ENV TMD inhibits secretion of cytokines and chemokines induced by TLR2.

<p>(<b>A–C</b>) RAW cells were incubated with/without the indicated peptides for 2 hours, then washed twice and activated with 500 ng hours, then washed twice and activated with 500 ng ng/ml of purified LTA. Media was collected after 5 hours (TNFα) or 24 hours (MCP-1 and IL-6). ELISA was performed using standard kits. (n = 3, ± represent SD) (<b>D</b>) THP-1 human macrophages were incubated using similar conditions as described in A, and TNFα levels were measured (* - p<0.05).</p

The gp41 TMD reduces serum cytokine levels in vivo and protects from consequential spleen damage.

<p>(<b>A–B</b>) Blood serum TNFα and IL-6 levels of PBS, gp41 TMD and gp41 mutant treated mice estimated 90 minutes and 210 minutes after LTA minutes and 210 minutes after LTA minutes after LTA/GLN injection (n = 9, ** - p<0.005, ***-p<0.0005). (<b>C</b>) H&E staining of spleen sections of control (top), LTA and no peptide (middle) and LTA and gp41 TMD treated (bottom) mice. Apoptotic foci in the white pulp are indicated in circles and red pulp as squares. Scale bars = 200 µm and 400 µm for left and right panels, respectively.</p

The gp41 TMD inhibits TLR2 induced signaling through ERK1,2 and cytokine secretion.

<p>(<b>A</b>) ERK1,2 phosphorylation in RAW264.7 cells was detected before and after LTA exposure. Cells were either pre-treated with indicated peptides for 2 hours prior to LTA addition or untreated (center). GAPDH shows equal loading. Here shown representative data of 3 typical experiments. (<b>B</b>) Quantification of the band intensity was determined for the p-ERK levels after 20 minutes LTA exposure. The WT peptide shows over 50 minutes LTA exposure. The WT peptide shows over 50% decrease in p-ERK levels while the mutant peptide shows approximately 15%. Statistical analysis was done by student T-test, (n = 4, ** p<0.01). (<b>C–D</b>) mRNA expression levels of MCP-1 (C) and TNFα (D) detected by real-time PCR. Expression levels were normalized to the expression of non-activated cells. The data is shown as the calculated 2^∧−dCT in the relativity of each gene to HPRT. (The figure is an average of three samples, * - p<0.05).</p

The gp41 TMD rescues mice in a LTA/GLN sepsis model and reduces liver damage.

<p>(<b>A</b>) Mice were inoculated with a lethal dosage of LTA/GLN and simultaneously treated with either the gp41 TMD peptide (▪) the gp41 mutant (♦) or PBS as control (•). Peptides were administrated at 5 mg mg/kg. (<b>B</b>) H&E staining of liver sections. Apoptotic foci are indicated in circles and arrows indicate hemorrhage. Cell death is multi focal as expected as a result of GLN treatment. Scale bar = 100 µm.</p