Wang Meijing, Wang Lina, Huang Chunyan, Wang I-Wen, Turrentine Mark W
Division of Cardiovascular and Thoracic Surgery, Department of Surgery, Indiana University School of Medicine, Indianapolis, Indiana.
Division of Cardiovascular and Thoracic Surgery, Department of Surgery, Indiana University School of Medicine, Indianapolis, Indiana.
J Surg Res. 2017 Jan;207:155-163. doi: 10.1016/j.jss.2016.08.073. Epub 2016 Aug 31.
Global myocardial ischemia-reperfusion (I/R) occurs during cardiac operations. This I/R injury leads to increased production of tumor necrosis factor α (TNF) instantly and upregulated expression of stromal cell-derived factor 1 α (SDF-1). On the basis of the published data from our laboratory and other groups, locally produced TNF contributes to cardiac dysfunction mainly via binding to its receptor (tumor necrosis factor receptor 1 [TNFR1]), whereas ischemia-induced myocardial SDF-1 mediates cardioprotection. Although TNF has been shown to work as an upstream initiator for induction of other cytokines and chemokines, there is no information regarding the interaction among TNF, TNFRs, and myocardial SDF-1 expression. In this study, given that TNF downregulated SDF-1 in vascular endothelial cells, we therefore hypothesized that TNF would have a negative effect on myocardial SDF-1 production, which is attributable to TNFR-initiated actions.
Using a Langendorff model, isolated male mouse hearts were infused with TNF for 45 min. Male adult mouse hearts from wild type, TNFR1 knockout (TNFR1KO), TNFR2KO, and TNFR1/2KO were subjected to global I/R. H9c2 cells with small interfering RNA transfection were used as an in vitro model. The levels of SDF-1 (protein and messenger RNA) were detected by enzyme-linked immunosorbent assay and quantitative reverse transcription-polymerase chain reaction . Protein kinases of IκB (nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor α) and c-jun N-terminal kinase were also determined using Western blot assay.
TNF infusion downregulated myocardial SDF-1 production in a dose-dependent manner in the hearts. In addition, using TNF significantly decreased SDF-1 expression in cardiomyoblasts (H9c2 cells), which was associated with reduced IκB level. Knockdown of TNFR1 or TNFR2 by small interfering RNAs neutralized TNF-suppressed SDF-1 in H9c2 cells. Furthermore, deletion of TNFR1/2 or TNFR2 increased SDF-1 production in the hearts after I/R.
Our study represents the initial evidence showing that TNF plays an inhibitory role in modulating myocardial SDF-1 production and blockade of TNF signaling by ablation of TNFR1 and TNFR2 genes increased SDF-1 expression in the heart. These data expand on TNF signaling-initiated mechanisms in myocardium, which may lend a more complete understanding of SDF-1 and TNFR-derived actions in hopes of advancing ischemic heart injury treatments.
在心脏手术过程中会发生全身性心肌缺血再灌注（I/R）。这种I/R损伤会立即导致肿瘤坏死因子α（TNF）生成增加以及基质细胞衍生因子1α（SDF-1）表达上调。根据我们实验室和其他研究小组已发表的数据，局部产生的TNF主要通过与其受体（肿瘤坏死因子受体1 [TNFR1]）结合导致心脏功能障碍，而缺血诱导的心肌SDF-1介导心脏保护作用。尽管TNF已被证明可作为诱导其他细胞因子和趋化因子的上游启动因子，但关于TNF、TNFRs和心肌SDF-1表达之间的相互作用尚无相关信息。在本研究中，鉴于TNF可下调血管内皮细胞中的SDF-1，因此我们推测TNF会对心肌SDF-1的产生产生负面影响，这归因于TNFR启动的作用。
使用Langendorff模型，将TNF注入离体雄性小鼠心脏45分钟。对来自野生型、TNFR1基因敲除（TNFR1KO）、TNFR2基因敲除（TNFR2KO）和TNFR1/2基因敲除的成年雄性小鼠心脏进行全身性I/R。将转染小干扰RNA的H9c2细胞用作体外模型。通过酶联免疫吸附测定和定量逆转录-聚合酶链反应检测SDF-1（蛋白质和信使RNA）水平。还使用蛋白质印迹法测定IκB（B细胞中κ轻链多肽基因增强子的核因子抑制剂α）和c-jun氨基末端激酶的蛋白激酶。
注入TNF以剂量依赖的方式下调心脏中心肌SDF-1的产生。此外，使用TNF可显著降低心肌成纤维细胞（H9c2细胞）中SDF-1的表达，这与IκB水平降低有关。通过小干扰RNA敲低TNFR1或TNFR2可中和H9c2细胞中TNF抑制的SDF-1。此外，缺失TNFR1/2或TNFR2可增加I/R后心脏中SDF-1 的产生。
我们的研究提供了初步证据，表明TNF在调节心肌SDF-1产生中起抑制作用，通过切除TNFR1和TNFR2基因阻断TNF信号传导可增加心脏中SDF-1的表达。这些数据扩展了心肌中TNF信号传导启动的机制，这可能有助于更全面地了解SDF-1和TNFR衍生的作用，以期推进缺血性心脏损伤的治疗。
