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Fig. 7 | Cell & Bioscience

Fig. 7

From: Remote ischemic preconditioning protects against spinal cord ischemia–reperfusion injury in mice by activating NMDAR/AMPK/PGC-1α/SIRT3 signaling

Fig. 7

HKL attenuates OGD/R-induced damage by preventing neuronal apoptosis and oxidative stress via the NMDAR/AMPK/PGC-1α/SIRT3 pathway in neurons. A The chemical structure of HKL. B The CCK8 kit was used to detect the viability of neurons that were treated with different concentrations of HKL with or without OGD/R (n = 6/group). * p < 0.05, compared to the former group. C, D Immunoblot images of SIRT3 in neurons treated with different concentrations of HKL. Relative expression was normalized to the level of the control treatment (n = 3/group). *p < 0.05, **p < 0.01, compared to the former group. E Representative brightfield images showing morphologic changes of OGD/R-treated primary neurons in the presence or absence of HKL. Scale bar = 50 μm. F Quantification of surviving neurons in figure E (n = 3/group). G The percentage of released LDH from OGD/R-treated KO or WT-derived neurons was assessed to determine neuronal injury in the presence or absence of HKL (n = 3/group). H Representative scatter plots of apoptotic neurons induced by OGD/R through PI/Annexin V double labeling in the presence or absence of HKL. I Quantification of apoptotic neurons (Annexin V-positive; n = 3/group) in panel G. J Relative activity of MnSOD, CAT, and GSH content in KO or WT-derived neurons after OGD/R treatment with or without HKL (n = 3/group). K Flow cytometry analysis of ROS production in KO or WT-derived neurons after OGD/R treatment in the presence or absence of HKL. L Quantification of ROS production in neurons of indicated groups (n = 3/group). Normalized to the level of WT neurons that were treated with OGD/R. M, N Western blotting was used to determine the effect of HKL on the expression of NMDAR2B, T-AMPK, p-AMPK, PGC-1α and SIRT3 in neurons that were subjected to OGD/R. Relative expression was normalized to the level of the control treatment (n = 3/group). Statistical analysis: mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001

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