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Role of endolysosomes in HIV-1 Tat-induced neurotoxicity.

Hui L, Chen X, Haughey NJ, Geiger JD - ASN Neuro (2012)

Bottom Line: Combined anti-retroviral therapeutic drugs effectively increase the lifespan of HIV-1-infected individuals who then have a higher prevalence of HAND (HIV-1 associated neurocognitive disorder).As early as 24 h after HIV-1 Tat was applied to neurons, HIV-1 Tat accumulated in endolysosomes, endolysosome morphology was affected and their size increased, endolysosome membrane integrity was disrupted, endolysosome pH increased, specific activities of endolysosome enzymes decreased and autophagy was inhibited, as indicated by the significant changes in three markers for autophagy.In contrast, statistically significant levels of HIV-1 Tat-induced neuronal cell death were observed only after 48 h of HIV-1 Tat treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of North Dakota School of Medicine and Health Sciences, Grand Forks, ND 58203, USA.

ABSTRACT
Combined anti-retroviral therapeutic drugs effectively increase the lifespan of HIV-1-infected individuals who then have a higher prevalence of HAND (HIV-1 associated neurocognitive disorder). Soluble factors including HIV-1 proteins released from HIV-1-infected cells have been implicated in the pathogenesis of HAND, and particular attention has been paid to the HIV-1 Tat (transactivator of transcription) protein because of its ability to directly excite neurons and cause neuronal cell death. Since HIV-1 Tat enters cells by receptor-mediated endocytosis and since endolysosomes play an important role in neuronal cell life and death, we tested here the hypothesis that HIV-1 Tat neurotoxicity is associated with changes in the endolysosome structure and function and also autophagy. Following the treatment of primary cultured rat hippocampal neurons with HIV-1 Tat or as controls mutant-Tat or PBS, neuronal viability was determined using a triple staining method. Preceding observations of HIV-1 Tat-induced neuronal cell death, we observed statistically significant changes in the structure and membrane integrity of endolysosomes, endolysosome pH and autophagy. As early as 24 h after HIV-1 Tat was applied to neurons, HIV-1 Tat accumulated in endolysosomes, endolysosome morphology was affected and their size increased, endolysosome membrane integrity was disrupted, endolysosome pH increased, specific activities of endolysosome enzymes decreased and autophagy was inhibited, as indicated by the significant changes in three markers for autophagy. In contrast, statistically significant levels of HIV-1 Tat-induced neuronal cell death were observed only after 48 h of HIV-1 Tat treatment. Our findings suggest that endolysosomes are involved in HIV-1 Tat-induced neurotoxicity and may represent a target for therapeutic intervention against HAND.

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HIV-1 Tat-elevated endolysosome pH in primary cultured neuronsEndolysosome pH was measured ratio-metrically using LysoSensor dye. HIV-1 Tat (100 nM) treatment for 1 and 2 days elevated significantly endolysosome pH (n = 20).
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Figure 3: HIV-1 Tat-elevated endolysosome pH in primary cultured neuronsEndolysosome pH was measured ratio-metrically using LysoSensor dye. HIV-1 Tat (100 nM) treatment for 1 and 2 days elevated significantly endolysosome pH (n = 20).

Mentions: The observation that HIV-1 Tat alters endolysosome morphology led us to determine next the extent to which HIV-1 Tat-affected endolysosome function. Since pH is critical for endolysosome function, we determined the extent to which HIV-1 Tat-affected endolysosome pH using LysoSensor dye, which permits ratio-metric assessment of pH changes in acidic organelles. We found that HIV-1 Tat, but not mutant Tat treatment for 1 or 2 days elevated significantly (n = 20, P<0.001) endolysosome pH in cultured hippocampal neurons (Figure 3). As endolysosome pH affects endolysosome enzyme activity, we next determined the protein levels and activity of endolysosome enzymes as evaluations of endolysosome function. Treatment of neurons with HIV-1 Tat for 1 or 2 days increased significantly protein levels of the endolysosome enzymes acid phosphatase (Figure 4A, n = 4, P<0.01 at 1 day and n = 4, P<0.05 at 2 days), cathepsin B (Figure 4C, n = 4, P<0.05) and cathepsin D (Figure 4E, n = 4, P<0.01 at 1 day and P<0.05 at 2 days). However, the specific activity levels of acid phosphatase (Figure 4B), cathepsin B (Figure 4D) and cathepsin D (Figure 4F) were decreased significantly (n = 4, P<0.001) in HIV-1 Tat-treated cultures.


Role of endolysosomes in HIV-1 Tat-induced neurotoxicity.

Hui L, Chen X, Haughey NJ, Geiger JD - ASN Neuro (2012)

HIV-1 Tat-elevated endolysosome pH in primary cultured neuronsEndolysosome pH was measured ratio-metrically using LysoSensor dye. HIV-1 Tat (100 nM) treatment for 1 and 2 days elevated significantly endolysosome pH (n = 20).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3379000&req=5

Figure 3: HIV-1 Tat-elevated endolysosome pH in primary cultured neuronsEndolysosome pH was measured ratio-metrically using LysoSensor dye. HIV-1 Tat (100 nM) treatment for 1 and 2 days elevated significantly endolysosome pH (n = 20).
Mentions: The observation that HIV-1 Tat alters endolysosome morphology led us to determine next the extent to which HIV-1 Tat-affected endolysosome function. Since pH is critical for endolysosome function, we determined the extent to which HIV-1 Tat-affected endolysosome pH using LysoSensor dye, which permits ratio-metric assessment of pH changes in acidic organelles. We found that HIV-1 Tat, but not mutant Tat treatment for 1 or 2 days elevated significantly (n = 20, P<0.001) endolysosome pH in cultured hippocampal neurons (Figure 3). As endolysosome pH affects endolysosome enzyme activity, we next determined the protein levels and activity of endolysosome enzymes as evaluations of endolysosome function. Treatment of neurons with HIV-1 Tat for 1 or 2 days increased significantly protein levels of the endolysosome enzymes acid phosphatase (Figure 4A, n = 4, P<0.01 at 1 day and n = 4, P<0.05 at 2 days), cathepsin B (Figure 4C, n = 4, P<0.05) and cathepsin D (Figure 4E, n = 4, P<0.01 at 1 day and P<0.05 at 2 days). However, the specific activity levels of acid phosphatase (Figure 4B), cathepsin B (Figure 4D) and cathepsin D (Figure 4F) were decreased significantly (n = 4, P<0.001) in HIV-1 Tat-treated cultures.

Bottom Line: Combined anti-retroviral therapeutic drugs effectively increase the lifespan of HIV-1-infected individuals who then have a higher prevalence of HAND (HIV-1 associated neurocognitive disorder).As early as 24 h after HIV-1 Tat was applied to neurons, HIV-1 Tat accumulated in endolysosomes, endolysosome morphology was affected and their size increased, endolysosome membrane integrity was disrupted, endolysosome pH increased, specific activities of endolysosome enzymes decreased and autophagy was inhibited, as indicated by the significant changes in three markers for autophagy.In contrast, statistically significant levels of HIV-1 Tat-induced neuronal cell death were observed only after 48 h of HIV-1 Tat treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of North Dakota School of Medicine and Health Sciences, Grand Forks, ND 58203, USA.

ABSTRACT
Combined anti-retroviral therapeutic drugs effectively increase the lifespan of HIV-1-infected individuals who then have a higher prevalence of HAND (HIV-1 associated neurocognitive disorder). Soluble factors including HIV-1 proteins released from HIV-1-infected cells have been implicated in the pathogenesis of HAND, and particular attention has been paid to the HIV-1 Tat (transactivator of transcription) protein because of its ability to directly excite neurons and cause neuronal cell death. Since HIV-1 Tat enters cells by receptor-mediated endocytosis and since endolysosomes play an important role in neuronal cell life and death, we tested here the hypothesis that HIV-1 Tat neurotoxicity is associated with changes in the endolysosome structure and function and also autophagy. Following the treatment of primary cultured rat hippocampal neurons with HIV-1 Tat or as controls mutant-Tat or PBS, neuronal viability was determined using a triple staining method. Preceding observations of HIV-1 Tat-induced neuronal cell death, we observed statistically significant changes in the structure and membrane integrity of endolysosomes, endolysosome pH and autophagy. As early as 24 h after HIV-1 Tat was applied to neurons, HIV-1 Tat accumulated in endolysosomes, endolysosome morphology was affected and their size increased, endolysosome membrane integrity was disrupted, endolysosome pH increased, specific activities of endolysosome enzymes decreased and autophagy was inhibited, as indicated by the significant changes in three markers for autophagy. In contrast, statistically significant levels of HIV-1 Tat-induced neuronal cell death were observed only after 48 h of HIV-1 Tat treatment. Our findings suggest that endolysosomes are involved in HIV-1 Tat-induced neurotoxicity and may represent a target for therapeutic intervention against HAND.

Show MeSH
Related in: MedlinePlus