Laboratory for Oxidative Stress (LabOS), Division of Molecular Medicine, Rudjer Boskovic Institute, HR-10000 Zagreb, Croatia.
Institute of Molecular Biosciences, Bio TechMed-Graz, University of Graz, 8010 Graz, Austria.
Cells. 2021 Jan 29;10(2):269. doi: 10.3390/cells10020269.
4-Hydroxynonenal (HNE) is a major aldehydic product of lipid peroxidation known to exert several biological effects. Normal and malignant cells of the same origin express different sensitivity to HNE. We used human osteosarcoma cells (HOS) in different stages of differentiation in vitro, showing differences in mitosis, DNA synthesis, and alkaline phosphatase (ALP) staining. Differentiated HOS cells showed decreased proliferation (H-thymidine incorporation), decreased viability (thiazolyl blue tetrazolium bromide-MTT), and increased apoptosis and necrosis (nuclear morphology by staining with 4',6-diamidino-2-phenylindole-DAPI). Differentiated HOS also had less expressed c-MYC, but the same amount of c-FOS (immunocytochemistry). When exposed to HNE, differentiated HOS produced more reactive oxygen species (ROS) in comparison with undifferentiated HOS. To clarify this, we measured HNE metabolism by an HPLC method, total glutathione (GSH), oxidized GSH (ox GSH), glutathione transferase activity (GST), proteasomal activity by enzymatic methods, HNE-protein adducts by genuine ELISA and fatty acid composition by GC-MS in these cell cultures. Differentiated HOS cells had less GSH, lower HNE metabolism, increased formation of HNE-protein adducts, and lower proteasomal activity, in comparison to undifferentiated counterpart cells, while GST and oxGSH were the same. Fatty acids analyzed by GC-MS showed that there is an increase in C20:3 in differentiated HOS while the amount of C20:4 remained the same. The results showed that the cellular machinery responsible for protection against toxicity of HNE was less efficient in differentiated HOS cells. Moreover, differentiated HOS cells contained more C20:3 fatty acid, which might make them more sensitive to free radical-initiated oxidative chain reactions and more vulnerable to the effects of reactive aldehydes such as HNE. We propose that HNE might act as natural promotor of decay of malignant (osteosarcoma) cells in case of their differentiation associated with alteration of the lipid metabolism.
4- 羟壬烯醛 (HNE) 是脂质过氧化的主要醛类产物，已知其具有多种生物学效应。同一来源的正常细胞和恶性细胞对 HNE 的敏感性不同。我们在体外使用不同分化阶段的人骨肉瘤细胞 (HOS)，这些细胞在有丝分裂、DNA 合成和碱性磷酸酶 (ALP) 染色方面表现出差异。分化的 HOS 细胞表现出增殖减少 (H-胸腺嘧啶掺入)、活力降低 (噻唑蓝溴化四唑 MTT)、凋亡和坏死增加 (核形态通过 4'，6-二脒基-2-苯基吲哚-DAPI 染色)。分化的 HOS 细胞也表达较少的 c-MYC，但 c-FOS 量相同 (免疫细胞化学)。当暴露于 HNE 时，与未分化的 HOS 相比，分化的 HOS 产生更多的活性氧 (ROS)。为了阐明这一点，我们使用 HPLC 方法测量 HNE 代谢、总谷胱甘肽 (GSH)、氧化 GSH (ox GSH)、谷胱甘肽转移酶活性 (GST)、通过酶法测定蛋白酶体活性、通过真正的 ELISA 测定 HNE-蛋白质加合物以及通过 GC-MS 测定脂肪酸组成在这些细胞培养物中。与未分化的对应细胞相比，分化的 HOS 细胞的 GSH 较少、HNE 代谢降低、HNE-蛋白质加合物形成增加、蛋白酶体活性降低，而 GST 和 oxGSH 相同。通过 GC-MS 分析的脂肪酸表明，分化的 HOS 中 C20:3 增加，而 C20:4 量保持不变。结果表明，负责保护细胞免受 HNE 毒性的细胞机制在分化的 HOS 细胞中效率较低。此外，分化的 HOS 细胞含有更多的 C20:3 脂肪酸，这可能使它们更容易受到自由基引发的氧化链式反应的影响，并且更容易受到 HNE 等活性醛的影响。我们提出，HNE 可能作为恶性 (骨肉瘤) 细胞分化时脂质代谢改变的自然促进剂发挥作用，导致细胞衰变。
