Figure 1

Apoptosis and hypoxia markers in normoxic and hypoxic ex vivo cultured lung cancer fragments. (A) Apoptosis is not increased in hypoxic ex vivo cultured NSCLC fragments. Representative images of cleaved caspase 3 staining for the assessment of apoptosis are shown: a hypoxic adenocarcinoma fragment cultured for three days in hypoxia without visible apoptosis and a fragment treated with 32 μM cisplatin as positive control. Right, apoptosis rates were determined by counting cleaved caspase 3 positive tumor cells in a blinded manner. Lines indicate mean +/- SEM. Groups were compared with the Mann–Whitney-U test. (B) HIF-1α and HIF-2α immunoreactivity in NSCLC fragments after three days of incubation in hypoxia or normoxia. Arrowhead: stroma cell, arrow: tumor cell. HIFs were evaluated in tumor cells and stroma cells semi-quantitatively in a blinded manner. Groups were compared with the paired Student’s t-test. (C) Pimonidazole staining indicating hypoxia in fragments cultured for one or three days in hypoxia or normoxia. Representative images are shown. All sections were counterstained with hematoxylin. (D) Carbonic anhydrase IX (CA IX) mRNA in hypoxic vs. normoxic NSCLC fragments (n = 14 patients). No difference in expression gives a value of zero. Significance was calculated with the single group Student’s t-test. Results are mean +/- SEM. NOX, normoxia; HOX, hypoxia.