Journal: Bioengineered

Article Title: Knockdown of forkhead box protein P1 alleviates hypoxia reoxygenation injury in H9c2 cells through regulating Pik3ip1/Akt/eNOS and ROS/mPTP pathway.

doi: 10.1080/21655979.2021.2016046

Figure Lengend Snippet: Figure 4. Foxp1 inhibits the PI3K/AKT/eNOS signaling pathway in HR-induced H9c2 cells. (a-e) Representative Western blot to determine the protein involved in survival and apoptotic signaling (including p-PI3K, PI3K, p-Akt, Akt, p-eNOS, eNOS, Bax, and Bcl-2) in HR-induced H9c2 cells, si-NC, si-Foxp1, and si-Foxp1 with LY294002 combination group, respectively. α-Tubulin was treated as an endogenous control (n = 3–5 for each group). (f) NO levels released in H9c2 cells (n = 6 for each group). (g) LDH activity tested in H9c2 cells (n = 6 for each group). LY, LY294002(50uM); L-NAME, NG-Nitro-L-arginine methyl Ester (100 μM); p, phosphorylation; t, total; PI3K, phosphatidylinositol 3-kinase; eNOS, endothelial nitric oxide synthase; NO, nitric oxide. HR, hypoxia/reoxygenation; * P < 0.05; ** P < 0.01; *** P < 0.001. n.s, no significance. One-way ANOVA followed by Tukey post-hoc tests for A to G.

Article Snippet: The addition of LY294002 (a PI3K inhibitor, 50 μM, MedChemExpress, NJ, USA), Atractyloside (ATR, mPTP-specific activator, 20 μM, MedChemExpress, NJ, USA), and NG-Nitro -L-arginine methyl Ester (L-NAME) (a NOS inhibitor, 100 μM, MedChemExpress, NJ, USA) into the culture medium was done 24 h before HR stimulation.

Techniques: Western Blot, Control, Activity Assay, Phospho-proteomics

Journal: Bioengineered

Article Title: Knockdown of forkhead box protein P1 alleviates hypoxia reoxygenation injury in H9c2 cells through regulating Pik3ip1/Akt/eNOS and ROS/mPTP pathway.

doi: 10.1080/21655979.2021.2016046

Figure Lengend Snippet: Figure 5. Downregulation of Foxp1 protected mitochondrial dysfunction in HR-induced H9c2 cells. (a) ROS production in H9c2 cells exposed to HR injury by flow cytometry, with the quantitative data right (n = 4 for each group). (b) JC-1 staining reveals alterations in mitochondrial membrane potential, with the quantitative data down (n = 6 for each group). The red fluorescence indicated normal ΔΨm of the cells. While the green fluorescence indicated that the ΔΨm decreased. (c) Sensitivity of mPTP opening to calcium. (d) Alterations in mPTP opening (A488 OD value of /max-min/). (e) Protein expression of mitochondrial apoptosis signaling (including Cytochrome c, cleaved-Caspase 3, and cleavage of Caspase 9) by Western blot (n = 3–5 for each group). (F to G) Quantitative data of E. α-Tubulin was utilized as an endogenous control. ROS, reactive oxygen species; mPTP, mitochondrial permeability transition pore; ΔΨm, mitochondrial membrane potential; HR, hypoxia/reoxygenation. NAC, N-acetyl-L-cysteine (5 mM); L-NAME, NG-Nitro-L-arginine methyl Ester(100uM); ATR, Atractyloside (20 μM). n = 3–6 for each group. ** P < 0.01; *** P < 0.001. One-way ANOVA followed by Tukey post-hoc tests for A to H.

Article Snippet: The addition of LY294002 (a PI3K inhibitor, 50 μM, MedChemExpress, NJ, USA), Atractyloside (ATR, mPTP-specific activator, 20 μM, MedChemExpress, NJ, USA), and NG-Nitro -L-arginine methyl Ester (L-NAME) (a NOS inhibitor, 100 μM, MedChemExpress, NJ, USA) into the culture medium was done 24 h before HR stimulation.

Techniques: Flow Cytometry, Staining, Membrane, Fluorescence, Expressing, Western Blot, Control, Permeability