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Abstract: FR-PO534

Activation of BKα Channel Prevented the High Glucose-Induced Oxidative Stress in Human Proximal Tubular Cells

Session Information

Category: Fluid, Electrolytes, and Acid-Base Disorders

  • 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic

Authors

  • Luo, Xiaomao, Emory University School of Medicine, Atlanta, Georgia, United States
  • Bian, Shuyang, Emory University School of Medicine, Atlanta, Georgia, United States
  • Huang, Yijin, Emory University School of Medicine, Atlanta, Georgia, United States
  • Hassounah, Faten, Emory University School of Medicine, Atlanta, Georgia, United States
  • Yue, Qiang, Emory University School of Medicine, Atlanta, Georgia, United States
  • Wang, Xiaonan H., Emory University School of Medicine, Atlanta, Georgia, United States
  • Cai, Hui, Emory University School of Medicine, Atlanta, Georgia, United States
Background

Hyperglycemia stimulates the production of reactive oxygen species (ROS) leading to oxidative stress. Oxidative stress could cause inflammation or fibrosis. Our previous study found that activation of the large conductance, calcium-activated potassium channels (BK channels) attenuated renal fibrosis in mice. We hypothesized that upregulation BK channel activity would suppress the high glucose (HG)-induced oxidative stress.

Methods


Human proximal tubular epithelial cell (HK-2) was cultured in DMEM/F12 medium. HEK-BKα cells (HEK293 stably expressing BKα) were maintained in DMEM with G418. Cells were cultured in the low glucose DMED (5.5mM glucose) for 24 hours before treatments. NS1916 (Sigma) was used to activate BKα channel. Two ROS products were measured: 1-hydrogen peroxide (H2O2) was tested by Amplex Red Hydrogen Peroxide Kit (cat # A22188, Molecular Probes): 2- DHE (Dihydroethidium) Assay (Abcam ab236206) was used to detect superoxide. Superoxide Dismutase (SOD), a defense of oxidative enzyme, was measured using colorimetric activity kit (Invitrogen EIASODC). Single channel recordings were used to analyze the activity of BKα channel.

Results

Using single channel recordings we showed that high glucose (HS, 25 mM D-glucose) inhibited BKα channel activity in HEK-BKα cells. As an osmolarity control, 25 mM mannitol does not change the BKα activity. To determine the effect of BKα activation on HG-induced ROS production HK2 cells was treated with HS (25mM) with or without NS1916 (BKα channel opener). H2O2 and superoxide productions were significantly increased with high glucose treatment, whereas NS1916 (20mM) prevented the HG-induced upregulation of H2O2 and ROS. In addition, HS treatment (25 mM) significantly decreased SOD activity in HK2 cells, whereas activation of BKα activity with NS1916 reversed the decreased SOD activity induced by HS.

Conclusion

High glucose inhibited BKα activity while increasing H2O2 and superoxide productions in HK2 cells. Activation of BKα channel activity attenuated HS-induced the ROS production. In addition, BKα reversed HS-induced down-regulation of SOD. These results suggest that activation of BKα channel has novel roles in preventing oxidative stress.

Funding

  • Veterans Affairs Support