Figure 1.

Cyclin G2 is a hypoxia-induced protein and abundant in pseudopalisade-forming glioma cells. (A and B) Hypoxic conditions enhance ccng2 mRNA transcription (A) and cyclin G2 protein expression (B). Cells were cultured in normoxic, deferoxamine mesylate (a reagent to mimic hypoxia)-added, or hypoxic conditions for 24 hours and subjected to reverse transcription-PCR or Western blot analysis. (C) Luciferase reporter assay with the human ccng2 promoter/pGL4.14 vector. U87MG cells were transfected with the reporter vector and cultured in normoxic or hypoxic conditions for 24 hours. Bars represent the SD. (D) ChIP assay showing that cyclin G2 expression is directly regulated by HIF-1α. (E) Cyclin G2 expression is dependent on HIF-1α accumulation. Cells treated with siRNA against HIF-1α show lower expression of cyclin G2 than scrambled siRNA-treated cells under hypoxic conditions. (F–M) Human glioblastoma specimens stained with hematoxylin and eosin (H&E) and immunostained with anti-cyclin G2 antibody. Other images are shown in Figure W4. (N) Cyclin G2 accumulates at the leading edge of glioma cells migrating from the necrotic foci to adjacent tissue. (O and P) Cyclin G2 is abundant in pseudopalisade-forming glioma cells (O) but not in inflammatory cells (CD68 positive) (P). (Q and R) Cyclin G2 expression is abundant in hypoxic or near necrotic regions. Dotted lines indicate the borders of pseudopalisades (F–I). The scale bars represent 200 µm (J–M), 50 µm (N), 100 µm (O and P), respectively. Numbers in B and E represent the ratio of densitometry analysis.