title: Human immunodeficiency virus type 1 enters brain microvascular endothelia by macropinocytosis dependent on lipid rafts and the mitogen-activated protein kinase signaling pathway.
abstract: Brain microvascular endothelial cells (BMVECs) present an incomplete barrier to human immunodeficiency virus type 1 (HIV-1) neuroinvasion. In order to clarify the mechanisms of HIV-1 invasion, we have examined HIV-1 uptake and transcellular penetration in an in vitro BMVEC model. No evidence of productive infection was observed by luciferase, PCR, and reverse transcriptase assays. Approximately 1% of viral RNA and 1% of infectious virus penetrated the BMVEC barrier without disruption of tight junctions. The virus upregulated ICAM-1 on plasma membranes and in cytoplasmic vesiculotubular structures. HIV-1 virions were entangled by microvilli and were taken into cytoplasmic vesicles through surface invaginations without fusion of the virus envelope with the plasma membrane. Subsequently, the cytoplasmic vesicles fused with lysosomes, the virions were lysed, and the vesicles diminished in size. Upon cell entry, HIV-1 colocalized with cholera toxin B, which targets lipid raft-associated GM1 ganglioside. Cholesterol-extracting agents, cyclodextrin and nystatin, and polyanion heparin significantly inhibited virus entry. Anti-CD4 had no effect and the chemokine AOP-RANTES had only a slight inhibitory effect on virus entry. HIV-1 activated the mitogen-activated protein kinase (MAPK) pathway, and inhibition of MAPK/Erk kinase inhibited virus entry. Entry was also blocked by dimethylamiloride, indicating that HIV-1 enters endothelial cells by macropinocytosis. Therefore, HIV-1 penetrates BMVECs in ICAM-1-lined macropinosomes by a mechanism involving lipid rafts, MAPK signaling, and glycosylaminoglycans, while CD4 and chemokine receptors play limited roles in this process.
authors.affiliation: Department of Medicine, Greater Los Angeles VA Medical Center, 675 Young Drive South, Los Angeles, CA 90073, USA.
authors.forename: Nancy Q
authors.initials: NQ
authors.lastname: Liu
authors.forename: Albert S
authors.initials: AS
authors.lastname: Lossinsky
authors.forename: Waldemar
authors.initials: W
authors.lastname: Popik
authors.forename: Xia
authors.initials: X
authors.lastname: Li
authors.forename: Chandrasekhar
authors.initials: C
authors.lastname: Gujuluva
authors.forename: Benjamin
authors.initials: B
authors.lastname: Kriederman
authors.forename: Jaclyn
authors.initials: J
authors.lastname: Roberts
authors.forename: Tatania
authors.initials: T
authors.lastname: Pushkarsky
authors.forename: Michael
authors.initials: M
authors.lastname: Bukrinsky
authors.forename: Marlys
authors.initials: M
authors.lastname: Witte
authors.forename: Martin
authors.initials: M
authors.lastname: Weinand
authors.forename: Milan
authors.initials: M
authors.lastname: Fiala
mesh_terms: D001812:Blood-Brain Barrier; D001921:Brain / Q000098:blood supply*; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000821:virology*; D015658:HIV Infections / Q000821:virology; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D020935:MAP Kinase Signaling System; D021962:Membrane Microdomains / Q000378:metabolism*; D008833:Microcirculation; D018613:Microscopy, Confocal; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008954:Models, Biological; D010873:Pinocytosis; D015398:Signal Transduction
chemical_list: D020928:Mitogen-Activated Protein Kinases