Faculty of Medicine, American University of Beirut, Beirut, Lebanon.
Department of Radiation Oncology, Miller School of Medicine, Sylvester Cancer Center, University of Miami, Miami, FL, USA.
FASEB J. 2020 Jun;34(6):7915-7926. doi: 10.1096/fj.201902179R. Epub 2020 Apr 15.
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.
细胞内分子途径参与放射性肾病仍知之甚少。肾小球内皮细胞是肾小球滤过屏障的结构和功能的关键组成部分，但很少有关于它们的损伤和修复的机制。目前的研究建立了永生化人肾小球内皮细胞（GEnC）对电离辐射（IR）的反应。我们研究了鞘脂类和脂质修饰酶神经酰胺磷酸二酯酶酸样 3b（SMPDL3b）在放射性 GEnC 损伤中的作用。在单次剂量的辐射后，长链和超长链神经酰胺物质的表达水平升高。相比之下，神经酰胺-1-磷酸（C1P）的水平呈时间依赖性下降，尽管 ceramide kinase（CERK）的 mRNA 和蛋白水平保持稳定。用 C1P 或敲低 SMPDL3b 处理部分恢复细胞存活并提供放射保护。我们还报告了 NADPH 氧化酶（NOXs），即 NOX1 和 NOX 衍生的活性氧（ROS）在放射性 GEnC 损伤中的新作用。将培养的内皮细胞暴露于辐射会导致 NOX 活性增加和超氧阴离子生成增加。用 NOX1 特异性 siRNA 沉默 NOX1 减轻了辐射诱导的氧化应激和细胞损伤。此外，我们报告了 NOX 和 SMPDL3b 之间的新联系。用 NOX 抑制剂 GKT 处理可减少辐射诱导的细胞损伤并恢复 SMPDL3b 的基础表达水平。我们的研究结果表明 SMPDL3b 作为一种潜在的治疗靶点在放射性肾损伤中的重要性。
