1 cells in the presence of bleomycin, either before or after ozone-induced oxidation of the variants. The oligomerization of SP-A variants was also examined. We found 1) cytokine levels induced by SP-A2 (1A, 1A0) were significantly higher than those by SP-A1 (6A2, 6A4) in the presence of bleomycin. 2) In the presence of bleomycin, ozone-induced oxidation significantly decreased the ability of 1A and 1A/6A4, but not of 6A4, to stimulate TNF-α production. 3) The synergistic effect of bleomycin/SP-A, either before or after oxidation, can be inhibited to the level of bleomycin alone by surfactant lipids. 4) Differences in oligomerization were also observed between SP-A1 and SP-A2. The results indicate that differences among SP-A variants may partly explain the individual variability of pulmonary complications observed during bleomycin chemotherapy and/or in an environment that may promote protein oxidation.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.