a. Scatter plot analysis of the changes in synaptic plasticity gene expression in clade B infected astrocytes.

<p>Pair wise comparison of control primary human astrocytes (No HIV infection) and clade B infected astrocytes by scatter plot analysis. Spots associated with individual human synaptic plasticity gene were collected and converted into log10 scale. The central line indicates unchanged gene expression. The synaptic plasticity genes with expression levels higher or lower in clade B infected astrocytes than the control cells are expected to produce dots that deviate from the centerline. <b>b. Scatter plot analysis of the changes in synaptic plasticity gene expression in clade C infected astrocytes.</b> Pair wise comparison of control primary human astrocytes (No HIV infection) and clade C infected astrocytes by scatter plot analysis. Spots associated with individual human synaptic plasticity gene were collected and converted into log10 scale. The central line indicates unchanged gene expression. The dots are allocated to positions that are above or below than the +3 fold or −3 fold line when the differences are greater than three folds.</p

Neuronal dendrite and spine measurement by Image J analysis.

<p>A typical dendrite segment from a pyramidal neuron is shown, and the six quantification parameters labeled as follows. 1) Total dendrite area is measured by drawing a box around the whole image; 2) dendrite diameter is obtained by drawing a line across the dendrite thickness at a place of average width; 3) spine density is the total number of spines divided by the dendrite length; 4) spine area is measured by drawing a box around the whole spine; 5) Dendrite length uses the broken line tool to measure the length and 6) Spine length. (Smith et al., 2009. Reversal of long-term dendritic spine alterations in Alzheimer disease models. Proc Natl Acad Sci U S A 106: 16877-16882. Copyright (2009) National Academy of Sciences, U.S.A.)</p

Apoptosis induction in HIV infected primary human astrocytes.

<p>Apoptosis induction was studied using double cell labeling with Annexin V-PE and 7-AAD. Seven days after infection, cells were collected and re-suspended in binding buffer containing Annexin V-PE and 7-AAD, and then processed for flow cytometry analysis. Representative flow figure for one of three experiments is shown here. In each box, cells in lower left corner (negative for 7-AAD and Annexin V-PE) are viable, cells in the upper right corner (positive for 7-AAD and Annexin V-PE) are necrotic or late apoptotic cells, while cells in the lower right corner (Annexin V-PE positive but negative to 7-AAD) are early apoptotic cells.</p

Human synaptic plasticity genes down-regulated in HIV-1 clade B & C infected primary human astrocytes (fold down regulation): Out of 84 genes analyzed, only the genes significantly (≥3 fold) down-regulate were shown in this table.

1<p>Immediate-Early Response Genes (IEGs);</p>2<p>Long Term Potentiation (LTP);</p>3<p>Long Term Depression (LTD);</p>4<p>Cell Adhesion;</p>5<p>Extracellular Matrix & Proteolytic Processing;</p>6<p>CREB Cofactors;</p>7<p>Neuronal Receptors;</p>8<p>Postsynaptic Density;</p>9<p>Others.</p

Confocal Images of DiI stained SK-N-MC cells.

<p>(4a) Control SK-N-MC cells: High number of long spines on the dendritic length, high dendrite diameter, total dendrite area and spine area. (4b) Clade B infected cells: Loss of spines on the dendrite length, decreased dendrite diameter, dendrite and spine area was observed than the control and clade C infected SK-N-MC cells. (4c) Clade C infected cells: Loss of number of spines, decreased dendrite diameter, dendrite area and spine area was observed than control cells.</p

Functional Grouping of Human Synaptic Plasticity genes.

<p>Functional Grouping of Human Synaptic Plasticity genes.</p

Effects of Alcohol and/or HDACi on Oxidative Stress and Antioxidant Defense Genes.

<p>Effects of Alcohol and/or HDACi on Oxidative Stress and Antioxidant Defense Genes.</p

Human synaptic plasticity genes up-regulated in HIV-1 clade B & C infected primary human astrocytes (fold up regulation).

1<p>Immediate-Early Response Genes (IEGs);</p>2<p>Late Response Genes;</p>3<p>Postsynaptic Density.</p

Dendrite morphology changes in HIV-1 clade B and clade C infected SK-N-MC neuroblastoma cells.

<p>SK-N-MC neuroblastoma cells were grown onto the glass coverslips and infected with the HIV clade B and C for 7 days. Cover slips were stained with the DiI stain and observed under confocal microscopy. Randomly selected pictures in each group of the cells were captured in confocal microscope. Image J software was used to analyze the total dendrite area (5a), spine density (5b), dendrite diameter (5c), spine area (5d), spine length (5e) and number of spines (5f).</p

Modulatory effects of Withanolide A and β-Amyloid_1-42 on human neuronal cells.

<p>A. Dose-response of Withanolide A showing growth-stimulatory effects in terms of % viability. B. MTT assay showing the inhibition of cell viability by β-Amyloid_1-42 and its reversal by Withanolide A at two concentrations on SK-N-MC cells. The values were expressed as percentage of cell viability compared to control cells and are the mean ± SD of four experiments. *** and ** indicates a statistically significant difference (p<0.0001) and (p<0.0018) respectively compared to controls. WA – Withanolide A; β-AMY - β-Amyloid_1-42</p