Filipa Gil Marques1, Esmeralda Poli2, João Malaquias3, Tânia Carvalho4, Ana Portêlo5, Afonso Ramires3, Fernando Aldeia3, Ruy Miguel Ribeiro6, Emília Vitorino7, Isabel Diegues2, Luís Costa8, João Coutinho3, Filomena Pina2, Marc Mareel9, Susana Constantino Rosa Santos10. 1. Angiogenesis Laboratory, Centro Cardiovascular da Universidade de Lisboa, Faculdade de Medicina, Universidade de Lisboa, Portugal. 2. Radiotherapy Service, Centro Hospitalar Universitário Lisboa Norte, Portugal. 3. Department of Surgery, Centro Hospitalar Universitário Lisboa Norte, Portugal. 4. Histology and Comparative Pathology Laboratory, Instituto de Medicina Molecular; Faculdade de Medicina, Universidade de Lisboa, Portugal. 5. Instituto de Medicina Molecular; Faculdade de Medicina, Universidade de Lisboa, Portugal. 6. Biomathematics Laboratory, Faculdade de Medicina, Universidade de Lisboa, Portugal. 7. Department of Pathology, Centro Hospitalar Universitário Lisboa Norte, Portugal. 8. Oncology Department, Centro Hospitalar Universitário Lisboa Norte; Instituto de Medicina Molecular; Faculdade de Medicina, Universidade de Lisboa, Portugal. 9. Department of Radiotherapy and Laboratory of Experimental Cancer Research, Ghent University Hospital, Belgium. 10. Angiogenesis Laboratory, Centro Cardiovascular da Universidade de Lisboa, Faculdade de Medicina, Universidade de Lisboa, Portugal. Electronic address: sconstantino@medicina.ulisboa.pt.
Abstract
PURPOSE: During radiotherapy the peritumoral tissues are daily exposed to subtherapeutic doses of ionizing radiation. Herein, the biological and molecular effects of doses lower than 0.8 Gy per fraction (LDIR), previously described as angiogenesis inducers, were assessed in human peritumoral tissues. MATERIAL AND METHODS: Paired biopsies of preperitoneal adipose tissue were surgically collected from 16 patients diagnosed with locally advanced rectal cancer who underwent neo-adjuvant radiotherapy. One of the biopsies is located in the vicinity of the region where the tumor received the prescribed dose of radiation, and thus exposed to LDIR; the other specimen, outside all beam apertures, was used as an internal calibrator (IC). Microvessel density (MDV) was quantified by immunohistochemistry and the expression of several pro-angiogenic genes was assessed by quantitative RT-PCR using exclusively endothelial cells (ECs) isolated by laser capture microdissection microscopy. RESULTS: LDIR activated peritumoral ECs by significantly up-regulating the expression of several pro-angiogenic genes such as VEGFR1, VEGFR2, ANGPT2, TGFB2, VWF, FGF2, HGF and PDGFC. Accordingly, the MVD was significantly increased in peritumoral tissues exposed to LDIR, compared to the IC. The patients that yielded a larger pro-angiogenic response, also showed the highest MVD. CONCLUSIONS: LDIR activate ECs in peritumoral tissues that are associated with increased MVD. Although the technological advances in radiotherapy have contributed to reduce the damage to healthy tissues over the past years, the anatomical regions receiving LDIR should be taken into account in the treatment plan report for patient follow-up and in future studies to correlate these doses with tumor dissemination.
PURPOSE: During radiotherapy the peritumoral tissues are daily exposed to subtherapeutic doses of ionizing radiation. Herein, the biological and molecular effects of doses lower than 0.8 Gy per fraction (LDIR), previously described as angiogenesis inducers, were assessed in human peritumoral tissues. MATERIAL AND METHODS: Paired biopsies of preperitoneal adipose tissue were surgically collected from 16 patients diagnosed with locally advanced rectal cancer who underwent neo-adjuvant radiotherapy. One of the biopsies is located in the vicinity of the region where the tumor received the prescribed dose of radiation, and thus exposed to LDIR; the other specimen, outside all beam apertures, was used as an internal calibrator (IC). Microvessel density (MDV) was quantified by immunohistochemistry and the expression of several pro-angiogenic genes was assessed by quantitative RT-PCR using exclusively endothelial cells (ECs) isolated by laser capture microdissection microscopy. RESULTS: LDIR activated peritumoral ECs by significantly up-regulating the expression of several pro-angiogenic genes such as VEGFR1, VEGFR2, ANGPT2, TGFB2, VWF, FGF2, HGF and PDGFC. Accordingly, the MVD was significantly increased in peritumoral tissues exposed to LDIR, compared to the IC. The patients that yielded a larger pro-angiogenic response, also showed the highest MVD. CONCLUSIONS: LDIR activate ECs in peritumoral tissues that are associated with increased MVD. Although the technological advances in radiotherapy have contributed to reduce the damage to healthy tissues over the past years, the anatomical regions receiving LDIR should be taken into account in the treatment plan report for patient follow-up and in future studies to correlate these doses with tumor dissemination.