ROS-Mediated Necroptosis Is Involved in Iron Overload-Induced Osteoblastic Cell Death

Figure 9

RIPK1 and RIPK3 contribute to iron overload-induced ROS. (a, b) The osteoblastic cells were transfected with siControl, siRIPK1, or siRIPK3 for 24 h; then, the expression of RIPK1 and RIPK3 was measured by western blot. Representative western blots of the expression of RIPK1 and RIPK3. Histogram analysis showing the relative protein levels of RIPK1 and RIPK3. Data are presented as the of three independent experiments ( vs. siControl, Student’s -tests). (c, d) The osteoblastic cells were treated with FAC (200 μM) for 120 h with siRIPK1 or siControl. Effect of siRIPK1 on ROS production by H2DCF-DA staining (c). Effect of siRIPK1 on iron overload-induced necrotic cell death by Annexin-V/PI double staining in the osteoblastic cells (d). (e, f) The osteoblastic cells were treated with FAC (200 μM) for 120 h with or without Nec-1 (20 μM). Effect of Nec-1 on ROS production by H2DCF-DA staining (e). Effect of Nec-1 on iron overload-induced necrotic cell death by Annexin-V/PI double staining in the osteoblastic cells (f). (g, h) The osteoblastic cells were treated with FAC (200 μM) for 120 h with siRIPK3 or siControl. Effect of siRIPK3 on ROS production by H2DCF-DA staining (g). Effect of siRIPK3 on iron overload-induced necrotic cell death by Annexin-V/PI double staining in the osteoblastic cells (h). Data are expressed as the from three independent experiments ( vs FAC 200, ANOVA test).

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