Western blots were performed as previously described [20 (link)]. The primary antibodies used were anti-MyHC (B103, 0.5 μg/mL, DHSB, Iowa City, IA, USA), anti-MYOD (ABP53067, 1:500, Abbkine, Wuhan, China), anti-MYH1A (F59, 0.5 μg/mL, DHSB, Iowa City, IA, USA), anti-MYH7B (S58, 0.5 μg/mL, DHSB, Iowa City, IA, USA), anti-NAMPT (bs-0272R, 1:500, Bioss, Beijing, China), anti-p-AMPK (ABN-PAB12602, 1:2000, Abnova, Taipei City, Taiwan, China), anti-AMPK (bs-1115R, 1:500, Bioss, Beijing, China), anti-PGC1α (66369–1-Ig, 1:5000, Proteintech, IL, USA), and anti-β-Tubulin (A01030, 1:10,000, Abbkine, Wuhan, China). ProteinFind Goat Anti-Mouse IgG (H + L), HRP Conjugate (HS201–01, 1:1000, TransGen, Beijing, China) and ProteinFind Goat Anti-Rabbit IgG (H + L), HRP Conjugate (HS101–01, 1:500, TransGen, Beijing, China) were used as a secondary antibody.
Immunofluorescence were performed using anti-Desmin (bs-1026R, 1:100, Bioss, Beijing, China) and anti-MyHC (B103, 2.5 μg/mL, DHSB, Iowa City, IA, USA), as previously described [20 (link)]. A fluorescence microscope (DMi8; Leica, Germany) was used to capture three randomly selected fields to visualize the area labeled with anti-MyHC.
Immunofluorescence were performed using anti-Desmin (bs-1026R, 1:100, Bioss, Beijing, China) and anti-MyHC (B103, 2.5 μg/mL, DHSB, Iowa City, IA, USA), as previously described [20 (link)]. A fluorescence microscope (DMi8; Leica, Germany) was used to capture three randomly selected fields to visualize the area labeled with anti-MyHC.