title: Circulating complexes of the vitamin D binding protein with G-actin induce lung inflammation by targeting endothelial cells.
abstract: This study investigated the actin scavenger function of the vitamin D binding protein (DBP) in vivo using DBP null (-/-) mice. Intravenous injection of G-actin into wild-type (DBP+/+) and DBP-/- mice showed that contrary to expectations, DBP+/+ mice developed more severe acute lung inflammation. Inflammation was restricted to the lung and pathological changes were clearly evident at 1.5 and 4h post-injection but were largely resolved by 24h. Histology of DBP+/+ lungs revealed noticeably more vascular leakage, hemorrhage and thickening of the alveolar wall. Flow cytometry analysis of whole lung homogenates showed significantly increased neutrophil infiltration into DBP+/+ mouse lungs at 1.5 and 4h. Increased amounts of protein and leukocytes were also noted in bronchoalveolar lavage fluid from DBP+/+ mice 4h after actin injection. In vitro, purified DBP-actin complexes did not activate complement or neutrophils but induced injury and death of cultured human lung microvascular endothelial cells (HLMVEC) and human umbilical vein endothelial cells (HUVEC). Cells treated with DBP-actin showed a significant reduction in viability at 4h, this effect was reversible if cells were cultured in fresh media for another 24h. However, a 24-h treatment with DBP-actin complexes showed a significant increase in cell death (95% for HLMVEC, 45% for HUVEC). The mechanism of endothelial cell death was via both caspase-3 dependent (HUVEC) and independent (HLMVEC) pathways. These results demonstrate that elevated levels and/or prolonged exposure to DBP-actin complexes may induce endothelial cell injury and death, particularly in the lung microvasculature.
authors.affiliation: Department of Pathology, Stony Brook University School of Medicine, Stony Brook, NY 11794-8691, USA.
authors.forename: Lingyin
authors.initials: L
authors.lastname: Ge
authors.affiliation: Department of Pathology, Stony Brook University School of Medicine, Stony Brook, NY 11794-8691, USA.
authors.forename: Glenda
authors.initials: G
authors.lastname: Trujillo
authors.affiliation: Center for Heart and Lung Research, The Feinstein Institute for Medical Research, Manhasset, NY 11030-3816, USA.
authors.forename: Edmund J
authors.initials: EJ
authors.lastname: Miller
authors.affiliation: Department of Pathology, Stony Brook University School of Medicine, Stony Brook, NY 11794-8691, USA. Electronic address: richard.kew@stonybrook.edu.
authors.forename: Richard R
authors.initials: RR
authors.lastname: Kew
mesh_terms: D000199:Actins / Q000008:administration & dosage / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002199:Capillary Permeability / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D004730:Endothelium, Vascular / Q000187:drug effects / Q000473:pathology*; D008168:Lung / Q000187:drug effects / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D046912:Multiprotein Complexes / Q000378:metabolism* / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000276:immunology*; D011014:Pneumonia / Q000139:chemically induced / Q000276:immunology*; D011485:Protein Binding; D014809:Vitamin D-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; D046912:Multiprotein Complexes; D014809:Vitamin D-Binding Protein; D053148:Caspase 3
keywords: 2′,7′-dichlorofluorescein diacetate; AO/EB; Actin; BAL; CTB; DBP; DCFDA; EA; EASS; EGM-2; Endothelial cells; HLMVEC; HUVEC; Inflammation; PARP; ROS; Tissue injury; Vitamin D binding protein; acridine orange/ethidium bromide; antibody-coated sheep erythrocytes; bronchoalveolar lavage; cell-titer blue viability assay; endothelial growth medium-2; extracellular actin scavenger system; human lung microvascular endothelial cell; human umbilical vein endothelial cell; poly ADP ribose polymerase; reactive oxygen species; vitamin D binding protein