Exposure of endothelial cells to recombinant human erythropoietin induces nitric oxide synthase activity.
BACKGROUND
Anemic patients with chronic renal failure receiving recombinant human erythropoietin (rHuEPO) therapy frequently develop hypertension through an unknown mechanism. We hypothesize that EPO receptors (EPORs) on endothelial cells (ECs) in various sites of vasculature may mediate the activities of nitric oxide synthase (NOS) and/or the release of endothelin-1 (ET-1), contributing to blood pressure changes. We tested this hypothesis using primary cultures of ECs obtained from human coronary artery (HCAEC), pulmonary artery (HPAEC), dermis (HDEC), and umbilical vein (HUVEC).


METHODS
EPORs were measured by 125I-EPO binding. The effect of EPO on EPOR, ET-1, and NOS mRNA levels was assessed by quantitative reverse transcription-polymerase chain reaction. Cellular NOS activity and ET-1 release into the medium was measured by the NOSdetect assay and by radioimmunoassay kits.


RESULTS
Short-term (4 h) treatment with EPO (4 U/mL) did not change the number or affinity of EPOR per cell. Neither were there any changes in the amount of EPOR, ET-1, and NOS transcripts (cDNA/microg of mRNA) nor in ET-1 release and NOS activity. In HUVEC only, 24-hour exposure to EPO caused a threefold increase in NOS transcript. In other cells, EPO treatment for six days increased NOS activity by twofold to fourfold.


CONCLUSIONS
We show that upon extended exposure, EPO induces NOS activity but does not affect ET-1 release. These findings indicate that the hypertensive effect of EPO is not likely to be caused by a direct effect on ECs.
Banerjee
D
D
The Lindsley F. Kimball Research Institute of The New York Blood Center, New York, NY 10021, USA. dbanerje@nybc.org
Rodriguez
M
M
Nag
M
M
Adamson
J W
JW
D001794:Blood Pressure / Q000187:drug effects*; D002478:Cells, Cultured; D019332:Endothelin-1 / Q000235:genetics / Q000378:metabolism; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000201:enzymology; D004790:Enzyme Induction / Q000187:drug effects; D004921:Erythropoietin / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D019001:Nitric Oxide Synthase / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D017467:Receptors, Erythropoietin / Q000235:genetics; D011994:Recombinant Proteins
D019332:Endothelin-1; D012333:RNA, Messenger; D017467:Receptors, Erythropoietin; D011994:Recombinant Proteins; D004921:Erythropoietin; D019001:Nitric Oxide Synthase