The expression of renin and the formation of angiotensin II in bovine aortic endothelial cells

One controversy in the field of vascular angiotensin generation has surroun ded the nature and particularly the sourer of vascular renin. This study in vestigated the expression of renin protein and its mRNA in aortic endotheli al cells using immunocytochemistry, Western blotting, in situ hybridization and reverse transcription PCR (RT-PCR). Using a monoclonal antibody agains t human renin, immunocytochemical analysis revealed positive immunoreactivi ty in the cytoplasm of cultured bovine aortic endothelial cells. Immunoblot ting of solubilized proteins separated by SDS-PAGE from cultured aortic end othelial cells identified two immunoreactive species with molecular masses of approximately 37-40 kDa. In situ hybridization showed that renin mRNA wa s localized in the cytoplasm of these cells. Using RT-PCR of RNA extracted from bovine aortic endothelial cells with primers specific for human renin, a clear single band was detected, which had the predicted size of 142 bp f or (pro)renin. Angiotensin II (Ang II) was assayed in conditioned medium (CM) from culture d bovine aortic endothelial cells, and in addition, the effects of Ang II a nd CM on the proliferation of aorta smooth muscle cells (ASMC) were also st udied. The results showed that CM contained Ang II equivalent to 15.05 +/- 4.67 pg/10(6) cells. Assay of smooth muscle cell proliferation by cell numb er, and by tritiated thymidine uptake, showed that proliferative responses in the presence of Ang II at a concentration of 10(-6) M were evident withi n 1 day of subculture, and cell numbers were nearly twice those of controls after 2 days. Thymidine incorporation into ASMC was also increased by Ang II in a dose-dependent manner and by endothelial cell CM. In both cases, st imulated proliferation was inhibited by the Ang II type 1 (AT1) receptor se lective antagonist, losartan. These findings suggest that these vascular en dothelial cells are a source of locally synthesized renin that may thus be involved in vascular Ang II generation. They also suggest that Ang II produ ced by the endothelial cells may be secreted and stimulate ASMC proliferati on via the AT1 receptor.