| Cancer is one of the most serious diseases in the world. The number of newly diagnosed cancer patients keeps increasing and the mortality rate of cancer also remains higher in China. Radiotherapy is still the major method in the curative treatment of cancer, therefore, it is urgent to explore the underlying mechanism of DNA damage and repair in cancer cell and to develop the radiation sensitization drugs to enhance the index of the radiotherapy. In vitro three-dimensional(3D) growth of cell model is more closely mimics the features of the environment in vivo and is being used increasingly in the field of biological and medical research. It has been demonstrated that 3D cultured cancer cells are more radioresistant compared to the cancer cells cultured in monolayer(2D). However, the mechanisms causing this difference remain unclear. It is of great important to explore the reason why 3D cultured cancer cells are radioresistant because it is helpful for us to understand the radiation response occurred in human body and to develop the novel sensitivity drugs for radiotherapy.Epigenetics regulation refers to the heritable changes underlying gene expression by regulating the structure and function of the genome that are not caused by alterations in the DNA sequence itself, It includes DN A methylation regulation, histone modifications regulation,micro RNA(mi RNA) regulation and reprogramming regulation etc. An increasing number of studies demonstrate that epigenetics regulation plays important roles in many cellular processes from gene expression to cellular proliferation. It has been demonstrated that radiation induces the epigenetics regualtion alteration. Meanwhile, the changes of epigenetics regulation also affect the cellular response to ionizing radiation. An important correlationship exists between epigenetics regulation and cellular radiosensitivity.In current study, we attempt to investigate the reasons causing the difference in radiosensitivity between 2D and 3D cultured cancer cells from the perspective of epigenetics regulation, such as reprogramming regulation, DNA methylation regulation, histone acetylation regulation, mi RN A regulation.We found that cells cultured in 3D upregulated the expression of the stem cell reprogramming factors, and with a higher fraction of Rholow cells, compared to the 2D cultured cells. We also found that 3D cultured cells downregualted the expr ession of DNA methyltransferase. It is well know that the downregulation of DNA methyltransferse is associated with the cellular reprogramming progress and cancer stem- like cells are radioresistance, compared to the normal cancer cells.In addition, the expression of histone acetyltransferase Tip60 in the 3D cultured cancer cells increased after X-ray irradiation compared to the 2D cultured cancer cells. After pre-treatment with MG149, an inhibitor of Tip60, increased the lethallty rate of 3D cultured cancer cells after X-ray irradiation, suggesting that MG149 may apply as a sensitization drug during the radiotherapy.Further, the expression of mi RNAs in 2D and 3D cultured cancer cells were measured after x-ray irradiation. We found that the expression of mi R-1915 was decreased in 2D cultured cells while it was decreased more significantly in 3D cultured cells after irradiation. The overexpression of mi R-1915 increased the radiosensitivity of cancer cells to X-ray irradiation. Our subsequently studies indicated that the overexpression of mi R-1915 downregulated the m RNA level and protein expression of Rad9 a, one of the DN A damage repair factors, which may result in the radiosensitivity of 2D cells by reducing the DNA repair ability.In summary, our results suggest that the differences of culture enviroment result in the alteration of epigenetic phenotype between 2D and 3D cultured cells, which affect the DNA repair ability and ultimately induce the different radiosensitivity between the 2D and 3D cultured cells. Our findings reveal the rad iation induced DNA damage and repair mechanisms in the 3D cultured cancer cells model, a more mimics the features of the in vivo environment in human body, from the perspective of epigenetics modulation. It provides the guidance and experience for the development of radiotherapy sensitization drugs. |
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