VASCULAR ENDOTHELIAL-CELL PROLIFERATION - REGULATION OF CELLULAR POLYAMINES

Objective: The aims were to investigate the requirement for and regula tion of cellular polyamines during vascular endothelial cell prolifera tion. Methods: Proliferation of cultured bovine pulmonary artery endot helial cells was determined after cellular polyamine depletion. In add ition, polyamine synthesis and uptake mechanisms were examined in the presence and absence of trophic stimuli and density dependent inhibiti on of cell proliferation. Results: Serum stimulated, subconfluent cell s ceased cell division following the inhibition of ornithine decarboxy lase (ODC), a rate limiting enzyme for polyamine biosynthesis. The add ition of 10 mu M putrescine, the product of the enzyme reaction cataly sed by ODC, completely reversed the inhibition of cell growth. Serum d eprivation reduced cytosolic ODC activity to near non-detectable level s. Readdition of 10% fetal bovine serum (FBS) resulted in transient in creases in ODC activity which preceded DNA synthesis and mitosis. Basi c fibroblast growth factor also stimulated ODC activity in a dose depe ndent manner with levels approximating the maximum obtainable with FBS . In contrast, platelet derived growth factor and epidermal growth fac tor did not stimulate ODC activity. Finally, mitogenic stimuli did not induce ODC activity in density arrested cells. The uptake of radiolab elled putrescine from the cell medium was time dependent and saturable . Kinetic studies from dividing cells revealed a single transport comp onent for putrescine uptake [maximum rate of uptake (V-max) = 11.2(SEM 2.0) pmol . 10(5) cells(-1). h(-1); Michaelis constant (K-m)= 1.1(0.3 ) mu M]. Putrescine uptake by density arrested cells was characterised by a 57% decrease in V-max with no change in K-m. Readdition of FBS t o serum deprived subconfluent cells, in the presence of ODC inhibitors , resulted in a rapid increase in the rate of putrescine uptake with V -max increasing by 533% over FBS alone by 48 h. Discussion: These data suggest that polyamines are essential for endothelial cell proliferat ion and that synthesis and uptake mechanisms are regulated according t o cell growth.