Fig 1: Upregulation of NRF2 expression by APPL1 overexpression after H/R. Expression levels of NRF2 and HO-1 in H/R-induced H9c2 cells were detected using western blotting. The experimental data are presented as the mean ± SD. *P<0.05, **P<0.01 and ***P<0.001. H/R, hypoxia-reoxygenation; NRF2, nuclear factor erythroid 2-related factor 2; HO-1, heme oxygenase-1; APPL1, adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1; ov, overexpression; NC, negative control.
Fig 2: Suppression of apoptosis by APPL1 overexpression. (A) Quantification of apoptotic rate of H9c2 cells induced by H/R (B) as determined by performing TUNEL staining. (C) Detection of cleaved caspase-3, cleaved PARP and Bcl2 expression levels. The experimental data are presented as the mean ± SD. *P<0.05, **P<0.01 and ***P<0.001. APPL1, adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1; H/R, hypoxia-reoxygenation; PARP, poly (ADP-ribose) polymerase 1; ov, overexpression; NC, negative control.
Fig 3: Decrease in ROS levels induced by APPL1 overexpression is mediated via AMP-activated protein kinase. (A) Quantification of ROS production (B) as determined by ROS staining. The experimental data are presented as the mean ± SD. ***P<0.001. ROS, reactive oxygen species; APPL1, adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1; H/R, hypoxia-reoxygenation; ov, overexpression; NC, negative control.
Fig 4: APPL1 overexpression increases cell viability after H/R and compound C reduces it via the AMP-activated protein kinase signaling pathway. (A) Cell viability and (B) LDH were detected using Cell Counting Kit-8 and LDH assays, respectively. The experimental data are presented as the mean ± SD. *P<0.05 and ***P<0.001. APPL1, adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1; LDH, lactate dehydrogenase; H/R, hypoxia-reoxygenation; ov, overexpression; NC, negative control.
Fig 5: APPL1 overexpression increases the phosphorylation levels of LKB1, AMPK and ACC, as well as SOD levels. (A) mRNA and (B) protein expression levels of APPL1 in H/R-induced H9c2 cells. (C) mRNA and (D) protein expression levels of APPL1 in H9c2 cells with or without ov-APPL1 transfection. (E) mRNA and (F) protein expression levels of APPL1 in H/R-induced H9c2 cells overexpressing APPL1. Determination of the expression levels of (G) p-LKB1, LKB1, p-AMPK, AMPK, p-ACC, ACC, (H) SOD2 and SOD3. (I) Determination of the activity levels of SOD. The experimental data are presented as the mean ± SD. *P<0.05, **P<0.01 and ***P<0.001. APPL1, adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1; ACC, acetyl-CoA carboxylase; H/R, hypoxia-reoxygenation; SOD, superoxide dismutase; LKB1, liver kinase B1; AMPK, AMP-activated protein kinase; p, phosphorylated; ov, overexpression; NC, negative control.
Supplier Page from Abcam for Anti-APPL antibody [EPR13569] - BSA and Azide free