Lakatos Petra, Szabó Éva, Hegedűs Csaba, Haskó György, Gergely Pál, Bai Péter, Virág László
Department of Medical Chemistry, Medical and Health Science Center, University of Debrecen, Debrecen, Hungary.
Biochim Biophys Acta. 2013 Mar;1833(3):743-51. doi: 10.1016/j.bbamcr.2012.12.003. Epub 2012 Dec 12.
Poly(ADP-ribosyl)ation (PARylation) is a NAD(+)-dependent protein modification carried out by PARP [poly(ADP-ribose) polymerase] enzymes. Here we set out to investigate whether PARylation regulates UVB-induced cell death in primary human keratinocytes. We used the benchmark PARP inhibitor 3-aminobenzamide (3AB) and a more potent and specific inhibitor PJ34 and found that UVB (0.05-0.2J/cm(2)) induced a dose dependent loss of viability that was prevented by 3AB but not by PJ34. Similarly to PJ34, two other new generation PARP inhibitors also failed to protect keratinocytes from UVB-induced loss of viability. Moreover, silencing PARP-1 in HaCaT human keratinocytes sensitized cells to UVB toxicity but 3AB provided protection to both control HaCaT cells and to PARP-1 silenced cells indicating that the photoprotective effect of 3AB is independent of PARP inhibition. Lower UVB doses (0.0125-0.05J/cm(2)) caused inhibition of proliferation of keratinocytes which was prevented by 3AB but augmented by PJ34. UVB-induced keratinocyte death displayed the characteristics of both apoptosis (morphology, caspase activity, DNA fragmentation) and necrosis (morphology, LDH release) with all of these parameters being inhibited by 3AB and apoptotic parameters slightly enhanced by PJ34. UVA also caused apoptotic and necrotic cell death in keratinocytes with 3AB protecting and PJ34 sensitizing cells to UVA-induced toxicity. 3AB prevented UVB-induced mitochondrial membrane depolarization and generation of hydrogen peroxide. In summary, PARylation is a survival mechanism in UV-treated keratinocytes. Moreover, 3-aminobenzamide is photoprotective and acts by a PARP-independent mechanism at a premitochondrial step of phototoxicity.
聚（ADP - 核糖）化（PARylation）是一种由PARP[聚（ADP - 核糖）聚合酶]酶催化的依赖NAD⁺的蛋白质修饰。在此，我们着手研究PARylation是否调节原代人角质形成细胞中紫外线B（UVB）诱导的细胞死亡。我们使用了作为基准的PARP抑制剂3 - 氨基苯甲酰胺（3AB）和一种更有效且特异的抑制剂PJ34，发现UVB（0.05 - 0.2J/cm²）诱导了剂量依赖性的活力丧失，3AB可阻止这种情况，但PJ34不能。与PJ34类似，另外两种新一代PARP抑制剂也未能保护角质形成细胞免受UVB诱导的活力丧失。此外，在HaCaT人角质形成细胞中沉默PARP - 1使细胞对UVB毒性敏感，但3AB对对照HaCaT细胞和PARP - 1沉默细胞均提供保护，这表明3AB的光保护作用独立于PARP抑制。较低剂量的UVB（0.0125 - 0.05J/cm²）导致角质形成细胞增殖受到抑制，3AB可阻止这种情况，但PJ34会增强这种抑制。UVB诱导的角质形成细胞死亡表现出凋亡（形态学、半胱天冬酶活性、DNA片段化）和坏死（形态学、乳酸脱氢酶释放）的特征，所有这些参数均被3AB抑制，而凋亡参数被PJ34轻微增强。紫外线A（UVA）也导致角质形成细胞发生凋亡和坏死性细胞死亡，3AB起到保护作用，而PJ34使细胞对UVA诱导的毒性敏感。3AB可阻止UVB诱导的线粒体膜去极化和过氧化氢的产生。总之，PARylation是紫外线处理的角质形成细胞中的一种存活机制。此外，3 - 氨基苯甲酰胺具有光保护作用，并且在光毒性的线粒体前步骤通过一种不依赖PARP的机制发挥作用。
