Nad-dependent 11[beta]-hydroxysteroid dehydrogenase in cultured human
colonic epithelial cells.
Reeves, W. Brian, M. D.
Division of Nephrology, University of Arkansas College of Medicine
and John L. McClellan Veterans Affairs Hospital, Little Rock,
Arkansas
APStracts 2:0078C, 1995.
The inactivation of physiologic glucocorticoids by 11[beta]-hydroxysteroid
dehydrogenase (11[beta]-HSD) confers mineralocorticoid specificity to
certain aldosterone target tissues. However, 11[beta]-HSD activity in a
human mineralocorticoid responsive tissue has never been
characterized. The present studies describe the features of 11[beta]-HSD
in the cultured human colonic epithelial cell line, T84. The 11[beta]-HSD
activity of T84 cells resided in the microsomal fraction and showed a
marked preference for NAD rather than NADP as cofactor. 200 [mu]M NAD or
NADP increased the conversion of corticosterone to 11
-dehydrocorticosterone by 24.1+2.1 and 0.5+0.7 pmol/mg protein/20 min,
respectively, indicating a greater than 40-fold preference for NAD
versus NADP. The Km values for corticosterone and cortisol were
11.3+1.5 nM and 79.8+10 nM, respectively. The T84 11[beta]-HSD was
inhibited by 11-dehydrocorticosterone in a non-competitive fashion
(Ki=180+9.6 nM) and by carbenoxolone in a competetive fashion
(Ki=17.4+1.3 nM). The expression of mineralocorticoid receptors in
these cells was demonstrated by RT-PCR of mRNA isolated from T84
cells and by [3H]-aldosterone binding studies. The coexpression
of this NAD-dependent isoform of 11[beta]-HSD and mineralocorticoid
receptors is consistent with the view that the NAD-dependent isoform
is responsible for the specificity of mineralocorticoid responses.
Received 18 October 1994; accepted in final form 9 December 1994.
APS Manuscript Number C623-4.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 28 February 1995.