Oncostatin M promotes excitotoxicity by inhibiting glutamate uptake in astrocytes: implications in HIV-associated neurotoxicity.
Elevated levels of oncostatin M (OSM), an interleukin-6 cytokine family member, have been observed in HIV-1-associated neurocognitive disorders (HAND) and Alzheimer's disease. However, the function of OSM in these disease conditions is unclear. Since deficient glutamate uptake by astrocytes is instrumental in HAND-associated neurotoxicity, we hypothesized that OSM impairs glutamate uptake in astrocytes and thereby promotes neuronal excitotoxicity.



Primary cultures of mouse cortical astrocytes, neurons, microglia, and BV2 cell line were used. The expression of glutamate transporters (GLAST/EAAT1 and GLT-1/EAAT2) was investigated using real-time PCR and Western blot, and their activity was assessed by measuring (3)H-D-aspartate uptake. Neuronal toxicity was measured using the colorimetric MTT (3-(4,5-dimethylthiazol-2-yl-) 2,5-diphenyltetrazolium bromide) assay and immunocytochemistry. A chimeric HIV-1 that infects murine cells (EcoHIV/NL4-3-GFP virus (EcoHIV)) was used to investigate whether the virus induces OSM, OSM receptor (OSMR)-β, glycoprotein 130 (gp130), GLT-1, GLAST (mRNA and protein), and OSM release (ELISA) in cultured BV2 cells, primary microglia, or astrocytes. Statistical analyses of the data were performed using one-way ANOVA (to allow multiple comparisons) and two-tailed Student's t test.



OSM treatment (10 ng/mL) time-dependently reduced GLAST and GLT-1 expression and inhibited (3)H-D-aspartate uptake in cultured astrocytes in a concentration-dependent manner, an effect prevented by the Janus kinase (JAK)/signal transducers and activators of transcription (STAT)3 inhibitor AG490. Down-regulation of astrocytic glutamate transport by OSM resulted in NMDA receptor-dependent excitotoxicity in cortical neurons. Infection with EcoHIV induced OSM gene expression and protein release in BV2 cells and microglia, but not in astrocytes. Conversely, EcoHIV caused a fivefold increase in OSMR-β mRNA (but not gp130) and protein in astrocytes, but not in microglia, which did not express OSMR-β protein. Finally, astrocytic expression of GLAST gene was unaffected by EcoHIV, whereas GLT-1 mRNA was increased by twofold.



We provide first evidence that activation of JAK/STAT3 signaling by OSM inhibits glutamate uptake in astrocytes, which results in neuronal excitotoxicity. Our findings with EcoHIV suggest that targeting OSMR-β signaling in astrocytes might alleviate HIV-1-associated excitotoxicity.
Department of Surgery, Division of Basic and Translational Research, University of Minnesota, Minneapolis, MN, USA.
Shamsudheen
S
Moidunny
Center for Neuroscience of Coimbra, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
Marco
M
Matos
Department of Neuroscience, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
Evelyn
E
Wesseling
Department of Surgery, Division of Basic and Translational Research, University of Minnesota, Minneapolis, MN, USA.
Santanu
S
Banerjee
Molecular Virology Division, St. Luke's-Roosevelt Hospital Center, New York, USA.
David J
DJ
Volsky
Center for Neuroscience of Coimbra, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
Rodrigo A
RA
Cunha
Center for Neuroscience of Coimbra, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
Paula
P
Agostinho
Department of Neuroscience, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
Hendrikus W
HW
Boddeke
Department of Surgery, Division of Basic and Translational Research, University of Minnesota, Minneapolis, MN, USA. royxx002@umn.edu.|Department of Pharmacology, University of Minnesota, Minneapolis, MN, USA. royxx002@umn.edu.
Sabita
S
Roy
D027322:Amino Acid Transport System X-AG / Q000378:metabolism; D000818:Animals; D000970:Antineoplastic Agents / Q000009:adverse effects* / Q000494:pharmacology / Q000633:toxicity; D001224:Aspartic Acid / Q000378:metabolism; D001253:Astrocytes / Q000187:drug effects* / Q000378:metabolism* / Q000821:virology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D004622:Embryo, Mammalian; D018690:Excitatory Amino Acid Agonists / Q000633:toxicity; D027342:Excitatory Amino Acid Transporter 2 / Q000378:metabolism; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D016202:N-Methylaspartate / Q000633:toxicity; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D053683:Oncostatin M / Q000009:adverse effects* / Q000494:pharmacology; D053681:Oncostatin M Receptor beta Subunit / Q000378:metabolism; D015682:Retroviridae Proteins, Oncogenic / Q000633:toxicity; D015398:Signal Transduction / Q000187:drug effects
D027322:Amino Acid Transport System X-AG; D000970:Antineoplastic Agents; D016207:Cytokines; D018690:Excitatory Amino Acid Agonists; D027342:Excitatory Amino Acid Transporter 2; D005904:Glial Fibrillary Acidic Protein; D053681:Oncostatin M Receptor beta Subunit; C508349:Osmr protein, mouse; D015682:Retroviridae Proteins, Oncogenic; C493396:Slc1a2 protein, mouse; C060954:p24 protein, Human T-lymphotropic virus 1; D053683:Oncostatin M; D001224:Aspartic Acid; D018698:Glutamic Acid; D016202:N-Methylaspartate
Astrocytes; Excitotoxicity; GLAST; GLT-1; Glutamate; HIV; Interleukin 6; NMDA; Oncostatin M