Conditioned media from hypoxic endothelial cells inhibit proliferation. Human microvascular endothelial cell 1 (HMEC-1) cells and C2C12 muscle cells were grown to confluence and then cultured further under normoxic or hypoxic conditions. Media were transferred undiluted to rapidly proliferating HMEC-1 cells in the presence of 10% FCS. Culture in DMEM with 10% FCS was used as a positive control, and culture with 10 μM mitomycin C added at baseline to block proliferation was used as a negative control. After 24 hours, further culture proliferation of HMEC-1 cells was measured using a haemocytometer, and the cells were imaged. (a) Exposure to conditioned media from HMEC-1 cells, but not from C2C12 cells, inhibited proliferation (results are expressed as total number of cells per well of a six-well plate/10-4). (b) Effects were more marked with media from hypoxic HMEC-1 cells which abolished proliferation and caused rounding up of the cells.