Nef is secreted in exosomes from Nef.GFP-expressing and HIV-1-infected human astrocytes
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HIV-1 infection of the central nervous system causes HIV-associated neurocognitive disorders, even in aviremic patients. Although astrocyte malfunction was associated to these disorders, their implication is overshadowed by contributions of microglia and macrophages. Astrocytes are infected with HIV-1 in vivo and express a relevant amount of viral protein Nef. Nef was shown to stimulate its own release in exosomes from diverse cell types, which in turn have damaging effects on neighboring cells. Using immunoblotting and electron microscopy, we showed that human astrocytes expressing Nef.GFP similarly release Nef in exosomes. Importantly, Nef.GFP expression increases the secretion of exosomes from human astrocytes up to 5.5-fold, as determined by total protein content and nanoparticle tracking analysis. Protein analysis of exosomes and viruses separated on iodixanol gradient further showed that native or pseudotyped HIV-1-infected human astrocytes release exosomes, which contain Nef. Our results provide the basis for future studies of the damaging role of Nef-exosomes produced by HIV-infected astrocytes on the central nervous system.
KeywordsAstrocytes HIV-1 Nef Extracellular vesicles Exosomes Central nervous system
We thank Prof. Robert Zorec, PhD, for the access to the Nucleofector 2b, Prof. Yifan Cheng, PhD, for help with TEM imaging and Anja Kejžar, PhD, for helping us with the NTA measurements. This work was supported by research grants J3-5499 and P1-0170 funded by the Slovenian Research Agency (ARRS).
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Conflict of interest
The authors declare that they have no conflict of interest.
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