This is supported by analyses performed on day 3, which demonstrated the presence of a few pimonidazole-stained cancer cells in L3 and a large number of such cells in L4 (Figure 3A,B and Supplementary Figure S5A,B), as well as an increase in the expression of HIF-1α and its target genes, Glut1, CA9, VEGF, and LDHA, as the distance from the media increased (i.e., from L1 to L4). Here, HIF1A is linked to cancer.