Na Hye-Kyung, Yang Hongkyung, Surh Young-Joon
Department of Food Science and Biotechnology, College of Knowledge-Based Services Engineering, Sungshin Women's University, Seoul, Korea.
Department of Future Applied Sciences, College of Natural Sciences, Sungshin Women's University, Seoul, Korea.
J Cancer Prev. 2020 Jun 30;25(2):100-110. doi: 10.15430/JCP.2020.25.2.100.
15-Deoxy-Δ-prostaglandin J (15d-PGJ), an endogenous ligand for PPARγ, has differential effects on cancer cell proliferation and survival depending on the dose and the type of cells. In the present study, we have investigated the effects of 15d-PGJ on apoptosis of the Ha- transformed human breast epithelial (MCF10A-) cells. When MCF10A- cells were treated with 15d-PGJ (10 μM) for 24 hours, they underwent apoptosis as evidenced by characteristic morphological features, an increased proportion of sub-G/G cell population, a typical pattern of annexin V/propidium iodide staining, perturbation of mitochondrial transmembrane potential (Δψ), and cleavage of caspase-3 and its substrate PARP. A pan-caspase inhibitor, Z-Val-Ala-Asp (OCH)-fluoromethyl ketone attenuated cytotoxicity and proteolytic cleavage of caspase-3 induced by 15d-PGJ. The 15d-PGJ-induced apoptosis was accompanied by enhanced intracellular accumulation of reactive oxygen species (ROS), which was abolished by the antioxidant -acetyl-L-cysteine (NAC). 15d-PGJ inhibited the DNA binding activity of NF-κB which was associated with inhibition of expression and catalytic activity of IκB kinase β (IKKβ). 15d-PGJ-mediated inhibition of IKKβ and nuclear translocation of phospho-p65 was blocked by NAC treatment. 9,10-Dihydro-PGJ, a non-electrophilic analogue of 15d-PGJ, failed to produce ROS, to inhibit NF-κB DNA binding, and to induce apoptosis, suggesting that the electrophilic α,β-unsaturated carbonyl group of 15d-PGJ is essential for its pro-apoptotic activity. 15d-PGJ-induced inactivation of IKKβ was also attributable to its covalent thiol modification at the cysteine 179 residue of IKKβ. Based on these findings, we propose that 15d-PGJ inactivates IKKβ-ΝF-κB signaling through oxidative or covalent modification of IKKβ, thereby inducing apoptosis in Ha- transformed human breast epithelial cells.
15-脱氧-Δ-前列腺素J（15d-PGJ）是过氧化物酶体增殖物激活受体γ（PPARγ）的内源性配体，根据剂量和细胞类型的不同，对癌细胞的增殖和存活有不同影响。在本研究中，我们研究了15d-PGJ对Ha-转化的人乳腺上皮（MCF10A-）细胞凋亡的影响。当MCF10A-细胞用15d-PGJ（10μM）处理24小时后，它们发生了凋亡，特征性形态学特征、亚G1/G1期细胞群体比例增加、膜联蛋白V/碘化丙啶染色的典型模式、线粒体跨膜电位（Δψ）的扰动以及半胱天冬酶-3及其底物聚（ADP-核糖）聚合酶（PARP）的裂解均证明了这一点。一种泛半胱天冬酶抑制剂Z-缬氨酸-丙氨酸-天冬氨酸（OCH）-氟甲基酮减弱了15d-PGJ诱导的细胞毒性和半胱天冬酶-3的蛋白水解裂解。15d-PGJ诱导的凋亡伴随着细胞内活性氧（ROS）积累的增强，而抗氧化剂N-乙酰-L-半胱氨酸（NAC）可消除这种增强。15d-PGJ抑制核因子κB（NF-κB）的DNA结合活性，这与抑制IκB激酶β（IKKβ）的表达和催化活性有关。NAC处理可阻断15d-PGJ介导的IKKβ抑制和磷酸化p65的核转位。9,10-二氢-PGJ是15d-PGJ的非亲电类似物，不能产生活性氧、抑制NF-κB DNA结合以及诱导凋亡，这表明15d-PGJ的亲电α,β-不饱和羰基对其促凋亡活性至关重要。15d-PGJ诱导的IKKβ失活也归因于其对IKKβ第179位半胱氨酸残基的共价硫醇修饰。基于这些发现，我们提出15d-PGJ通过对IKKβ的氧化或共价修饰使IKKβ-NF-κB信号失活，从而诱导Ha-转化的人乳腺上皮细胞凋亡。
