Yanger_Supplemental information Supplemental Information
-Supplementary Tables 1 and 2
-Supplementary Figures S1-S8
-Supplementary Figure Legends
-Supplementary Methods
-References for Supplementary Methods
基底油Supplementary Table 1. List of Antibodies
Supplementary Table 2. List of primer quences
山西高原Gene Forward(5’ to 3’)Rever(3’ to 5’)干捞面
GAPDH ATGTTCCA GTATGA CTCCACTCA CG GAA GA CA CCA GTA GA CTCCACA CA CK19 GA CCTA GCCAA GATCCTGA GT TCA GCTCCTCAATCCGA GCA EpCAM GCGGCTCA GA GA GA CTGT CCAA GCATTTA GACGCCA GTTT Sox9 ACTCTGGGCAA GCTCTGGA G CGAA GGGTCTCTTCTCGCTCT HNF1βCCCA GCAATCTCA GAA CCTC AGGCTGCTA GCCACA CTGTT AAAGCCTATCATGGA GAA GA GGCG GGAATGCCGGGA GCTATCTTTCTT
Supplementary Figure Legends
消防安全检查内容
春节对联春联Fig. S1. The AAV2/8 rotype specifically transduces hepatocytes.
AAV2/8 virus containing Cre recombina under the control of the ubiquitous cytomegalovirus (CMV) promoter (AAV8-CMV-Cre) or the hepatocyte-specific thyroid binding globulin (TBG) promoter (AAV8-TBG-Cre) were ud to determine the specificity, nsitivity, and tropism of viral infection. The images depict YFP+ lineage-labelled cells either 2 weeks (A) or 10-months (C) after injection of AAV
捏彩泥8-CMV-Cre virus into Rosa YFP animals. Efficient marking of hepatocytes (HNF4α+ cells) with no marking of BECs (CK19+ cells) was obrved under both conditions (B, D), demonstrating that viral tropism (and not the TBG promoter) accounts for hepatocyte-specific labelling. AAV8-TBG-Cre also results in persistent specific labelling of hepatocytes 9 months after infection; again there is no labelling of CK19+ BECs with this virus (E,F). (G) Additionally, all YFP+ cells resulting from AAV8-TBG-Cre recombination (3948 cells counted) were HNF4α+.Scale bar=25μm.
Fig. S2. Notch signalling mediates hepatocyte reprogramming.
(A)Infection of Rosa NICD mice with AAV8-TBG-GFP did not lead to induction of the biliary markers Sox9 or OPN in hepatocytes, and no ectopic CK19+ cells within the hepatic lobule were obrved, demonstrating that AAV8 itlf does not induce the reprogramming phenotype. (B) Infection of Rosa NICD mice with AAV8-TBG-Cre leads to an “intermediate” cell phenotypecharacterized by expression of BEC markers in hepatocytes.HNF1β is expresd exclusively in CK19+BECs in normal liver, but staining was detected in CK19-hepatocytes 1 week following Notch activation.(C) “Zone 3” hepatocytes located clo to the central vein(CV;元宵节几月几号
blood vesl outlined with white dotted line, zone 3 hepatocytes outlined with yellow dotted line) do n
ot an “intermediate” phenotype following AAV8-TBG-Cre injection in Rosa NICD animals. Sox9 and OPN staining are not obrved in the cells despite active Notch signalling (demonstrated by induction of Hes1). (D) Clonal analysis demonstrating that Notch-induced reprogramming is cell autonomous. In Rosa NICD/YFP mice infected with low-do AAV8-TGB-Cre, Sox9 and OPN expression is detected exclusively in YFP+ hepatocytes. (E)PCKζ and Ac-tub (boxed) staining is detected at the apical surfaces of BECs (-AAV). Following the induction of Notch signalling (+AAV), PCKζ and Ac-tub staining is obrved in HNF4α+ hepatocytes that have coalesced to form a central lumen (arrowhead). Scale bar=25μm.
Fig. S3. DDC treatment is associated with significant cellular turnover.
(A) Administration of a DDC diet leads to an ADC respon characterized by the expansion of small BEC-like cells from the peri-portal region into the lobule. Animals given BrdU in the drinking water for two weeks with either regular chow (Vehicle) or in combination with a DDC-containing diet (DDC) exhibit marked proliferation of BECs, marked by CK19 (B, arrowheads), as well as hepatocytes, marked by HNF4α (C, arrowheads). (D) DDC treatment also leads to a significant increa in hepatocyte cell death, as detected by staining for cleaved caspa-3. Scale bar=50μm.
Fig.S4. Hepatocytes express biliary markers shortly after Notch activation/injury.星期天早上
(A)CK19lo are not obrved in normal liver but are apparent at 2 weeks following Notch activation and continue to increa in number and staining intensity over time.(B)“Intermediate” cells emerge early following treatment with DDC (2 weeks). Most YFP+ cells with co-staining
