The effect of Ca2+ on the conversion of cortisol to its inert metabolite co
rtisone, the reaction catalyzed by the microsomal enzyme 11 beta -hydroxyst
eroid dehydrogenase type 2 (11 beta -HSD2), was investigated in human place
ntal microsomes. Placental microsomal 11 beta -HSD2 activity, as determined
by the rate of conversion of cortisol to cortisone, was inhibited up to 50
% by increasing free Ca2+ concentrations from 22 to 268 nM. The Ca2+-induce
d inhibition was reversible since chelation of endogenous Ca2+ with EGTA in
creased 11 beta -HSD2 activity up to 200%. Ca2+ decreased the maximal veloc
ity (V-max) of the 11 beta -HSD2 catalyzed conversion of cortisol to cortis
one without altering the K-m of 11 beta -HSD2 for cortisol, indicating that
Ca2+ modulates the catalytic efficiency rather than the substrate binding
of 11 beta -HSD2. Moreover, the Ca2+-induced inhibition does not appear to
involve altered cofactor (NAD(+)) binding since the inhibition of microsoma
l 11 beta -HSD2 activity by a sub-maximal concentration of free Ca2+ was no
t overcome by increasing the concentration of NAD(+). These findings in the
microsomes were then extended to an intact cell system, JEG-3 cells, an es
tablished model for human placental trophoblasts. In these cells, an increa
se in cytosolic free Ca2+ concentration ([Ca2+](i)) elicited by a known phy
siological stimulus, PGF(2 alpha), was accompanied by a 40% decrease in the
level of 11 beta -HSD2 activity. Furthermore, the PGF(2 alpha)-induced inh
ibition of 11 beta -HSD2 activity was abrogated when increases in [Ca2+](i)
were blocked with the intracellular Ca2+ chelator, BAPTA. Collectively, th
ese results demonstrate for the first time that Ca2+ inhibits human placent
al 11 beta -HSD2 activity by a post-translational mechanism not involving s
ubstrate or cofactor binding. (C) 2001 Academic Press.