Circulating Exosomal Proteins are linked to Neuropathogenesis in SIV‐infected Rhesus Macaque: A Proteomic Approach

Abstract

BackgroundCombined antiretroviral therapy suppresses HIV replication, but 30‐60% of patients suffer from HIV‐1 associated neurocognitive disorders (HAND). Studies are uncovering the role of extracellular vesicles (EVs), especially exosomes, in HIV CNS infection. Using proteomics, we investigated possible circulating exosomal protein links to neuropathogenesis in SIV‐infected rhesus macaque (RM).MethodsExosomes were isolated from sera of SIV‐infected (SIV‐Exo) and uninfected (CTL‐Exo) RM (N = 3/group) by QIAGEN exoEasy kit and characterized by the qNano‐IZON system. Proteomic analysis of the isolated exosomes was performed using liquid chromatography/mass spectrometry (LC‐MS/MS).ResultsqNano‐IZON analysis indicated that isolated EVs were predominantly exosomes (particle size < 150 nm). In the LC‐MS/MS study, 5,654 proteins were quantified, with 236 proteins (~ 4%) significantly differentially expressed (DE) between CTL‐Exo and SIV‐Exo. Two or more unique peptides were detected in 85% (4777/5654) of quantified proteins, and in 89% (211/236) of significant DE proteins, indicating the depth of analysis. We quantified most of the exosome‐associated proteins (tetraspanins, enzymes, lipid rafts, cytoskeletal, and endosome‐specific proteins) reported in previous studies. The heat‐maps and hierarchical clustering indicated that proteins involved in latent viral reactivation (heat shock transcription factor 1), inflammation (complement factor H, antioxidants, glycoproteins), unfolded protein response (UPR) (proteasome activators, cochaperones), neuropathology (amyloid beta [Aβ] precursor, chromogranin‐A and ‐B), and signaling (cytoskeleton regulators, cyclin‐H, mTOR complex 2, CD74) were expressed at significantly higher levels in SIV‐Exo than CTL‐Exo. However, proteins involved in mitochondrial (Mt) fission (Mt‐fission 1, ‐fission factor, and fission regulator 1), and ATP production (Mt Complex‐I, ‐IV, and ‐V), that play a critical role in the brain energy supply, were significantly decreased in SIV‐Exo. Moreover, exosomal‐proteins involved in autophagy‐mediated degradation (autophagy related 9A, ‐2B, lysosomal associated membrane protein 2), endosomal recycling (sorting nexin 4) and exocytosis (synaptogyrin), sprouting angiogenesis (jumonij domain‐containing 6), and cytoskeleton organization (calponin) were also expressed at significantly lower levels in SIV‐Exo than CTL‐Exo.ConclusionsOur novel findings suggest that circulating exosomal proteins are associated with viral reactivation, inflammation, UPR, mitochondrial dysfunction, defective autophagy, and Aβ and Tau pathology that may elucidate the etiology of HAND, and possibly provide novel therapeutic targets.