Böhrnsen F, Godek F, Kiesel J, Kramer F J, Brockmeyer P, Schliephake H
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany.
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany; Department of Preventive Dentistry, Periodontology and Cardiology, University Medical Center Göttingen, Germany.
J Craniomaxillofac Surg. 2017 May;45(5):731-740. doi: 10.1016/j.jcms.2017.02.009. Epub 2017 Feb 17.
TGF-β1 signaling modulates epithelial mesenchymal transitions (EMT) of head and neck squamous cell carcinoma (HNSCC). Bone marrow mesenchymal stromal cells (BMSC) are able to exert a regulating influence on the expression of markers of EMT in HNSCC cells. It was thus the aim of this study to test the hypothesis that TGF-β1 modulates the interactions of tumor transition between BMSCs and HNSCC, affecting the expression of E-cadherin, Vimentin, Snail, Twist, MMP14 and beta-catenin. Furthermore, we analyzed alterations in the AKT-signaling of tumor and stroma cells.
BMSCs were isolated from iliac bone marrow aspirates and co-cultured in trans-well permeable membrane wells with tumor cells of the established HNSCC cell line PCI-13. Following the induction with TGF-β1 under serum free conditions the expression of Vimentin and E-Cadherin was assessed via immunofluorescence. A quantitative RT-PCR analysis of tumor transition markers E-cadherin, Vimentin, Snail, Twist, MMP14 and beta-catenin was performed. Changes in AKT-Signaling were identified via protein analysis.
In non-induced co-cultures, BMSC were able to suppress Vimentin in PCI-13 as a marker of tumor transition. In TGF-β1 induced co-cultures PCI-13 significantly increased the expression of Vimentin, Twist, Snail, MMP14, GSK3a, PRAS40, 4E-BP1, and AMPKa compared to monolayer controls. TGF-β1 co-cultured BMSC demonstrated a significant increase of Snail, PRAS40, mTOR, GSK3a/b, Bad, PDK1 and 4E-BP1.
TGF-β1 was able to attenuate the modulating influence of BMSC in co-culture and drive the co-culture towards a progressive tumor transition, affecting the expression of markers of EMT, AKT-Signaling and proliferative checkpoints.
转化生长因子-β1（TGF-β1）信号传导调节头颈部鳞状细胞癌（HNSCC）的上皮-间质转化（EMT）。骨髓间充质基质细胞（BMSC）能够对头颈部鳞状细胞癌细胞中EMT标志物的表达产生调节作用。因此，本研究旨在验证以下假设：TGF-β1调节BMSC与HNSCC之间的肿瘤转化相互作用，影响E-钙黏蛋白、波形蛋白、Snail、Twist、基质金属蛋白酶14（MMP14）和β-连环蛋白的表达。此外，我们分析了肿瘤细胞和基质细胞中AKT信号传导的变化。
从髂骨髓穿刺物中分离出BMSC，并与已建立的HNSCC细胞系PCI-13的肿瘤细胞在Transwell可渗透膜孔中共培养。在无血清条件下用TGF-β1诱导后，通过免疫荧光评估波形蛋白和E-钙黏蛋白的表达。对肿瘤转化标志物E-钙黏蛋白、波形蛋白、Snail、Twist、MMP14和β-连环蛋白进行定量逆转录聚合酶链反应（RT-PCR）分析。通过蛋白质分析确定AKT信号传导的变化。
在未诱导的共培养中，BMSC能够抑制PCI-13中作为肿瘤转化标志物的波形蛋白。与单层对照相比，在TGF-β1诱导的共培养中，PCI-13中波形蛋白、Twist、Snail、MMP14、糖原合成酶激酶3α（GSK3α）、脯氨酸丰富的AKT底物40（PRAS40）、真核起始因子4E结合蛋白1（4E-BP1）和AMP活化蛋白激酶α（AMPKα）的表达显著增加。TGF-β1共培养的BMSC中Snail、PRAS40、雷帕霉素靶蛋白（mTOR）、GSK3α/β、凋亡前体蛋白Bad、磷酸肌醇依赖性蛋白激酶1（PDK1）和4E-BP1显著增加。
TGF-β1能够减弱共培养中BMSC的调节作用，并促使共培养向进行性肿瘤转化发展，影响EMT标志物的表达、AKT信号传导和增殖检查点。
