Krzykawska-Serda Martyna, Miller Richard C, Elas Martyna, Epel Boris, Barth Eugene D, Maggio Mathew, Halpern Howard J
Department of Biophysics Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, ul. Gronostajowa 7, 30-387, Krakow, Poland.
Department of Radiation and Cellular Oncology, The University of Chicago, Chicago, IL, USA.
Adv Exp Med Biol. 2017;977:319-325. doi: 10.1007/978-3-319-55231-6_42.
Rapid expansion of tumor cells that outpace existing vasculature essential for nutrient and oxygen support as well as waste removal, correlates with profound changes in the microenvironment including angiogenesis, vasodilation, glucose metabolism, and cell cycle perturbations. Since hypoxic cells are up to three times more radioresistant than normoxic cells, identification of hypoxic populations to predict radiotherapeutic outcome is important. The consequences of hypoxia and activated proteins contribute to radioresistant tumors and radiotherapeutic failure. Stereotactic MCa4 tumor tissue biopsies from mouse tumors that were guided by electron paramagnetic resonance (EPR) O imaging were examined for hypoxia-induced proteins. The oxygen broadening of narrow EPR spectral lines or, equivalently, the increase in relaxation rates of electron magnetization, report pO with 1-2 torr resolution in image voxels less than 1 mm. The pO reporter molecule OX063d64 (trityl) was used to acquire the data described here. Trityl appears to be selectively retained in tumors with a half-life of ~30 min. We used an inversion recovery electron spin echo (IRESE) to measure the T1 rate of the trityl inside the tumor bearing leg. We estimate our uncertainty in pO measurement to be 1-3 torr per voxel. Three hypoxic cell biomarkers, hypoxic-induced factor 1-alpha (HIF-1α), vascular endothelial growth factor (VEGF), and carbonic anhydrase IX (CA9), were examined using the ELISA assay. Quantification of these proteins based on results from the ELISA immunoassay kits indicate a strong correlation between EPR pO-identified hypoxic fractions (<10 torr) and HIF-1α, VEGF, and CA9. We clearly demonstrate that hypoxic regions in tumors generate substantial amounts of HIF- 1α, VEGF, and CA9 protein.
肿瘤细胞的快速增殖超过了对营养和氧气支持以及废物清除至关重要的现有脉管系统，这与包括血管生成、血管舒张、葡萄糖代谢和细胞周期紊乱在内的微环境的深刻变化相关。由于缺氧细胞的放射抗性比正常氧合细胞高三倍，识别缺氧群体以预测放射治疗结果很重要。缺氧和活化蛋白的后果导致肿瘤放射抗性和放射治疗失败。对由电子顺磁共振（EPR）氧成像引导的小鼠肿瘤的立体定向MCa4肿瘤组织活检进行缺氧诱导蛋白检查。窄EPR谱线的氧展宽，或者等效地，电子磁化弛豫率的增加，在小于1毫米的图像体素中以1 - 2托的分辨率报告pO。pO报告分子OX063d64（三苯甲基）用于获取此处描述的数据。三苯甲基似乎选择性地保留在肿瘤中，半衰期约为30分钟。我们使用反转恢复电子自旋回波（IRESE）来测量荷瘤腿内三苯甲基的T1速率。我们估计我们在pO测量中的不确定性为每个体素1 - 3托。使用ELISA测定法检测了三种缺氧细胞生物标志物，即缺氧诱导因子1 - α（HIF - 1α）、血管内皮生长因子（VEGF）和碳酸酐酶IX（CA9）。基于ELISA免疫测定试剂盒的结果对这些蛋白质进行定量，表明EPR pO识别的缺氧分数（<10托）与HIF - 1α、VEGF和CA9之间存在很强的相关性。我们清楚地证明肿瘤中的缺氧区域会产生大量的HIF - 1α、VEGF和CA9蛋白。
