Seynhaeve Ann L B, Rens Joost A P, Schipper Debby, Eggermont Alexander M M, Ten Hagen Timo L M
Laboratory of Experimental Surgical Oncology, Department of Surgical Oncology, Erasmus MC, Rotterdam, The Netherlands.
Laboratory of Experimental Surgical Oncology, Department of Surgical Oncology, Erasmus MC, Rotterdam, The Netherlands.
Surgery. 2014 Mar;155(3):545-53. doi: 10.1016/j.surg.2013.10.019. Epub 2013 Oct 15.
We demonstrated previously that the administration of tumor necrosis factor alpha (TNF-α) for the treatment of solid tumors enhanced the response to chemotherapy by augmenting intratumoral drug accumulation. TNF-α changes the integrity of the endothelial cell monolayer in combination with interferon gamma (IFN-γ), which is further enhanced by the addition of peripheral blood mononuclear cells (PBMCs). The improved effect of PBMCs was mostly induced by the endogenous production of interleukin-1beta (IL-1ß) after TNF-α stimulation. In the current study, we demonstrate that exposing endothelial cells to TNF-α and PBMCs mediates the loss of vascular endothelial (VE)-cadherin, an important adherens junction protein for maintaining endothelial integrity, through endogenous IL-1ß. This loss increases permeability of the endothelial layer, thereby explaining the augmented passage of chemotherapeutics into the tumor.
Human umbilical vein endothelial cells were exposed to TNF-α, IFN-γ, PBMCs, or IL-1ß, and the effects on the endothelial integrity were assessed by morphological changes and permeability changes with the use of fluorescein isothiocyanate-labeled bovine serum albumin flux. The loss of VE-cadherin was assessed using immunofluorescence, western blotting, and polymerase chain reaction.
Incubating endothelial cells with TNF-α, IFN-γ, and PBMCs increased cell elongation, gap formation, and subsequently the permeability of fluorescein isothiocyanate-labeled bovine serum albumin compared with control or TNF-α and IFN-γ-treated cells (P < .05). When PBMCs were replaced with IL-1ß, identical changes were observed. These changes in integrity were associated with a loss of VE-cadherin at the membrane.
We conclude that VE-cadherin is lost at the membrane when endothelial cells are exposed to TNF-α, IFN-γ, and PBMCs, which results in loss of integrity. IL-1ß can mimic the effects of PBMCs, indicating a dominant role of endogenously produced IL-1ß in this process.
我们之前已证明，给予肿瘤坏死因子α（TNF-α）用于实体瘤治疗可通过增加瘤内药物蓄积来增强对化疗的反应。TNF-α与干扰素γ（IFN-γ）联合可改变内皮细胞单层的完整性，而添加外周血单个核细胞（PBMCs）可进一步增强这种改变。PBMCs的改善作用主要由TNF-α刺激后内源性白细胞介素-1β（IL-1ß）的产生所诱导。在本研究中，我们证明，使内皮细胞暴露于TNF-α和PBMCs会通过内源性IL-1ß介导血管内皮（VE）-钙黏蛋白的丢失，VE-钙黏蛋白是维持内皮完整性的一种重要黏附连接蛋白。这种丢失会增加内皮层的通透性，从而解释了化疗药物进入肿瘤的增加。
将人脐静脉内皮细胞暴露于TNF-α、IFN-γ、PBMCs或IL-1ß，并通过形态学变化和使用异硫氰酸荧光素标记的牛血清白蛋白通量评估通透性变化来评估对内皮完整性的影响。使用免疫荧光、蛋白质印迹和聚合酶链反应评估VE-钙黏蛋白的丢失。
与对照或TNF-α和IFN-γ处理的细胞相比，用TNF-α、IFN-γ和PBMCs孵育内皮细胞会增加细胞伸长、间隙形成，随后增加异硫氰酸荧光素标记的牛血清白蛋白的通透性（P <.05）。当用IL-1ß替代PBMCs时，观察到相同的变化。这些完整性变化与膜上VE-钙黏蛋白的丢失有关。
我们得出结论，当内皮细胞暴露于TNF-α、IFN-γ和PBMCs时，膜上的VE-钙黏蛋白会丢失，这导致完整性丧失。IL-1ß可模拟PBMCs的作用，表明内源性产生的IL-1ß在此过程中起主导作用。
