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HIV Infection Induces Extracellular Cathepsin B Uptake and Damage to Neurons.

ABSTRACT: HIV-associated neurocognitive disorders prevail in 20-50 percent of infected individuals. Macrophages transmigrate through the blood brain barrier during HIV-1 infection, triggering neuronal dysfunction. HIV-infected macrophages secrete cathepsin B (CATB), and serum amyloid p component (SAPC), inducing neuronal apoptosis by an unknown mechanism. We hypothesized that HIV infection facilitates CATB/SAPC secretion from macrophages followed by neuronal internalization, promoting dysfunction. SK-N-SH neuronal cells were exposed to active recombinant histidine-tagged cathepsin B (His-CATB). His-CATB entry was tracked by intracellular flow cytometry, and neuronal dysfunction was verified by western blot. Macrophage-derived extracellular vesicles (EVs) were tested for the presence of CATB and SAPC. Neurons internalized His-CATB, an effect that was partially decreased by pre-treatment with anti-CATB antibody. Pre-treatment with CATB and SAPC antibodies decreased cleavage of caspase-3 and restored synaptophysin in neurons. Neurons exposed to macrophage-conditioned media differentially internalized His-CATB, dependent on the HIV replication levels. Finally, CATB and SAPC were secreted in EVs. We report for the first time that CATB is secreted from macrophages both free and in EVs, and is internalized by neurons. Moreover, HIV-replication levels modulate the amount of CATB neuronal uptake, and neuronal dysfunction can be decreased with CATB antibodies. In conclusion, the CATB/SAPC complex represents a novel target against HIV-associated neurocognitive disorders.

SUBMITTER: Cantres-Rosario YM

PROVIDER: S-EPMC6541605 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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HIV Infection Induces Extracellular Cathepsin B Uptake and Damage to Neurons.

Cantres-Rosario Yisel M YM Ortiz-Rodríguez Sarah C SC Santos-Figueroa Aemil G AG Plaud Marines M Negron Karla K Cotto Bianca B Langford Dianne D Melendez Loyda M LM

Scientific reports 20190529 1

HIV-associated neurocognitive disorders prevail in 20-50 percent of infected individuals. Macrophages transmigrate through the blood brain barrier during HIV-1 infection, triggering neuronal dysfunction. HIV-infected macrophages secrete cathepsin B (CATB), and serum amyloid p component (SAPC), inducing neuronal apoptosis by an unknown mechanism. We hypothesized that HIV infection facilitates CATB/SAPC secretion from macrophages followed by neuronal internalization, promoting dysfunction. SK-N-SH ...[more]

PMID: 31142756

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Project description:IntroductionMitochondrial dysregulation is a key event in HIV-1 infection. Recent studies have suggested that age-related neurodegenerative disorders are associated with increased mitochondrial DNA (mtDNA) damage. As accelerated ageing was found in HIV-1 patients, we hypothesized that HIV-1 infection or HIV-1 proteins can lead to mtDNA damage. Unrepaired mtDNA impairs mitochondrial function, which can lead to oxidative stress and cell death. Investigations of mechanisms of mtDNA damage are limited by the lack of available human models.MethodsWe compared mtDNA or nDNA (nuclear DNA) damage in human cortical neurons and PBMC cells. Primary neuronal cultures were incubated with conditioned media from HIV-1 infected PBMC, or HIV-1 viral proteins Tat or Vpr. Total genomic DNA (nuclear and mtDNA) was isolated using the QIAamp Kit. Nuclear and mtDNA were amplified using the long q-PCR/Gene Amp XL Kit. Real-Time RT-PCR using mitochondrial energy metabolism array was performed to assess mitochondrial energy metabolism markers. Superoxide dismutase (SOD) activity in neuronal cells was measured by the OxiSelect SOD Activity Assay. Reactive oxygen species (ROS) were determined by the confocal microscopy. ATP levels were analyzed using ATP determination biochemical assay. Mitochondrial, cytoplasmic and nuclear proteins were studied by quantitative western-blot assay.ResultsWe show that both treatment of neuronal cells with HIV-1 conditioned media, or infection of PBMC with HIV-1 increase mtDNA damage in cells. mtDNA damage was also seen in neuronal cells, incubated with HIV-1 proteins, Tat and Vpr. Next, we confirmed that mtDNA damage was also increased in neuronal cells transfected by Tat expressing plasmids. We showed that mtDNA was not damaged in neuronal cells following treatment with heat inactivated HIV-1 or Tat protein. Further, we demonstrated that HIV-1 or Tat caused more mtDNA damage compared to nuclear DNA damage in neuronal cells. Finally, we showed that Tat dysregulates RNA expression of several genes regulating mitochondrial energy metabolism, suggesting involvement of Tat in mitochondrial bioenergetics in human neurons. Finally, our hypothesis was confirmed by qWestern analysis of mitochondrial and apoptotic proteins demonstrating the accumulation of apoptotic Bax and Bad proteins in mitochondrial fraction of Tat-treated neuronal cells, suggesting toxic effects of Tat on mitochondrial survival.ConclusionWe showed an increase of mtDNA damage in primary neurons, treated with HIV-1 proteins and in PBMC, infected with HIV-1. Increased mtDNA damage can lead to neurodegeneration, and cause neuronal apoptosis. Our system presents a suitable model to study mtDNA changes during HIV-1 infection.

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HIV Infection Induces Extracellular Cathepsin B Uptake and Damage to Neurons.

Publications

HIV Infection Induces Extracellular Cathepsin B Uptake and Damage to Neurons.

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