title: Cytoplasmic HIV-RNA in monocytes determines microglial activation and neuronal cell death in HIV-associated neurodegeneration.
abstract: Despite highly active antiretroviral therapy, HIV-associated neurocognitive disorders (HAND) are still highly prevalent. Direct neurotoxicity of microglia activated by HIV-infected monocytes independent from viral replication may account for this observation. To investigate underlying molecular and viral determinants, human monocytoid cells (U937) transduced with HIV-particles were co-cultured with primary human microglia or astrocytes. Using genetically-engineered HIV-particles key steps of infection were examined. Levels of pro-inflammatory/neurotoxic cytokines were investigated in co-culture supernatants by flow cytometry. Neurotoxicity mediated by the supernatants was analysed using primary cortical rat neurons. To corroborate our findings, cytokine profiles in cerebrospinal fluid (CSF) of neuropsychologically asymptomatic HIV positive (HIV(+)) patients (n=45) were correlated with neurofilament H (NfH) as surrogate of neuronal/axonal degeneration. In contrast to direct exposure of HIV to microglia, only the presence of HIV-transduced monocytoid cells strongly activated human microglia as evidenced by enhanced secretion of CXCL10, CCL5, CCL2, and IL-6 (1.3-7.1-fold; p<0.01) leading to two-fold increased neurotoxicity (p<0.001). In direct comparison, astrocyte activation by HIV-transduced monocytoid cells was limited. Using different mutant HIV-particles we show that the presence of cytoplasmic HIV-RNA in monocytoid cells is the viral determinant for this unique microglial activation pattern and subsequent neuronal cell death; reverse transcription and expression of viral genes were not essential. In CSF of presymptomatic HIV(+) patients, CXCL10, CCL5 and IL-6 were correlated with NfH as surrogate marker of neurodegeneration as well as CSF-pleocytosis. In conclusion, cytosolic viral RNA in monocytes is mandatory for subsequent microglial activation and neurotoxicity; activated astrocytes may augment neuroinflammation. In addition, neuroinflammation and neurodegeneration occur even in preclinical HIV(+) patients and are associated with cytokines regulated in vitro. Our data may aid in the development of biomarkers and glia-directed therapeutic approaches of HAND.
authors.affiliation: Department of Neurology, St. Josef-Hospital, Ruhr-University Bochum, Gudrunstr. 56, 44791 Bochum, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: Faissner
authors.affiliation: Department of Neurology, St. Josef-Hospital, Ruhr-University Bochum, Gudrunstr. 56, 44791 Bochum, Germany.
authors.forename: Björn
authors.initials: B
authors.lastname: Ambrosius
authors.affiliation: Department of Neurology, St. Josef-Hospital, Ruhr-University Bochum, Gudrunstr. 56, 44791 Bochum, Germany.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Schanzmann
authors.affiliation: Department of Molecular and Medical Virology, Ruhr-University Bochum, Universitätsstr. 150, 44801 Bochum, Germany.
authors.forename: Bastian
authors.initials: B
authors.lastname: Grewe
authors.affiliation: Department of Dermatology, St. Josef-Hospital, Ruhr-University Bochum, Gudrunstr. 56, 44791 Bochum, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Potthoff
authors.affiliation: Institute of Molecular Virology, Ulm University, Meyerhofstr. 1, 89081 Ulm, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Münch
authors.affiliation: Department of Neurosurgery, University Duisburg-Essen, Hufelandstr. 55, 45147 Essen, Germany.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Sure
authors.affiliation: Institute of Clinical and Molecular Virology, University Erlangen, Schloßgarten 4, 91054 Erlangen, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Gramberg
authors.affiliation: Institute of Clinical and Molecular Virology, University Erlangen, Schloßgarten 4, 91054 Erlangen, Germany.
authors.forename: Sabine
authors.initials: S
authors.lastname: Wittmann
authors.affiliation: Department of Dermatology, St. Josef-Hospital, Ruhr-University Bochum, Gudrunstr. 56, 44791 Bochum, Germany.
authors.forename: Norbert
authors.initials: N
authors.lastname: Brockmeyer
authors.affiliation: Department of Molecular and Medical Virology, Ruhr-University Bochum, Universitätsstr. 150, 44801 Bochum, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Uberla
authors.affiliation: Department of Neurology, St. Josef-Hospital, Ruhr-University Bochum, Gudrunstr. 56, 44791 Bochum, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Gold
authors.affiliation: Department of Molecular and Medical Virology, Ruhr-University Bochum, Universitätsstr. 150, 44801 Bochum, Germany; Fraunhofer Institute for Cell Therapy and Immunology, Perlickstr. 1, 04103 Leipzig, Germany. Electronic address: thomas.grunwald@izi.fraunhofer.de.
authors.forename: Thomas
authors.initials: T
authors.lastname: Grunwald
authors.affiliation: Department of Neurology, St. Josef-Hospital, Ruhr-University Bochum, Gudrunstr. 56, 44791 Bochum, Germany. Electronic address: andrew.chan@rub.de.
authors.forename: Andrew
authors.initials: A
authors.lastname: Chan
mesh_terms: D000818:Animals; D002453:Cell Cycle / Q000502:physiology; D016923:Cell Death / Q000502:physiology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D002555:Cerebrospinal Fluid / Q000737:chemistry / Q000821:virology; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D004622:Embryo, Mammalian; D005333:Fetus / Q000166:cytology; D006678:HIV / Q000235:genetics*; D016655:HIV Core Protein p24 / Q000235:genetics / Q000378:metabolism; D015658:HIV Infections / Q000134:cerebrospinal fluid; D006801:Humans; D017628:Microglia / Q000166:cytology / Q000502:physiology*; D009000:Monocytes / Q000166:cytology / Q000378:metabolism* / Q000821:virology*; D009474:Neurons / Q000502:physiology / Q000821:virology; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism*; D051381:Rats
chemical_list: D016207:Cytokines; D016655:HIV Core Protein p24; D012367:RNA, Viral
keywords: Cytokines; Glial activation; HIV RNA; HIV related neurological diseases; Microglia; Neurodegeneration; Neuroinflammation