Song Chunjuan, Song Chunjing, Tong Fan
Department of Physiology and Functional Genomics, College of Medicine, University of Florida, Gainesville, Florida, USA.
Department of Experimental Education, Tianjin University of Traditional Chinese Medicine, Tianjin, China.
Cytotherapy. 2014 Oct;16(10):1361-70. doi: 10.1016/j.jcyt.2014.04.006. Epub 2014 Jun 26.
Bone marrow-derived mesenchymal stromal cells (BMSCs) are being extensively investigated as cellular therapeutics for many diseases, including cardiovascular diseases. Although preclinical studies indicated that BMSC transplantation into infarcted hearts improved heart function, there are problems to be resolved, such as the low survival rate of BMSCs during the transplantation process and in the ischemic region with extreme oxidative stress. Autophagy plays pivotal roles in maintaining cellular homeostasis and defending against environmental stresses. However, the precise roles of autophagy in BMSCs under oxidative stress remain largely uncharacterized.
BMSCs were treated with H2O2, and autophagic flux was examined by means of microtubule-associated protein 1A/1B-light chain 3 II/I ratio (LC3 II/I), autophagosome formation and p62 expression. Cytotoxicity and cell death assays were performed after co-treatment of BMSCs by autophagy inhibitor (3-methyladenine) or autophagy activator (rapamycin) together with H2O2.
We show that short exposure (1 h) of BMSCs to H2O2 dramatically elevates autophagic flux (2- to 4-fold), whereas 6-h prolonged oxidative treatment reduces autophagy but enhances caspase-3 and caspase-6-associated apoptosis. Furthermore, we show that pre- and co-treatment with rapamycin ameliorates H2O2-induced caspase-3 and caspase-6 activation and cell toxicity but that 3-methyladenine exacerbates H2O2-induced cell apoptotic cell death.
Our results demonstrate that autophagy is critical for the survival of BMSCs under oxidative conditions. Importantly, we also suggest that the early induction of autophagic flux is possibly a self-defensive mechanism common in oxidant-tolerant cells.
骨髓间充质干细胞（BMSCs）作为多种疾病（包括心血管疾病）的细胞治疗手段正受到广泛研究。尽管临床前研究表明，将BMSCs移植到梗死心脏可改善心脏功能，但仍存在一些问题有待解决，如BMSCs在移植过程中以及在具有极端氧化应激的缺血区域的低存活率。自噬在维持细胞稳态和抵御环境应激中起关键作用。然而，氧化应激下自噬在BMSCs中的精确作用仍 largely未明确。
用H2O2处理BMSCs，通过微管相关蛋白1A/1B轻链3 II/I比值（LC3 II/I）、自噬体形成和p62表达检测自噬通量。在用自噬抑制剂（3-甲基腺嘌呤）或自噬激活剂（雷帕霉素）与H2O2共同处理BMSCs后进行细胞毒性和细胞死亡检测。
我们发现，BMSCs短时间（1小时）暴露于H2O2会显著提高自噬通量（2至4倍），而6小时的延长氧化处理会降低自噬，但会增强与caspase-3和caspase-6相关的凋亡。此外，我们发现雷帕霉素预处理和共同处理可改善H2O2诱导的caspase-3和caspase-6激活以及细胞毒性，但3-甲基腺嘌呤会加剧H2O2诱导的细胞凋亡性细胞死亡。
我们的结果表明，自噬对氧化条件下BMSCs的存活至关重要。重要的是，我们还表明自噬通量的早期诱导可能是耐氧化细胞中常见的一种自我防御机制。
