Atherogenic L5 LDL induces cardiomyocyte apoptosis and inhibits KATP channels through CaMKII activation

Figures

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  Fig. 1. Increased apoptosis and decreased cell viability in NRCMs treated with L5. a, Early apoptosis of NRCMs treated with L5, shown by representative images of Hoechst (blue, nuclei), Annexin V-Alexa 488 (green), and PI (red) staining. b, Dose-dependent curve showing the effects of increasing concentrations of L5 and L1 on the percentage of cells staining positive for Annexin V (n = 6 per group). c, Relative apoptosis levels of NRCMs after the indicated treatment, determined by using a caspase-3/7 activity assay (n = 7 per group). d, Cardiomyocyte viability was analyzed after the indicated treatment by using an MTT kit, n = 6 per group. *P < 0.05, **P < 0.01 vs. the PBS-treated group

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  Fig. 2. Prolonged APD75 in NRCMs after L5 treatment. a, Representative traces of the APD75 in NRCMs treated with L5. b, Comparison of the APD75 at 0.1 Hz, 1 Hz, and 2.5 Hz. n = 5 per group. *P < 0.05 vs. baseline

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  Fig. 3. Decreased IKATP density in NRCMs treated with L5. a, Representative traces of IKATP in NRCMs treated with Pin-baseline (20 μM pinacidil, IKATP opener), Pin+L5 (7.5 μg/mL), Pin-washout, and Pin+Gli (100 μM glibenclamide, IKATP blocker). b, Time-course analysis showing the effect of L5 on IKATP. c, The perfusion of NRCMs with L5 reduced IKATP, which partially recovered after 5 min of washout (n = 5 per group, **P < 0.01,*P < 0.05 vs. Pin-baseline, #P < 0.05 vs. Pin+L5). d, The overnight incubation of NRCMs with L5 decreased the expression level of Kir6.2, but no change was seen after overnight incubation with L1, n = 3 per group. *P < 0.05 vs. PBS

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  Fig. 4. Increased CaMKII activity and ROS production in NRCMs treated with L5. a, L5 increased CaMKII activity in NRCMs, whereas L1 did not (n = 8 per group). b, L5 increased the expression level of NOX2/gp91phox in NRCMs, whereas L1 did not (n = 3 per group). c, L5 induced CaMKII phosphorylation, and the CaMKII inhibitor KN93 decreased L5-induced CaMKII phosphorylation. The ROS scavenger Mn (III) TBAP did not affect L5-induced CaMKII phosphorylation (n = 3 per group). *P < 0.05, **P < 0.01 vs. PBS

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  Fig. 5. The attenuation of L5’s effect on NRCMs by inhibitors of ROS and CaMKII. a, The ROS scavenger Mn (III) TBAP and CaMKII inhibitor KN93 prevented the L5-induced decrease in the cell viability of NRCMs. b, Mn (III) TBAP and KN93 attenuated the increased apoptosis of NRCMs caused by L5. n = 6 per group. **P < 0.01 vs. L5