Abstract 3846: The effect of x-ray radiation on spheroids cultured in a 3D matrix under different oxygen conditions .
Authors: Dornish John Michael , Markussen Christine , Andersen Therese 
Published: August 2014

Abstract
The use of 3-dimensional (3D) matrices for cell culture has become popular as it provides more representative models of the biological situation in tissues, organs and tumors than the traditional 2D cell culture. NovaMatrix®-3D is an alginate-based cell culture system comprising an alginate foam matrix and an alginate immobilization solution. The cells are first suspended in the alginate solution and applied to the foam. This induces in situ gelation by diffusion of calcium ions to the added alginate and the cells are immobilized inside the pores of the foam. Alginate is not recognized by cell receptors and is considered inert towards cells. Cells form 3-dimensional structures such as spheroids, and the system may be utilized as an in vitro tumor model. The growth of spheroids inside the 3D matrix can be determined by evaluating changes in spheroid volume. Hypoxic cells are more resistant towards traditional cancer treatments such as chemotherapy and radiation. This study used cells cultured in a 3D matrix to evaluate the effect of different doses of x-ray radiation when cells were cultured under different oxygen concentrations. NHIK 3025 cells (human cervical carcinoma) were cultured in 2D and then established as spheroids in NovaMatrix®-3D under 20% O2 and 5% O2 continuous culture conditions. The growth rate of spheroids was first determined without x-ray radiation of the cells. In subsequent experiments the cells were irradiated after 8 days of culture at a dose of 5 Gy or 15 Gy. In all experiments the spheroid growth rate was monitored by determining the volume of the spheroids by vital staining and confocal imaging at regular intervals over 30 days. The volumes were obtained by measuring the diameter of the spheroids in both x- and y-directions. The radii were calculated from the average of these diameters and were then used to calculate the volumes. Spheroids were formed under both normoxic and hypoxic conditions and it was possible to determine growth curves from the volume measurements. The sigmoidal growth curves were similar for both normoxic and hypoxic conditions without radiation treatment. An x-ray radiation dose of 5 Gy was not sufficient to affect the growth rate of the spheroids showing that NHIK 3025 cells are more radiation resistant when grown as 3D spheroids than in traditional 2D culture. When the spheroids were irradiated with 15 Gy, a decrease in growth rate was observed. The spheroids cultured under 20% oxygen were affected more, i.e. the growth curves showed a significant reduction in volume increase, compared to the spheroids cultured under 5% oxygen. This study indicates that hypoxia increases the resistance of NHIK 3025 spheroids against x-ray radiation, and demonstrates the utility of a 3D cell culture matrix (NovaMatrix®-3D) in dose response experiments.
This work was funded by the METOXIA project no.222741 under the 7th Research Framework Program of the European Union.
Citation Format: John Michael Dornish, Christine Markussen, Therese Andersen. The effect of x-ray radiation on spheroids cultured in a 3D matrix under different oxygen conditions . [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3846. doi:10.1158/1538-7445.AM2013-3846