Division of Immunology and Immunotherapy, Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Radiation Oncology, University Clinic, University of Navarra, Pamplona, Spain.
Division of Immunology and Immunotherapy, Center for Applied Medical Research, University of Navarra, Pamplona, Spain.
Int J Radiat Oncol Biol Phys. 2017 Feb 1;97(2):389-400. doi: 10.1016/j.ijrobp.2016.10.043. Epub 2016 Nov 7.
PURPOSE/OBJECTIVES: The goal of this study was to assess the effects of ionizing radiation on the expression of the integrin ligands ICAM-1 and VCAM that control leucocyte transit by lymphatic endothelial cells.
MATERIALS/METHODS: Confluent monolayers of primary human lymphatic endothelial cells (LEC) were irradiated with single dose of 2, 5, 10 or 20 Gy, with 6 MeV-x-rays using a Linear-Accelerator. ICAM-1 and VCAM expression was determined by flow cytometry. Human tissue specimens received a single dose of 20 Gy with 15 MeV-x-rays. MC38, B16-OVA or B16-VEGF-C tumors grown in C57BL/6 mice were irradiated with single dose of 20Gy using a Linear-Accelerator fitted with a 10mm Radiosurgery collimator. Clinical samples were obtained from patients previous and 4 weeks after complete standard radiotherapy. ICAM-1 and VCAM expression was detected in all tissue specimens by confocal microscopy. To understand the role of TGFβ in this process anti-TGFβ blocking mAb were injected i.p. 30min before radiotherapy. Cell adhesion to irradiated LEC was analyzed in adhesion experiments performed in the presence or in the absence of anti- TGFβ and /or anti-ICAM1 blocking mAb.
We demonstrate that lymphatic endothelial cells in tumor samples experience induction of surface ICAM-1 and VCAM when exposed to ionizing radiation in a dose- and time-dependent manner. These effects can be recapitulated in cultured LEC, and are in part mediated by TGFβ. These data are consistent with increases in ICAM-1 and VCAM expression on LYVE-1+ endothelial cells in freshly explanted human tumor tissue and in mouse transplanted tumors after radiotherapy. Finally, ICAM-1 and VCAM expression accounts for enhanced adherence of human T lymphocytes to irradiated LEC.
Our results show induction of ICAM-1 and VCAM on LVs in irradiated lesions and offer a starting point for elucidating the biological and therapeutic implications of targeting leukocyte traffic in combination to immunotherapy.
本研究旨在评估电离辐射对整合素配体 ICAM-1 和 VCAM 表达的影响，这些配体控制白细胞通过淋巴管内皮细胞的迁移。
用 6 MeV-x 射线，通过线性加速器对单层汇合的原代人淋巴管内皮细胞（LEC）进行单次 2、5、10 或 20 Gy 照射。通过流式细胞术测定 ICAM-1 和 VCAM 表达。用 15 MeV-x 射线对人体组织标本进行单次 20 Gy 照射。将 MC38、B16-OVA 或 B16-VEGF-C 肿瘤植入 C57BL/6 小鼠，用配备 10mm 放射外科准直器的线性加速器进行单次 20 Gy 照射。临床标本取自患者在完全标准放疗前和 4 周后。通过共聚焦显微镜检测所有组织标本中的 ICAM-1 和 VCAM 表达。为了了解 TGFβ 在这个过程中的作用，在放疗前 30 分钟通过腹腔注射抗 TGFβ 阻断 mAb。在存在或不存在抗 TGFβ 和/或抗 ICAM1 阻断 mAb 的情况下，在粘附实验中分析细胞对照射的 LEC 的粘附。
我们证明，淋巴管内皮细胞在肿瘤样本中暴露于电离辐射时，会以剂量和时间依赖的方式诱导表面 ICAM-1 和 VCAM 的表达。这些效应可以在培养的 LEC 中重现，部分是由 TGFβ 介导的。这些数据与在新鲜离体人肿瘤组织和放疗后小鼠移植瘤中 LYVE-1+内皮细胞中 ICAM-1 和 VCAM 表达增加一致。最后，ICAM-1 和 VCAM 的表达解释了人 T 淋巴细胞对照射的 LEC 的粘附增强。
我们的结果表明，在照射的病变中诱导了 LVs 中的 ICAM-1 和 VCAM，为阐明靶向白细胞迁移与免疫治疗联合的生物学和治疗意义提供了起点。
