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Fig. 1 | Orphanet Journal of Rare Diseases

Fig. 1

From: In vitro functional rescue by ivacaftor of an ABCB11 variant involved in PFIC2 and intrahepatic cholestasis of pregnancy

Fig. 1

Characterisation of the p.A257V variant of ABCB11 and effect of ivacaftor on taurocholate (TC) transport activity. A Structure of wild type (wt) human ABCB11 in an inward-open state solved at 3.5 Å using cryogenic electron microscopy (PDB ID: 6LR0 [18]). The fourth transmembrane helix (TM4) is colored in blue and the Ala257 residue is depicted in red. B MDCK cells stably expressing Abcb11-GFP (wt or A257V) and Ntcp-cMyc were lysed and analysed by immunoblotting. This panel is representative of at least six independent experiments. C Electrophoretic patterns from B were separately quantified and their relative amounts were calculated and normalized to β-actin. Results are means (± SEM) of at least three experiments. ns: not significant (Student t test). D Abcb11-GFP (wt or A257V) and Ntcp-cMyc were stably expressed in MDCK cells. After immunolabelling, Abcb11 (green) and Ntcp (red) were visualized by confocal microscopy. Bottom, centre and right panels show x–z, x–y and y–z plane images, respectively. Bars: 10 μm. E Abcb11-wt-GFP and Abcb11-A257V-GFP were transiently expressed in HepG2 cells. After immunolabelling, Abcb11 (green) and ABCC2 (red) were visualised by confocal microscopy. *Bile canaliculus. Bars: 10 μm. F Vectorial transport of [3H]TC in MDCK cells stably expressing Abcb11 (wt or A257V) and Ntcp in the absence (−) or presence of the indicated concentrations of ivacaftor. The dashline indicates [3H]TC transport measured in MDCK cells expressing Ntcp alone. Means (± SEM) of at least six independent experiments for each tested condition are shown. ****P < .0001 versus wt; and $$P < .01; $$$P < .005 versus non-treated A257V cells (one-way ANOVA)

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