Distinct gene expression profiles between human preterm-derived and adult-derived intestinal organoids exposed to Enterococcus faecalis: a pilot study

the NTCPmediated as well as Oatp1a/1bmediated bile salt uptake machinery is rapidly downregulated in inflammatory conditions, which may explain the modestly elevated taurocholic acid (TCA) elevation seen in CCL4/antiNTCP injected mice. Because our research interests centre around hepatobiliairy transport processes, including NTCP function, we studied NTCP cell surface presence and activity in LX2 cells without relying on antibodies (online supplemental material). We performed bile salt uptake assays in the presence and absence of the specific and wellcharacterised NTCP inhibitory peptide Myrcludex B (also called bulevirtide). In contrast to U2OS cells overexpressing human NTCP, HTCA as well as TauroNORTHCA24DBD uptake was at background levels in LX2 cells, independent of TGFß-induced activation, and could not be further blocked with Myrcludex B (figure 1A,B). TGFβ-activated LX2 cells did not show any plasma membrane labelling on incubation with FITClabelled Myrcludex B (figure 2), in contrast to NTCPpositive U2OS cells. This suggests that no, also no lowabundance subsets of, NTCPpositive cells were present in this cell population. Our bile salt uptake data are in sharp contrast to the data provided by the authors, where the culture medium containing 100 μM TCA is completely depleted from TCA after 45 min, supposedly due to uptake activity of NTCP. If we (safely) assume that the height of the cells is <1% of the total height of the culture medium, the intracellular TCA concentration would need to reach at least 10 mM inside these NTCPpositive stellate cells, an unlikely scenario as this is above the critical micelle concentration of TCA. In conclusion, we believe that a direct role of NTCP activity in stellate cells contributing to fibrosis is not proven by the experiments presented by Salhab et al. More investigations using specific inhibitors and better controlled conditions are necessary before we can pursue NTCP in stellate cells as a promising antifibrotic target.