Anti nmnat1

Paraffin-embedded liver sections were handled using standard methods as described previously [27 ]. The following antibodies were used for immunoblotting: anti-PPARα (Thermo Fisher Scientific), anti-4-HNE (Northwest Life Science Specialties, Vancouver, WA), anti-CHOP (Cell Signaling Technology, Danvers, MA), anti-catalase (EMD Millipore, Burlington, MA), anti-CYP2E1 (Abcam, Cambridge, MA), anti-NADSYN (Antibodies-online, Limerick, PA), anti-TDO2 (Thermo Fisher Scientific), and anti-NMNAT1 (Lifespan Biosciences, Seattle, WA). Methyl green was used to counterstain cell nuclei for 4-HNE staining.

Mouse livers homogenates were loaded onto 12% sodium dodecyl sulfate-polyacrylamide gel, transblotted onto polyvinylidene difluoride membranes (Bio-Rad, Hercules, CA), blocked with 5% nonfat dry milk, and then incubated with the following antibodies, including anti-PPARα, anti-TDO2 (Thermo Fisher Scientific), anti-catalase (EMD Millipore), anti-CYP2E1, anti-ADH, anti-ALDH2, anti-NQO1 (Abcam), anti-GSTM1 (Proteintech, Rosemont, IL), anti-ATF4, anti-GAPDH (Cell Signaling Technology), anti-NADSYN (Antibodies-online), anti-NMNAT1 (Lifespan Biosciences) and anti-β-actin (Sigma-Aldrich, St. Louis, MO). The membranes were then incubated with HRP-conjugated goat anti-rabbit IgG, goat anti-mouse IgG, or rabbit anti-goat IgG (Thermo Fisher Scientific). The bound complexes were detected by enhanced chemiluminescence and quantified by densitometry analysis.

Liver nuclear extracts were prepared using a nuclear extraction kit from Cayman Chemical. PPARα activity in the extracts was assessed by detecting double-stranded DNA-bound PPARα with specific antibodies in an ELISA format (Cayman Chemical). Catalase activity in whole liver homogenates was examined using a catalase activity assay kit (BioVision). The following antibodies were used for immunoblotting: anti-PPARα (Thermo Fisher Scientific), anti-4-HNE (Northwest Life Science Specialties, Vancouver, WA), anti-CHOP (Cell Signaling Technology, Danvers, MA), anti-catalase (EMD Millipore, Burlington, MA), anti-CYP2E1 (Abcam, Cambridge, MA), anti-NADSYN (Antibodies-online, Limerick, PA), anti-TDO2 (Thermo Fisher Scientific), and anti-NMNAT1 (Lifespan Biosciences, Seattle, WA). Methyl green was used to counterstain cell nuclei for 4-HNE staining.