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Abstract: PO0292

Oxidative Stress and Heme Metabolism in Red Blood Cells of Hemodialysis Patients

Session Information

Category: Anemia and Iron Metabolism

  • 200 Anemia and Iron Metabolism

Authors

  • Dias, Gabriela Ferreira, Renal Research Institute, New York, New York, United States
  • Andrade, Gabriela Bohnen, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
  • Grobe, Nadja, Renal Research Institute, New York, New York, United States
  • Tao, Xia, Renal Research Institute, New York, New York, United States
  • Pecoits-Filho, Roberto, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
  • Kotanko, Peter, Renal Research Institute, New York, New York, United States
  • Moreno-Amaral, Andrea Novais, Pontificia Universidade Catolica do Parana, Curitiba, PR, Brazil
Background

We have previously described that indoxyl sulfate promotes red blood cells (RBC) ROS generation through organic anion transporter 2 as well as NADPH oxidase activity-dependent and GSH-independent mechanisms (Dias et al., 2018). However, there is little information regarding pathways of antioxidant balance to protect RBC from extensive oxidative stress that occurs during hemodialysis (HD). Intracellular free heme is degraded by Heme Oxygenase 1 (HO-1), which is regarded as the major cytoprotective enzyme (Maines, 1988; Gozzelino et al., 2010). In the current study, we assessed HO-1 activity and ROS production in RBC from healthy subjects and hemodialysis (HD) patients before and after HD.

Methods

Blood was drawn from 6 healthy individuals (CON-RBC) and 6 HD patients (HD-RBC) before (pre/HD-RBC) and after high flux HD (post/HD-RBC). Isolated RBC were stained with DCFH-DA (Abcam) for ROS measurements. To quantify HO-1, RBC were incubated with anti-HO-1 antibody (Abcam) and m-IgGκ BP-CFL 488 (Santa Cruz Biotechnology) as a secondary antibody. Samples were analyzed by flow cytometry.

Results

Our results show a 4-fold increase in ROS levels in pre/HD-RBC compared to CON-RBC. ROS levels were even further increased by 1.65-fold after HD treatment in post/HD-RBC (Figure 1). Both pre/HD-RBC and post/HD-RBC showed a similarly significant increase of 3.3-fold in HO-1 compared to CON-RBC. (Figure 1).

Conclusion

High levels of HO-1 may represent a defense against oxidative stress that occurs in ESKD and particularly during the HD session. Further research is needed to evaluate whether HO-1 overexpression could accelerate heme degradation and contribute to renal anemia.

Figure 1. Reactive oxygen species (ROS) and Heme Oxygenase-1 levels in red blood cells (RBC) from healthy individuals (CON-RBC) and patients pre (Pre/HD-RBC) and post (Post/HD-RBC) hemodialysis. Data are shown as median and interquartile range of the Mean Fluorescence Intensity (MFI). ***p<0.001 and **p<0.01 represent the difference compared to CON-RBC. &p<0.001 represents the difference between pre- and post/HD-RBC.

Funding

  • Private Foundation Support