Inhibition of intracellular Ca2+ mobilisation by low antiproliferative concentrations of thapsigargin in human vascular smooth-muscle cells.
Low nanomolar concentrations of thapsigargin, a modulator of intracellular Ca2+ ([Ca2+]i) pools, inhibit vascular smooth-muscle cell (VSMC) proliferation. Because the mechanisms underlying this effect have not been defined, the effect of antiproliferative concentrations of thapsigargin on [Ca2+]i in fura-2-loaded VSMCs was studied by using dynamic video imaging of [Ca2+]i. After seeding on coverslips, human VSMCs were incubated for 1-48 h with thapsigargin before loading with fura-2 or during imaging. Mobilisation of [Ca2+]i was stimulated with 1 microM ionomycin in Ca2+-free medium and the increase in [Ca2+]i detected by using Ca2+ imaging techniques. Continuous exposure of cells to low concentrations of thapsigargin (which failed measurably to increase in [Ca2+]i) reduced the ionomycin response in a time- and dose-dependent manner (100% inhibition at 10 nM thapsigargin after 1 h exposure). After exposure of cells to 10 nM thapsigargin for 1 h followed by washing and further incubation for < or = 72 h, there was a time-dependent recovery of the ionomycin response. Because the concentrations of thapsigargin and exposure times are identical to those that inhibit replication in VSMCs, it is proposed that depletion of [Ca2+]i pools mediates the inhibitory effect of thapsigargin on VSMC proliferation.
Bristol Heart Institute, Bristol Royal Infirmary, University of Bristol, England, UK.
S D
SD
Birkett
J Y
JY
Jeremy
S M
SM
Watts
N
N
Shukla
G D
GD
Angelini
C A
CA
McArdle
D002118:Calcium / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects*; D003952:Diagnostic Imaging; D004305:Dose-Response Relationship, Drug; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects*; D012501:Saphenous Vein / Q000187:drug effects; D019284:Thapsigargin / Q000494:pharmacology*; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors
D015759:Ionomycin; D019284:Thapsigargin; D007770:L-Lactate Dehydrogenase; D002118:Calcium; D013936:Thymidine