Morphology and cellular integrity of HUVEC treated with different cAMP derivates. (A) Stimulation of HUVEC for 72 h with cAMP or 8-Br-cAMP induced a regular monolayer but led to extensive vacuolization (white arrows) that may represent signs of cytotoxicity (a,b). Cells stimulated with 8-Br-cAMP/Na or pCPT-cAMP grew in a dense monolayer with regularly arranged cells and visible cell borders (c,d) without any signs of cytotoxicity. Shown are representative micrographs of at least three experiments with cells derived from different donors. (B) All four cAMP derivates induced enhanced cellular cytotoxicity in the first 24 h whereas after 72 h the ratio of dead and viable cells is comparable or, for pCPT-cAMP, even below that of untreated cells. TNF-α/CPT treated HUVEC served as positive control. (n = 3) (C) Exposure to cAMP, 8-Br-cAMP or 8-Br-cAMP/Na induced a slight reduction in intracellular ATP level over the whole time period. Treatment with pCPT-cAMP resulted in a partial decrease of cellular ATP concentrations after 24 h but showed no differences to untreated cells after 72 h. (n = 3) (D) All four cAMP derivates induced a slight decrease in caspase3/7 activity after 24 h. After 48 h and 72 h level of activated caspases are comparable to those of untreated cells (n = 4). Scale bar represent 25 μm.