Fig 1: APP expression in exemplary subjects.Comparison of three exemplary BA cases (a–c) that were available for all analyses. Immunostaining of liver APP performed using rabbit monoclonal antibody to APP. Bound antibodies were visualized using HRP-DAB (3,3′-diaminobenzidine tetrahydrochloride) staining. APP-positive areas within hepatocytes are brown in color. BA bile atresia, APP amyloid precursor protein, PCR polymerase chain reaction, mRNA messenger ribonucleic acid, ELISA enzyme-linked immunosorbent assay.
Fig 2: Representative immunofluorescence imaging of APP in control and biliary atresia liver.Figure showing immunofluorescence imaging of control liver (upper row) and bile atresia liver (lower row) of amyloid precursor protein (APP). Note the expression of APP in small intrahepatic bile ducts in bile atresia (arrowhead lower row), while there was no detectable expression of APP in intrahepatic bile ducts of the control liver (arrowhead upper row). APP was visualized using an Alexa Fluor 555 fluorescent secondary antibody. Autofluorescence imaging was generated using a GFP filter cube. Nuclei were stained with DAPI. Magnification 200×, scale bar equals 100 µm.
Fig 3: Amyloidosis-related gene expression.Dot plots showing normalized log2 mRNA counts of amyloid-metabolism-related genes in 4 different outcome groups (a) and in subjects with severe (Ishak fibrosis score >4) vs. less severe (Ishak fibrosis score <5) liver fibrosis (b). Additionally, APP-levels are compared between different Ishak fibrosis scores (c). LTx_LT, late liver transplantation (≥1 year); LTx_ET, early liver transplantation (<1 year); SNL, survival with the native liver; JF_SNL, jaundice-free survival with the native liver. Statistical significance was categorized as *(p < 0.05), **(p < 0.01), ***(p < 0.001), and ****(p < 0.0001).
Fig 4: Plasma quantification of APP.a Dot plots showing plasma levels of APP in bile atresia (BA) subjects vs. age-matched children with inguinal hernia (control). b Dot plots showing APP plasma levels in subjects with different fibrosis severity assessed by Ishak fibrosis score. Data presented as mean (SD), and P values ≤ 0.05 were considered statistically significant. Statistical significance was categorized as *(p < 0.05), **(p < 0.01), ***(p < 0.001), and ****(p < 0.0001).
Fig 5: Immunostaining of liver APP performed using rabbit monoclonal antibody to APP.Bound antibodies were visualized using HRP-DAB (3,3′-diaminobenzidine tetrahydrochloride) staining. APP-positive areas are specifically stained in brown. Images were taken after automated whole-slide imaging using the APERIO CS2 scanner (Leica Biosystems, Wetzlar, Germany) and ImageScope software version 12.3.3.5048 (Leica Biosystems). Healthy liver tissue was used as a control. BA bile atresia. APP amyloid precursor protein. Statistical significance was categorized as *(p < 0.05), **(p < 0.01), ***(p < 0.001), and ****(p < 0.0001).
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