Mechanical strain increases endothelin-1 gene expression via protein kinase C pathway in human endothelial cells.
Vascular endothelial cells (ECs) are constantly subjected to mechanical strain due to relaxation and contraction of vessel walls. The effects of cyclical strain on endothelin-1 (Et-1) secretion and Et-1 mRNA levels in human umbilical vein ECs were examined. Cultured ECs grown on a flexible membrane base were deformed by negative pressure (16 kPa at 60 cycles/min). Cells subjected to strain showed increased Et-1 secretion (0.54 ng/hr/10(6) cells) compared with unstrained control cells (0.22 ng/hr/10(6) cells). Northern blot analysis of cells strained for 2 hours or longer demonstrated a sustained elevated Et-1 mRNA level at more than double the level in unstrained controls. This strain-induced ET-1 mRNA level returned to its basal level 2 hours after the release of strain. Cells treated with actinomycin D before or during strain treatment showed no strain-induced gene expression. Pretreatment of ECs with a protein kinase C (PKC) inhibitor, Calphostin C, strongly inhibited the strain-induced Et-1 gene expression. Pretreatment of ECs with cAMP- or cGMP-dependent protein kinase inhibitors (KT5720 or KT5823) only partially inhibited the increased Et-1 mRNA levels in strain-treated cells. EGTA strongly inhibited the Et-1 gene expression. The intracellular calcium chelator BAPTA/AM also showed an inhibitory effect on Et-1 mRNA levels. We conclude that mechanical strain can stimulate the secretion of Et-1 from ECs by increasing Et-1 mRNA levels via transcription, and that this gene induction is mediated predominantly via the PKC pathway and requires extracellular Ca2+. This strain-induced Et-1 gene expression in ECs may contribute to the regulation of vascular tone and structure in normal and pathological states of the cardiovascular system.
Wang
D L
DL
Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan, Republic of China.
Wung
B S
BS
Peng
Y C
YC
Wang
J J
JJ
D002478:Cells, Cultured; D003609:Dactinomycin / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D016232:Endothelins / Q000235:genetics* / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology*; D015870:Gene Expression; D006801:Humans; D009281:Naphthalenes; D011083:Polycyclic Compounds / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger / Q000378:metabolism; D013314:Stress, Mechanical
D016232:Endothelins; D009281:Naphthalenes; D011083:Polycyclic Compounds; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D003609:Dactinomycin; D004533:Egtazic Acid; D011493:Protein Kinase C; C058819:calphostin C; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid