Cetrullo Silvia, D'Adamo Stefania, Guidotti Serena, Borzì Rosa Maria, Flamigni Flavio
Dipartimento di Scienze Biomediche e Neuromotorie, Università di Bologna, via Irnerio 48, 40126 Bologna, Italy.
Dipartimento di Scienze Biomediche e Neuromotorie, Università di Bologna, via Irnerio 48, 40126 Bologna, Italy; Dipartimento di Scienze Mediche e Chirurgiche, Università di Bologna, via Massarenti 9, 40136 Bologna, Italy.
Biochim Biophys Acta. 2016 Jun;1860(6):1181-91. doi: 10.1016/j.bbagen.2016.03.002. Epub 2016 Mar 3.
Hydroxytyrosol (HT), a major phenolic antioxidant found in olive oil, can afford protection from oxidative stress in several types of non-tumoral cells, including chondrocytes. Autophagy was recently identified as a protective process during osteoarthritis (OA) development and critical for survival of chondrocytes. Therefore we have investigated the possibility to modulate chondrocyte autophagy by HT treatment.
DNA damage and cell death were estimated in human C-28/I2 and primary OA chondrocytes exposed to hydrogen peroxide. Autophagic flux and mitophagy were monitored by measuring levels and location of autophagy markers through western blot, immunostaining and confocal laser microscopy. Late autophagic vacuoles were stained with monodansylcadaverine. The involvement of sirtuin 1 (SIRT-1) was evaluated by immunohistochemistry, western blot and gene silencing with specific siRNA.
HT increases markers of autophagy and protects chondrocytes from DNA damage and cell death induced by oxidative stress. The protective effect requires the deacetylase SIRT-1, which accumulated in the nucleus following HT treatment. In fact silencing of this enzyme prevented HT from promoting the autophagic process and cell survival. Furthermore HT supports autophagy even in a SIRT-1-independent manner, by increasing p62 transcription, required for autophagic degradation of polyubiquitin-containing bodies.
These results support the potential of HT as a chondroprotective nutraceutical compound against OA, not merely for its antioxidant ability, but as an autophagy and SIRT-1 inducer as well.
HT may exert a cytoprotective action by promoting autophagy in cell types that may be damaged in degenerative diseases by oxidative and other stress stimuli.
羟基酪醇（HT）是橄榄油中一种主要的酚类抗氧化剂，能够为包括软骨细胞在内的多种非肿瘤细胞提供抗氧化应激保护。自噬最近被确定为骨关节炎（OA）发展过程中的一种保护机制，对软骨细胞的存活至关重要。因此，我们研究了通过HT处理调节软骨细胞自噬的可能性。
在暴露于过氧化氢的人C-28/I2细胞和原发性OA软骨细胞中评估DNA损伤和细胞死亡情况。通过蛋白质免疫印迹、免疫染色和共聚焦激光显微镜检测自噬标志物的水平和定位，监测自噬流和线粒体自噬。用单丹磺酰尸胺对晚期自噬泡进行染色。通过免疫组织化学、蛋白质免疫印迹和特异性siRNA基因沉默评估沉默调节蛋白1（SIRT-1）的作用。
HT增加自噬标志物水平，并保护软骨细胞免受氧化应激诱导的DNA损伤和细胞死亡。这种保护作用需要脱乙酰酶SIRT-1，HT处理后该酶在细胞核中积累。事实上，沉默该酶可阻止HT促进自噬过程和细胞存活。此外，HT通过增加含多聚泛素体自噬降解所需的p62转录，甚至以不依赖SIRT-1的方式支持自噬。
这些结果支持HT作为一种抗OA的软骨保护营养化合物的潜力，不仅因其抗氧化能力，还因其作为自噬和SIRT-1诱导剂的作用。
HT可能通过促进自噬在可能因氧化和其他应激刺激而在退行性疾病中受损的细胞类型中发挥细胞保护作用。
