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Modulation of radiation-induced damage of human glomerular endothelial cells by SMPDL3B
Alaa Abou Daher1, Marina Francis1, Patrick Azzam1
1Faculty of Medicine, American University of Beirut, Beirut, Lebanon.
Abstract:
The intracellular molecular pathways involved in radiation-induced nephropathy are still poorly understood. Glomerular endothelial cells are key components of the structure and function of the glomerular filtration barrier but little is known about the mechanisms implicated in their injury and repair. The current study establishes the response of immortalized human glomerular endothelial cells (GEnC) to ionizing radiation (IR). We investigated the role of sphingolipids and the lipid-modifying enzyme sphingomyelin phosphodiesterase acid-like 3b (SMPDL3b) in radiation-induced GEnC damage. After delivering a single dose of radiation, long and very-long-chain ceramide species, and the expression levels of SMPDL3b were elevated. In contrast, levels of ceramide-1-phosphate (C1P) dropped in a time-dependent manner although mRNA and protein levels of ceramide kinase (CERK) remained stable. Treatment with C1P or knocking down SMPDL3b partially restored cell survival and conferred radioprotection. We also report a novel role for the NADPH oxidase enzymes (NOXs), namely NOX1, and NOX-derived reactive oxygen species (ROS) in radiation-induced GEnC damage. Subjecting cultured endothelial cells to radiation was associated with increased NOX activity and superoxide anion generation. Silencing NOX1 using NOX1-specific siRNA mitigated radiation-induced oxidative stress and cellular injury. In addition, we report a novel connection between NOX and SMPDL3b. Treatment with the NOX inhibitor, GKT, decreased radiation-induced cellular injury and restored SMPDL3b basal levels of expression. Our findings indicate the importance of SMPDL3b as a potential therapeutic target in radiation-induced kidney damage.
Insights
Radiation damages kidney cells by altering sphingolipids and increasing oxidative stress via NOX1. Targeting SMPDL3b and NOX1 may protect against radiation-induced kidney injury.
Area of Science:
- Cell Biology
- Molecular Biology
- Radiology
Background:
- Radiation-induced nephropathy mechanisms are unclear.
- Glomerular endothelial cells (GEnC) are crucial for kidney filtration but their radiation response is poorly understood.
- Sphingolipids and SMPDL3b's role in GEnC radiation injury requires investigation.
Purpose of the Study:
- To investigate the role of sphingolipids and SMPDL3b in radiation-induced GEnC damage.
- To explore the involvement of NADPH oxidase (NOX) enzymes and reactive oxygen species (ROS) in this process.
- To identify potential therapeutic targets for mitigating radiation-induced kidney damage.
Main Methods:
- Exposed immortalized human GEnC to ionizing radiation (IR).
- Measured sphingolipid species, SMPDL3b, and CERK levels.
- Utilized C1P treatment, SMPDL3b knockdown, and NOX1-specific siRNA.
- Assessed NOX activity, ROS generation, and cell survival.
Main Results:
- IR increased long/very-long-chain ceramides and SMPDL3b expression, while decreasing C1P.
- SMPDL3b knockdown or C1P treatment improved GEnC survival post-IR.
- IR elevated NOX activity and ROS production, with NOX1 silencing mitigating damage.
- NOX inhibition reduced GEnC injury and restored SMPDL3b levels.
Conclusions:
- SMPDL3b and NOX-derived ROS are key mediators of radiation-induced GEnC injury.
- Targeting SMPDL3b or NOX1 offers potential radioprotective strategies for the kidney.
- This study highlights SMPDL3b as a promising therapeutic target for radiation nephropathy.
