Ab78540

Formalin fixed and paraffin embedded liver fibrosis tissue arrays were purchased from US Biomax (LV805a) (Rockville, MD). Tissue array patient information is shown in Supplementary Table 2. IHF was performed according to a previous report34 (link). Thin cryosections (4 μm) of liver tissue were fixed in acetone for immunohistofluorescence, stained with the indicated antibodies. Antibodies used in IHF were: anti-CUGBP1 (Santa Cruz Biotechnology, SC-20003, 1: 50), α-SMA (Santa Cruz Biotechnology, SC-32251, 1: 50), Reelin (Abcam, ab78540, 1: 50), IFN-γ (Abcam, ab133566, 1: 50), Cytoglobin (Abcam, ab57713, 1: 50), CUGBP1 conjugated to Alexa Fluor 488 (Abcam, ab129115, 1: 100), IFN-γ conjugated to PE-CF594 (BD Biosciences, 562303, 1: 50), goat anti-mouse IgG2a conjugated to Alexa Fluor 594 (Invitrogen, A-21135, 1: 500), goat anti-mouse IgG1 conjugated to Alexa Fluor 647 (Invitrogen, A-21240, 1: 500), goat anti-rabbit IgG conjugated to Alexa Fluor 594 (Invitrogen, R37117, 1: 500), goat anti-mouse IgG2b conjugated to Alexa Fluor 488 (Invitrogen, A-21141, 1: 500). The sections were then stained with DAPI and examined with a confocal laser scanning microscope (Leica, Wetzlar, Germany). Sirius Red staining was performed by Servicebio (Wuhan, China). The liver fibrosis stage was assessed by Ishak scale35 (link).

Affected hippocampal tissues were lysed in ice-cold RIPA buffer (Beyotime, China), containing PMSF (Beyotime, China) and phosphatase inhibitor (Solarbio, China). Total protein concentration was determined by the BCA protein assay (Beyotime, China). Aliquots of homogenate with equal protein concentration were separated by 10% SDS-PAGE gel and then transferred to polyvinyl difluoramine membrane. After blocking in 5% non-fat milk for 1.5 h at room temperature, the membranes were then incubated with primary antibodies, including mouse anti-reelin (1:1,000, ab78540, abcam), mouse anti-tau (tau-5, 1:5,000, ab80579, abcam), and rabbit anti-tau (phospho-S199/202, 1:5,000, ab4864, abcam) at 4°C overnight. The blots were then incubated with anti-rabbit or anti-mouse IgG conjugated to HRP for 2 h at room temperature and visualized using the enhanced chemiluminescence.

The following primary antibodies were used for immunofluorescence staining: goat anti-tdTomato (1:500; catalog #AB8181-200, SICGEN; RRID:AB_2722750); rabbit anti-TRP73 (1:500; catalog #ab40658, Abcam; RRID:AB_776999); mouse anti-RELN (1:500; catalog #MAB5364, MilliporeSigma; RRID:AB_1293544); mouse anti-RELN (1:500; catalog #ab78540, Abcam; RRID:AB_1603148); rabbit anti-CALB1 (1:500; catalog #CB38, Swant; RRID:AB_10000340); mouse anti-DCX (1:25; catalog #sc-271390, Santa Cruz Biotechnology; RRID:AB_10610966); and goat anti-EGFP/YFP (yellow fluorescent protein; 1:500; catalog #AB0020-500, SICGEN; RRID:AB_2333100). The secondary antibodies used were as follows: donkey anti-goat Alexa Fluor 555 (1:1000; catalog #A21432, Thermo Fisher Scientific; RRID:AB_2535853); donkey anti-rabbit Alexa Fluor 488 (1:1000; catalog #A21206, Thermo Fisher Scientific; RRID:AB_2535792); and donkey anti-mouse Alexa Fluor 647 (1:1000; catalog #A31571, Thermo Fisher Scientific; RRID:AB_162542).