title: Glutaminase-containing microvesicles from HIV-1-infected macrophages and immune-activated microglia induce neurotoxicity.
abstract: BACKGROUND
  HIV-1-infected and/or immune-activated microglia and macrophages are pivotal in the pathogenesis of HIV-1-associated neurocognitive disorders (HAND). Glutaminase, a metabolic enzyme that facilitates glutamate generation, is upregulated and may play a pathogenic role in HAND. Our previous studies have demonstrated that glutaminase is released to the extracellular fluid during HIV-1 infection and neuroinflammation. However, key molecular mechanisms that regulate glutaminase release remain unknown. Recent advances in understanding intercellular trafficking have identified microvesicles (MVs) as a novel means of shedding cellular contents. We posit that during HIV-1 infection and immune activation, microvesicles may mediate glutaminase release, generating excessive and neurotoxic levels of glutamate.
  
  
  RESULTS
  MVs isolated through differential centrifugation from cell-free supernatants of monocyte-derived macrophages (MDM) and BV2 microglia cell lines were first confirmed in electron microscopy and immunoblotting. As expected, we found elevated number of MVs, glutaminase immunoreactivities, as well as glutaminase enzyme activity in the supernatants of HIV-1 infected MDM and lipopolysaccharide (LPS)-activated microglia when compared with controls. The elevated glutaminase was blocked by GW4869, a neutral sphingomyelinase inhibitor known to inhibit MVs release, suggesting a critical role of MVs in mediating glutaminase release. More importantly, MVs from HIV-1-infected MDM and LPS-activated microglia induced significant neuronal injury in rat cortical neuron cultures. The MV neurotoxicity was blocked by a glutaminase inhibitor or GW4869, suggesting that the neurotoxic potential of HIV-1-infected MDM and LPS-activated microglia is dependent on the glutaminase-containing MVs.
  
  
  CONCLUSIONS
  These findings support MVs as a potential pathway/mechanism of excessive glutamate generation and neurotoxicity in HAND and therefore MVs may serve as a novel therapeutic target.
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA. beiqing.wu2@unmc.edu.
authors.forename: Beiqing
authors.initials: B
authors.lastname: Wu
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA. yhuan1@unmc.edu.|Center for Translational Neurodegeneration and Regenerative Therapy, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, 200025, China. yhuan1@unmc.edu.
authors.forename: Yunlong
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA. alexander.braun@unmc.edu.
authors.forename: Alexander L
authors.initials: AL
authors.lastname: Braun
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA. zenghan.tong@unmc.edu.
authors.forename: Zenghan
authors.initials: Z
authors.lastname: Tong
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA. runze.zhao@unmc.edu.
authors.forename: Runze
authors.initials: R
authors.lastname: Zhao
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA.|Center for Translational Neurodegeneration and Regenerative Therapy, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, 200025, China.
authors.forename: Yuju
authors.initials: Y
authors.lastname: Li
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA.
authors.forename: Fang
authors.initials: F
authors.lastname: Liu
authors.affiliation: Laboratory of Neuroimmunology and Regenerative Therapy, Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA. jzheng@unmc.edu.|Departments of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, 68198-5930, USA. jzheng@unmc.edu.|Center for Translational Neurodegeneration and Regenerative Therapy, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, 200025, China. jzheng@unmc.edu.
authors.forename: Jialin C
authors.initials: JC
authors.lastname: Zheng
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D005972:Glutaminase / Q000378:metabolism*; D015497:HIV-1; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000378:metabolism* / Q000821:virology*; D017628:Microglia / Q000276:immunology / Q000378:metabolism / Q000821:virology*; D009474:Neurons / Q000378:metabolism / Q000821:virology*; D017207:Rats, Sprague-Dawley
chemical_list: D008070:Lipopolysaccharides; D005972:Glutaminase