Italian Liver Foundation (FIF), Bldg Q-AREA Science Park Basovizza, SS14 Km 163,5, 34149 Trieste, Italy.
Department of Life Sciences, University of Trieste, 34127 Trieste, Italy.
Int J Mol Sci. 2020 Oct 30;21(21):8101. doi: 10.3390/ijms21218101.
Severe hyperbilirubinemia can cause permanent neurological damage in particular in neonates, whereas mildly elevated serum bilirubin protects from various oxidative stress-mediated diseases. The present work aimed to establish the intracellular unconjugated bilirubin concentrations (iUCB) thresholds differentiating between anti- and pro-oxidant effects.
Hepatic (HepG2), heart endothelial (H5V), kidney tubular (HK2) and neuronal (SH-SY5Y) cell lines were exposed to increasing concentration of bilirubin. iUCB, cytotoxicity, intracellular reactive oxygen species (ROS) concentrations, and antioxidant capacity (50% efficacy concentration (EC)) were determined.
Exposure of SH-SY5Y to UCB concentration > 3.6 µM (iUCB of 25 ng/mg) and >15 µM in H5V and HK2 cells (iUCB of 40 ng/mg) increased intracellular ROS production ( < 0.05). EC of the antioxidant activity was 21 µM (iUCB between 5.4 and 21 ng/mg) in HepG2 cells, 0.68 µM (iUCB between 3.3 and 7.5 ng/mg) in SH-SY5Y cells, 2.4 µM (iUCB between 3 and 6.7 ng/mg) in HK2 cells, and 4 µM (iUCB between 4.7 and 7.5 ng/mg) in H5V cells.
In all the cell lines studied, iUCB of around 7 ng/mg protein had antioxidant activities, while iUCB > 25 ng/mg protein resulted in a prooxidant and cytotoxic effects. UCB metabolism was found to be cell-specific resulting in different iUCB.
严重的高胆红素血症会对新生儿等造成永久性神经损伤，而轻度升高的血清胆红素可预防多种氧化应激介导的疾病。本研究旨在建立区分胆红素抗/促氧化作用的细胞内未结合胆红素浓度（iUCB）阈值。
用不同浓度胆红素处理 HepG2 肝细胞、H5V 心脏内皮细胞、HK2 肾小管细胞和 SH-SY5Y 神经元细胞系。检测细胞内未结合胆红素浓度（iUCB）、细胞毒性、细胞内活性氧（ROS）浓度和抗氧化能力（50%有效浓度（EC））。
SH-SY5Y 细胞暴露于 UCB 浓度>3.6 μM（iUCB 为 25ng/mg）和 H5V 和 HK2 细胞>15 μM（iUCB 为 40ng/mg）时，细胞内 ROS 生成增加（<0.05）。HepG2 细胞的抗氧化活性 EC 为 21 μM（iUCB 在 5.4 至 21ng/mg 之间），SH-SY5Y 细胞为 0.68 μM（iUCB 在 3.3 至 7.5ng/mg 之间），HK2 细胞为 2.4 μM（iUCB 在 3 至 6.7ng/mg 之间），H5V 细胞为 4 μM（iUCB 在 4.7 至 7.5ng/mg 之间）。
在所研究的所有细胞系中，iUCB 约为 7ng/mg 蛋白时具有抗氧化活性，而 iUCB>25ng/mg 蛋白时则具有促氧化和细胞毒性作用。发现 UCB 代谢具有细胞特异性，导致不同的 iUCB。
