Gfap d1f4q

Antibodies were utilized for immunohistochemistry and western blot analyses. Primary antibodies against GFAP (D1F4Q) and Aβ (D54D2), PS1 (D39D1) and BACE (D10E5) were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). The Iba1 antibody (019-19741) was purchased from Wako Chemicals USA, Inc. (Richmond, VA, USA). The CD68 (MCA1957) antibody was purchased from Bio-Rad Laboratories, Inc. (Hercules, CA, USA). The anti-APP, Y188 (ab32136) antibody was purchased from Abcam (Cambridge, MA, USA). The GAPDH (sc32233) antibody was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Elite Vectastain ABC reagents, Vector VIP, biotinylated anti-rabbit, and anti-mouse secondary antibodies were purchased from Vector Laboratories, Inc (Burlingame, CA, USA).

Liver histology and immunohistochemistry were performed in paraffin or cryostat sections as previously published.²⁸ Liver specimens were fixed with a 10% buffered formalin solution, embedded in paraffin, and processed for hematoxylin and eosin (H&E) staining. To access the percentage of steatosis, livers were fixed in 4% parafomaldehyde in PBS, embedded in OCT, and processed for Oil Red O staining. Immunohistochemistry was performed using Mac-1 (M1/70) and Ly-6G (1A8) from BD Biosciences, MMP-9 (AF909; R&D Systems), and Tnc (MTn-12: Abcam) antibodies at optimal dilutions. Triple staining of liver tissues and isolated HSCs was detected by immunofluorescence with Alexa Fluor 488 (green) for Tnc and Alexa Fluor 594 (red) for GFAP (D1F4Q; Cell Signaling), Alexa Fluor 594 (red) for CD31 (Santa Cruz Biotechnology) and Alexa Fluor 594 (red) for CD68 (Abcam). Vectashield mounting media with DAPI (Vector Laboratories) was used for nuclear staining. Sections were blindly evaluated by counting 10 high-powered fields (HPFs)/section in triplicate.