Amiloride facilitates plasmid entry <i>in vitro</i>.

<p>Cy5-pEGFP entry into cell lines with or without 1 mM amiloride was analyzed after 2 h and is shown as the percentage of cells that were Cy5⁺. Expression of GFP was analyzed at day 3 and is shown as percentage of EGFP⁺Cy5⁺, on RAW264.7 (A, B, C), JAWSII (D, E), and DC2.4 (F, G). Data represent one of three independent experiments.</p

Amiloride enhances adaptive immunity against HBV S2.

<p>Naïve C57 mice were immunized s.c. with pcD-S2 at various concentrations with or without amiloride in the hind footpad four times using the immunization scheme as shown (A). Seven days after the last immunization, animals (n = 3) were used to test anti-S2 IgG antibody titer (B) and delayed hypersensitivity (DTH) response after re-stimulation with 1 µg sAg <i>s.c.</i> in a hind footpad for 24 h (C). PBS was added as negative control. *, statistical significance among all groups. Another 3 animals were used to test HBV S208–215 specific lysis <i>in vitro</i> (D) and <i>in vivo</i> (E). Hepatocytes from HBV Alb1 trangenic mice were used as targets mixed with effectors from the immunized mice, or transferred into the immunized mice before the analysis of specific lysis <i>in vitro</i> (F) and <i>in vivo</i> (G). *, statistical significance between +/− amiloride.</p

Amiloride increases the frequency of triple positive CD8 T cells.

<p>Splenocytes from mice immunized with pcD-S2 with or without amiloride (n = 3) were re-stimulated in vitro with 10 µg/ml S208–215 for 12 h (A-C), or 10 µg/ml HBsAg for 24 h (D), then cytokine secretion was blocked by monensin for 6 h. PMA or Ionomycin stimulating splenocyte of pcD-S2 immunized mice was added as positive controls. Cells stained with anti-CD3 and anti-CD8 were gated and then used for intracellular staining with single or multiple fluorescent-labeled antibodies. A shows the proportion of responsive cells in the total CD8⁺ T cells with or without amiloride treatment. These responsive cells were designated as either IFN-γ⁺, perforin⁺, or granzymeB⁺ cells. B shows the cytokine expression pattern in the responsive CD8 T cells with or without amiloride treatment. C shows the dose dependent effects of amiloride on the frequency of (IFN-γ⁺perforin⁺granzymeB⁺) triple positive cells. D shows the change in frequency of triple positive cells in response to HBsAg re-stimulation. The change in frequency of triple positive cells after co-culture with peritoneal macrophages followed by re-stimulation with S208–215 (E), or with spleno-DC (F) are also shown. Data represent three independent experiments with similar results.</p

Amiloride accelerates plasmid entry <i>in vivo</i>.

<p>Naïve C57 mice (n = 3) were immunized with Cy5-pEGFP <i>s.c.</i> in hind footpad with (right side) or without (left side) amiloride. Lymph node cells were collected after 4 h (Cy5⁺ cells) and after 24 h (GFP⁺ cells) and examined for proportion (B, D) and subtype (C, E) of stained cells from both draining and control lymph nodes. * in B and D, statistical significance between control and draining lymph nodes in all treated groups.</p