IntroductionCancer patients often suffer from local tumor recurrence and distant tumor metastases after radiation therapy. It has been widely accepted that these problems are caused by the existence of radioresistant cancer cells in malignant solid tumors. In vitro experiments indicated that oxygen-availability and cell cycle status each influences radiosensitivity of cancer cells; cancer cells obtain radioresistance when they are in S phase or when surrounding environment is hypoxic. However, how and where these two factors influence each other in highly heterogeneous malignant tumors and produce radioresistant cancer cells remain largely unknown.
MethodsHeLa cells and male Balb/c nude mice were used in our research. Immunohistochemistry was also performed with HeLa tumor xenografts to analyze the spatial relationship between the extrinsic and intrinsic factors and its impact on radioresistant characteristics of cancer cells in diverse tumor microenvironments.
ResultsWe first perform IHC staining analyses with intrinsic and extrinsic hypoxia markers, HIF-1a and pimonidazole, respectively, and with an S phase marker, Cyclin A. Hypoxic regions were detected with both hypoxia markers in distal regions from perfusion-positive tumor blood vessels. HIF-1a-positive area was slightly but definitelycloser to blood vessels than the pimonidazole-positive area, consistent with previous reports. Cancer cells with high level of Cyclin A expression were detected predominantly in pimonidazole-positive regions rather than in HIF-1a-positive regions and normoxic regions.
We examined whether cells in pimonidazole-positive hypoxic regions are actually resistant to radiation in vivo. HeLa tumor xenografts were treated with 4 Gy of X-radiation, and 30 min later surgically excised for immunohistochemical analysis with the combination of antibodies against pimonidazole and gH2AX. The number of gH2AX foci was obviously decreased in pimonidazole-positive regions than proximal regions to tumor blood vessels.
Conclusions and DiscussionsThere is tight relationship between cell cycle status and tumor microenvironments, and decreasing in radiation-induced DNA damage in Pimonidazole-positive Regions of solid Tumors. Our results indicate that hypoxic but HIF-1a-negative conditions increase S phase cancer cells in pimonidazole-positive perinecrotic regions. Cancer cells in pimonidazole-positive regions are more radioresistant than the other populations, and therefore suffered from less DNA damage after radiation treatment.
This project is supported by 973 key project (No:2011CB503704) and National Natural Science Foundation of China (No: 60871068;No: 60971055) and International Science & Technology CooperationProject (No: 2010DFA31900).