title: CD36 mediates H2O2-induced calcium influx in lung microvascular endothelial cells.
abstract: Elevated levels of reactive oxygen species and intracellular Ca2+play a key role in endothelial barrier dysfunction in acute lung injury. We previously showed that H2O2-induced increases in intracellular calcium concentrations ([Ca2+]i) in lung microvascular endothelial cells (LMVECs) involve the membrane Ca2+channel, transient receptor potential vanilloid-4 (TRPV4) and that inhibiting this channel attenuated H2O2-induced barrier disruption in vitro. We also showed that phosphorylation of TRPV4 by the Src family kinase, Fyn, contributes to H2O2-induced Ca2+influx in LMVEC. In endothelial cells, Fyn is tethered to the cell membrane by CD36, a fatty acid transporter. In this study, we assessed the effect of genetic loss or pharmacological inhibition of CD36 on Ca2+responses to H2O2H2O2-induced Ca2+influx was attenuated in LMVEC isolated from mice lacking CD36 (CD36-/-). TRPV4 expression and function was unchanged in LMVEC isolated from wild-type (WT) and CD36-/-mice, as well as mice with deficiency for Fyn (Fyn-/-). TRPV4 immunoprecipitated with Fyn, but this interaction was decreased in CD36-/-LMVEC. The amount of phosphorylated TRPV4 was decreased in LMVEC from CD36-/-mice compared with WT controls. Loss of CD36 altered subcellular localization of Fyn, while inhibition of CD36 fatty acid transport with succinimidyl oleate did not attenuate H2O2-induced Ca2+influx. Lastly, we found that CD36-/-mice were protected from ischemia-reperfusion injury in vivo. In conclusion, our data suggest that CD36 plays an important role in H2O2-mediated lung injury and that the mechanism may involve CD36-dependent scaffolding of Fyn to the cell membrane to facilitate TRPV4 phosphorylation.
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland; ksuresh2@jhmi.edu.
authors.forename: Karthik
authors.initials: K
authors.lastname: Suresh
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: Laura
authors.initials: L
authors.lastname: Servinsky
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: Jose
authors.initials: J
authors.lastname: Reyes
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: Clark
authors.initials: C
authors.lastname: Undem
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: Joel
authors.initials: J
authors.lastname: Zaldumbide
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: Otgonchimeg
authors.initials: O
authors.lastname: Rentsendorj
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: Sruti
authors.initials: S
authors.lastname: Modekurty
authors.affiliation: Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland; and.
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Dodd-O
authors.affiliation: Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland.
authors.forename: Alan
authors.initials: A
authors.lastname: Scott
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: David B
authors.initials: DB
authors.lastname: Pearse
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.
authors.forename: Larissa A
authors.initials: LA
authors.lastname: Shimoda
mesh_terms: D000818:Animals; D018955:CD36 Antigens / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism; D017353:Gene Deletion; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D008077:Lipoproteins, LDL / Q000494:pharmacology; D008168:Lung / Q000098:blood supply*; D008810:Mice, Inbred C57BL; D055806:Microvessels / Q000166:cytology*; D019301:Oleic Acid / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051076:Proto-Oncogene Proteins c-fyn / Q000378:metabolism; D015427:Reperfusion Injury / Q000378:metabolism / Q000473:pathology; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism; D050916:TRPV Cation Channels / Q000378:metabolism
chemical_list: D018955:CD36 Antigens; D005227:Fatty Acids; D008077:Lipoproteins, LDL; D050916:TRPV Cation Channels; C486576:Trpv4 protein, mouse; C032629:acetyl-LDL; D019301:Oleic Acid; D006861:Hydrogen Peroxide; C494960:Fyn protein, mouse; D051076:Proto-Oncogene Proteins c-fyn; D002118:Calcium
keywords: ROS; acute lung injury; calcium