Перегляд вихідного коду сторінки UNC2881: An Unequivocable Efficiency!

In this assay, larvae are fed the quenched fluorescent phospholipid PED6, which is cleaved in the intestine, activating the fluorescent signal ( Farber et al., 2001). The fluorescent metabolites are then transported through the biliary system to the gallbladder ( Fig.?1E). PED6 fluorescence can be observed in the intestines of s445 mutant larvae ( Fig.?1F, asterisk). However, in s445 mutant larvae, no PED6 signal is observed in the gallbladder, despite the presence of a morphologically identifiable [http://en.wikipedia.org/wiki/MERTK UNC2881] gallbladder and extrahepatic bile duct ( Fig.?1H). Based on these data, we hypothesized that the defect in PED6 metabolite transportation is due to an impairment of the intrahepatic biliary network. To test this hypothesis, we undertook time course observations of intrahepatic biliary network formation in s445 mutants. Tg(Tp1-MmHbb:EGFP)um14 is expressed by intrahepatic biliary epithelial cells in the zebrafish liver ( [http://www.selleckchem.com/products/SRT1720.html see more] Lorent et al., 2010?and?Parsons et al., 2009). At 80?hpf, we did not observe any difference in Tg(Tp1-MmHbb:EGFP)um14 expression in the s445 mutant liver ( Figs. 2A and D), suggesting that the intrahepatic biliary network initially formed. At 100?hpf, the Tg(Tp1-MmHbb:EGFP)um14 positive intrahepatic biliary network is highly branched in wild-type larvae. However, in contrast, by 100?hpf in s445 mutant larvae, Tg(Tp1-MmHbb:EGFP)um14 expression largely disappears from the liver, with only weak fluorescence detected in the central part of the liver ( Fig.?2E). At 120?hpf, Tg(Tp1-MmHbb:EGFP)um14 fluorescence cannot be detected in the s445 mutant liver ( Fig.?2F). These data suggest that the intrahepatic biliary network forms initially [http://www.selleckchem.com/products/Adrucil(Fluorouracil).html selleck screening library] but disappears by 120?hpf in s445 mutant larvae. To better understand the defects caused by the s445 mutation, we visualized the three-dimensional structure of the intrahepatic biliary network. Immunofluorescence staining for Alcam, the intrahepatic biliary network marker ( Sakaguchi et al., 2008), was used to visualize the intrahepatic biliary network ( Fig.?3A). Consistent with the time course observations, we did not observe any difference in Alcam or Tg(Tp1-MmHbb:EGFP)um14 expression in s445 mutant larvae at 80?hpf ( Figs. 3A and B; Supplementary Fig.?2), supporting the idea that biliary epithelial cells initially differentiate in s445 mutant larvae. However, in s445 mutant larvae at 100?hpf, Tg(Tp1-MmHbb:EGFP)um14 expression in the liver was significantly different from wild-type siblings ( Fig.?3D). In the wild-type livers, the Tg(Tp1-MmHbb:EGFP)um14 expressing intrahepatic biliary network added new branches ( Fig.?3C). In contrast, the intrahepatic biliary network appears to be degenerating in the s445 mutant livers ( Fig.?3D). In s445 mutant larvae, we detected fluorescent TUNEL-positive/Tg(Tp1-MmHbb:EGFP)um14-positive cells in the liver (average 6.6 cells per liver, SD?=?4.9, n?=?5, p?