Thapsigargin inhibits angiogenesis in the rat isolated aorta: studies on the role of intracellular calcium pools.
OBJECTIVE
Since the role of Ca2+ in angiogenesis is not fully understood, we investigated the effect of thapsigargin (TG: depletes intracellular Ca2+ pools) and other Ca2+ modulators [ionomycin, calcium ionophore A23187 and dibutyrylhydroquinone (DBHQ)] on in vitro angiogenesis by rat aortic rings.


METHODS
Aortae from Sprague-Dawley rats were cut into 2-mm rings, embedded in a fibrin clot and cultured for 15 days in serum-free medium containing drugs and the microvessels counted. Rings were also pre-treated with TG and Ca2+ modulators for 1 h prior to embedding and culture. Viability was examined by the measurement of lactic acid dehydrogenase release. Rings were also treated with hydrocortisone and lavendustin A (a tyrosine kinase inhibitor), as positive controls. The effect of TG on the proliferation and migration of human umbilical artery endothelial cells (HUVECs) was studied in parallel.


RESULTS
TG significantly inhibited microvessel formation and HUVEC proliferation and migration in a dose-dependent manner, all at <10 nmol/l, without affecting viability. In contrast, ionomycin, A23187 and DBHQ were cytotoxic at inhibitory concentrations. Continual exposure to hydrocortisone and lavendusin A also inhibited angiogenesis without affecting viability.


CONCLUSIONS
Since low concentrations of TG deplete intracellular Ca2+ stores, it is concluded that these pools play a central role in mediating angiogenesis.
Shukla
N
N
Department of Anaesthesia and Cardiac Surgery, Bristol Royal Infirmary, University of Bristol, Bristol, UK.
Freeman
N
N
Gadsdon
P
P
Angelini
G D
GD
Jeremy
J Y
JY
D000704:Analysis of Variance; D000818:Animals; D001011:Aorta; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors*; D002455:Cell Division / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D046508:Culture Techniques; D003864:Depression, Chemical; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006854:Hydrocortisone / Q000494:pharmacology; D006873:Hydroquinones / Q000494:pharmacology; D007424:Intracellular Fluid / Q000378:metabolism*; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D008833:Microcirculation; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D010636:Phenols / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley; D019284:Thapsigargin / Q000494:pharmacology*; D014469:Umbilical Arteries
D004791:Enzyme Inhibitors; D006873:Hydroquinones; D007476:Ionophores; D010636:Phenols; D000001:Calcimycin; C062898:lavendustin A; D015759:Ionomycin; D019284:Thapsigargin; D011505:Protein-Tyrosine Kinases; D000252:Calcium-Transporting ATPases; D002118:Calcium; D006854:Hydrocortisone