The Role of C-X-C Chemokine Receptor Type 4 (CXCR4) in Cell Adherence and Spheroid Formation of Human Ewing’s Sarcoma Cells under Simulated Microgravity

Abstract

We studied the behavior of Ewing’s Sarcoma cells of the line A673 under simulated microgravity (s-µg). These cells express two prominent markers—the oncogene EWS/FLI1 and the chemokine receptor CXCR4, which is used as a target of treatment in several types of cancer. The cells were exposed to s-µg in a random-positioning machine (RPM) for 24 h in the absence and presence of the CXCR4 inhibitor AMD3100. Then, their morphology and cytoskeleton were examined. The expression of selected mutually interacting genes was measured by qRT-PCR and protein accumulation was determined by western blotting. After 24 h incubation on the RPM, a splitting of the A673 cell population in adherent and spheroid cells was observed. Compared to 1 g control cells, EWS/FLI1 was significantly upregulated in the adherent cells and in the spheroids, while CXCR4 and CD44 expression were significantly enhanced in spheroids only. Transcription of CAV-1 was upregulated and DKK2 and VEGF-A were down-regulated in both, adherent in spheroid cells, respectively. Regarding, protein accumulation EWS/FLI1 was enhanced in adherent cells only, but CD44 decreased in spheroids and adherent cells. Inhibition of CXCR4 did not change spheroid count, or structure. Under s-µg, the tumor marker EWS/FLI1 is intensified, while targeting CXCR4, which influences adhesion proteins, did not affect spheroid formation.