The ischemic zone of the left ventricle (LV) heart tissue was snap frozen and ground with mortar and pestle in liquid nitrogen. The powdered tissue was then suspended in RIPA lysis buffer.
29 (link) H9c2 cells were used for Western blot analysis and cells were suspended in lysis buffer. Proteins were separated by reducing SDS‐PAGE and transferred to a PVDF membrane. The following antibodies were used for Western blot: p‐AMPK T172 (1:1000, Cell Signaling, #2535), p‐ULK1 S555 (1:1000, Cell Signaling, #5869), GAPDH (1:2000, ThermoFisher, #MA5‐15738), Adiponectin (1:1000, ImmunoDiagnostics, #12010), ATG5‐12 (1:1000, Novus Biologicals, #NB110‐53818), ATG7 (1:1000, Santa Cruz, #sc‐376212), Beclin1 (1:1000, Cell Signaling, #3738), LC3B (1:1000, Cell Signaling, #2775), p62 (1:1000, Cell Signaling, #5114), BAX (1:1000, Cell Signaling, #2772), cleaved Caspase 3 (1:1000, Cell Signaling, #9661), bcl‐2 (1:1000, Cell Signaling, #3489), and β‐actin (1:2000, ThermoFisher, #PA1‐46296) were used. The quantification of signals was performed by densitometry of scanned autoradiographs with the aid of ImageJ (version 1.4v).
29 (link) H9c2 cells were used for Western blot analysis and cells were suspended in lysis buffer. Proteins were separated by reducing SDS‐PAGE and transferred to a PVDF membrane. The following antibodies were used for Western blot: p‐AMPK T172 (1:1000, Cell Signaling, #2535), p‐ULK1 S555 (1:1000, Cell Signaling, #5869), GAPDH (1:2000, ThermoFisher, #MA5‐15738), Adiponectin (1:1000, ImmunoDiagnostics, #12010), ATG5‐12 (1:1000, Novus Biologicals, #NB110‐53818), ATG7 (1:1000, Santa Cruz, #sc‐376212), Beclin1 (1:1000, Cell Signaling, #3738), LC3B (1:1000, Cell Signaling, #2775), p62 (1:1000, Cell Signaling, #5114), BAX (1:1000, Cell Signaling, #2772), cleaved Caspase 3 (1:1000, Cell Signaling, #9661), bcl‐2 (1:1000, Cell Signaling, #3489), and β‐actin (1:2000, ThermoFisher, #PA1‐46296) were used. The quantification of signals was performed by densitometry of scanned autoradiographs with the aid of ImageJ (version 1.4v).
