Figure 4: ABCA1 has a pivotal function in extracellular IPP release.

(a) Intracellular and extracellular IPP levels generated by untreated (ZA-) and ZA-treated (ZA+) DC incubated for 24 h in fresh medium (CTRL) or with 10 μM probucol to inhibit ABCA1 function. Probucol marginally increased intracellular IPP but significantly decreased extracellular IPP release in ZA-treated DC (**P<0.01; ANOVA). The bars represent the mean±s.e.m. of four experiments. (b) Western blot analysis of ABCA1 expression in untreated (ZA-) and ZA-treated (ZA+) DC after Abca1-silencing with siRNA (scrambled [scr] non-targeting siRNA; siAbca1: Abca1-silencing siRNA). The blots are representative of one out of the three experiments. β-tubulin was employed as the control of equal protein loading. (c) Intracellular and extracellular IPP levels in DC incubated for 48 h in fresh medium (CTRL) or in the presence of scr or Abca1-silencing siRNA (siAbca1). ZA (1 μM) was added in the last 24 h. Extracellular IPP release is significantly lower in Abca1-silenced ZA-treated DC. Bars represent the mean±s.e.m. of six experiments (**P<0.01; ***P<0.001; ANOVA). (d) Vγ9Vδ2 T cell proliferation after 7-day PBMC stimulation with supernatants from DC treated as reported in c. Supernatant from Abca1-silenced ZA-treated DC is unable to induce Vγ9Vδ2 T cell proliferation (**P<0.01; Wilcoxon–Mann–Whitney). The bars represent the mean±s.e.m. of six experiments. ANOVA, analysis of variance.