title: Surfactant protein A2 (SP-A2) variants expressed in CHO cells stimulate phagocytosis of Pseudomonas aeruginosa more than do SP-A1 variants.
abstract: Surfactant protein A (SP-A) enhances phagocytosis of Pseudomonas aeruginosa. Two functional genes, SP-A1 and SP-A2, encode human SP-A. As we showed before, baculovirus-mediated insect cell-expressed SP-A2 enhances the association of P. aeruginosa with rat alveolar macrophages (rAMs) more than does SP-A1. However, true phagocytosis (internalization) was not shown, and insect cell derived proteins lack or are defective in certain mammalian posttranslational modifications that may be important for SP-A1 and SP-A2 activity and specificity. Here we used SP-A1 (6A(2), 6A(4)) and SP-A2 (1A(0), 1A(1)) allele variants expressed by CHO (Chinese hamster ovary) mammalian cells to study their effect on association and/or internalization of P. aeruginosa by rAMs and/or human AMs (hAMs) and to study if phagocytosis can be modulated differentially and/or more effectively by CHO cell-expressed SP-A variants than by insect-cell expressed SP-A variants. For cell association and internalization assessments, light microscopy and fluorescence-activated cell sorter analyses were used, respectively. We found the following for the first time. (i) SP-A2 variants enhanced phagocytosis (cell association and/or internalization) of P. aeruginosa more than SP-A1 variants did, and the cell association correlated with internalization. (ii) Differences in the activities of SP-A variants were observed in the following order: 1A(1)>1A(0)>6A(2)>6A(4). (iii) rAMs, although more active than hAMs, are an appropriate model, as SP-A2 variants exhibited activity higher than that seen for SP-A1 variants with either rAMs or hAMs. (iv) CHO cell-expressed SP-A was considerably more active than insect cell-expressed variants. We conclude that SP-A2 variants stimulate phagocytosis of P. aeruginosa more effectively than SP-A1 variants and that posttranslational modifications positively influence the phagocytic activity of SP-A.
authors.affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, 500 University Drive, Hershey, PA 17033, USA.
authors.forename: Anatoly N
authors.initials: AN
authors.lastname: Mikerov
authors.forename: Guirong
authors.initials: G
authors.lastname: Wang
authors.forename: Todd M
authors.initials: TM
authors.lastname: Umstead
authors.forename: Mario
authors.initials: M
authors.lastname: Zacharatos
authors.forename: Neal J
authors.initials: NJ
authors.lastname: Thomas
authors.forename: David S
authors.initials: DS
authors.lastname: Phelps
authors.forename: Joanna
authors.initials: J
authors.lastname: Floros
mesh_terms: D000818:Animals; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D016676:Macrophages, Alveolar / Q000276:immunology / Q000378:metabolism; D008297:Male; D010587:Phagocytosis / Q000235:genetics / Q000276:immunology; D011499:Protein Processing, Post-Translational; D011550:Pseudomonas aeruginosa / Q000276:immunology; D037662:Pulmonary Surfactant-Associated Protein A / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D037662:Pulmonary Surfactant-Associated Protein A; C445842:SFTPA1 protein, human; C445843:SFTPA2 protein, human