Structure-dependent inhibition of human and rat 11β-hydroxysteroid dehydrogenase 2 activities by phthalates

Article abstract of DOI:10.1016/j.cbi.2009.09.014

Phthalates are diesters of phthalic acid and an alcohol moiety. Phthalates have been classified as endocrine disruptors and have a broad range of effects with unknown mechanisms. Some of the effects of phthalate are consistent with disruptions of normal glucocorticoid homeostasis, and in particular, with defective function of 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2). In the present study, we tested 12 phthalate diesters and four monoesters for the inhibition of human and rat kidney 11β-HSD2. We examined the modes of inhibition and looked for a relationship between the potency for inhibition and the chemical structures. Of the phthalate diesters we tested, dipropyl phthalate (DPrP) and di-n-butyl phthalate (DBP) significantly inhibited both human and rat 11β-HSD2 activities. The IC₅₀s were 85.59 μM for DPrP and 13.69 μM for DBP when calculated for rat 11β-HSD2. As diesters, 8 of the phthalates did not affect 11β-HSD2 enzyme activity. Compared to the diesters that were inhibitory, the 8 non-inhibitory phthalates, had either fewer carbons, that is 1 or 2 carbons in the alcohol moiety, or more carbons, 5-10, as a branched or unbranched chain in the alcohol moeity. However, phthalates could be inhibitors with six carbons in the alcohol moiety if the carbons were cyclized, as in dicyclohexyl phthalate (DCHP), which inhibited rat 11β-HSD2 with an IC₅₀ of 32.64 μM. Thus, whether a phthalate is an inhibitor may reflect the size and shape of the compound. Although the diesters are the compounds used in manufacturing and present as environmental contaminants, it is the monoester metabolites that are detected in human serum and urine. We showed that mono (2-ethylhexyl) phthalate (MEHP) significantly inhibited human (IC₅₀ = 110.8 ± 10.9) and rat (121.8 ± 8.5 μM) 11β-HSD2 activity even though its parent compound, di(2-ethylhexyl) phthalate (DEHP) did not. MEHP was a competitive inhibitor of 11β-HSD2 enzymatic activity. We conclude that phthalates of a certain size act as competitive inhibitors.

Full text of DOI:10.1016/j.cbi.2009.09.014

Chemico-Biological Interactions 183 (2010) 79–84
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Chemico-Biological Interactions
journal homepage: www.elsevier.com/locate/chembioint
Structure-dependent inhibition of human and rat 11␤-hydroxysteroid
dehydrogenase 2 activities by phthalatesଝ
Binghai Zhaoa,b,c, Yanhui Chuc, Yadong Huangd, Dianne O. Hardyb,
Shaoqiang Line,∗∗, Ren-Shan Geb,e,∗
a School of Public Health, Jilin University, Changchun, Jilin 130118, PR China
b Population Council, 1230 York Avenue, New York, NY 10065, USA
c Heilongjiang Key Laboratory of Anti-fibrosis Biotherapy, Mu Dan Jiang Medical College, Heilongjiang, PR China
d Institutes of Life and Health Engineering, Jinan University, Guangzhou, Guangdong 510632, PR China
e School of Pharmacy, Wenzhou Medical College, Wenzhou, Zhejiang 325000, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 28 July 2009
Received in revised form
15 September 2009
Accepted 18 September 2009
Available online 26 September 2009
Phthalates are diesters of phthalic acid and an alcohol moiety. Phthalates have been classified as endocrine
disruptors and have a broad range of effects with unknown mechanisms. Some of the effects of phthalate
are consistent with disruptions of normal glucocorticoid homeostasis, and in particular, with defec-
tive function of 11␤-hydroxysteroid dehydrogenase 2 (11␤-HSD2). In the present study, we tested
12 phthalate diesters and four monoesters for the inhibition of human and rat kidney 11␤-HSD2. We
examined the modes of inhibition and looked for a relationship between the potency for inhibition
and the chemical structures. Of the phthalate diesters we tested, dipropyl phthalate (DPrP) and di-n-
butyl phthalate (DBP) significantly inhibited both human and rat 11␤-HSD2 activities. The IC50s were
85.59 ␮M for DPrP and 13.69 ␮M for DBP when calculated for rat 11␤-HSD2. As diesters, 8 of the phtha-
lates did not affect 11␤-HSD2 enzyme activity. Compared to the diesters that were inhibitory, the 8
non-inhibitory phthalates, had either fewer carbons, that is 1 or 2 carbons in the alcohol moiety, or
more carbons, 5–10, as a branched or unbranched chain in the alcohol moeity. However, phthalates
could be inhibitors with six carbons in the alcohol moiety if the carbons were cyclized, as in dicy-
clohexyl phthalate (DCHP), which inhibited rat 11␤-HSD2 with an IC50 of 32.64 ␮M. Thus, whether a
phthalate is an inhibitor may reflect the size and shape of the compound. Although the diesters are
the compounds used in manufacturing and present as environmental contaminants, it is the monoester
metabolites that are detected in human serum and urine. We showed that mono (2-ethylhexyl) phthalate
(MEHP) significantly inhibited human (IC50 = 110.8 10.9) and rat (121.8 8.5 ␮M) 11␤-HSD2 activity
even though its parent compound, di(2-ethylhexyl) phthalate (DEHP) did not. MEHP was a competitive
inhibitor of 11␤-HSD2 enzymatic activity. We conclude that phthalates of a certain size act as competitive
inhibitors.
Keywords:
Phthalates
11␤-HSD2
Enzyme inhibition
Kidney microsome
© 2009 Published by Elsevier Ireland Ltd.
1. Introduction
reports that exposures to phthalates may be linked to abnormal
reproductive development and the metabolic syndrome includ-
Phthalate diesters of phthalic acid are used as plasticizers
and solvents in polyvinyl chloride (PVC) products, children’s toys,
cosmetics, soaps, shampoos, medical tubing, catheters and intra-
venous delivery sets. Public concern has arisen in response to
ing obesity and insulin resistance in humans [1–3]. In most
species, phthalate diesters are rapidly hydrolyzed within the gut
by hydrolases into phthalate monoesters. Metabolites such as
monomethyl phthalate (MMP), and mono(2-ethylhexyl) phthalate
(MEHP), (derived from Fig. 1)[4] are thought to be more toxic than
the parent diesters, dimethyl phthalate (DMP) and di(2-ethylhexyl)
phthalate (DEHP) [4]. Among phthalates, DEHP is the most abun-
dantly used. Although daily exposure to humans in the general
population is only in the range from 5.8 to 19 ␮g/kg/day [5], the
exposure during medical infusion may be up to 167.9 mg/day [5].
Neonates may be exposed to high levels of phthalate through con-
tact with medical devices. Epidemiological studies have shown
statistical correlations between urinary concentrations of phtha-
ଝ
Matthew P. Hardy and R.-S. Ge were supported in part by NIH R01 ES10233 and
Y. Chu by Heilongjiang Natural Science Foundation D2007-99.
∗
Corresponding author at: Population Council, 1230 York Avenue, New York, NY
10065, USA. Tel.: +1 212 327 8754; fax: +1 212 327 7678.
∗∗
Corresponding author.
E-mail address: shaotsiang@163.com (S. Lin),
rge@popcbr.rockefeller.edu (R.-S. Ge).
0009-2797/$ – see front matter © 2009 Published by Elsevier Ireland Ltd.
doi:10.1016/j.cbi.2009.09.014

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B. Zhao et al. / Chemico-Biological Interactions 183 (2010) 79–84
Fig. 1. Structures of DEHP and DMP and their respective phthalate monesters. Di(2-ethylhexyl) phthalate (DEHP) and dimethyl phthalate (DMP) are converted into their
respective metabolites, mono (2-ethylhexyl) phthalate (MEHP) and monomethyl phthalate (MMP).
Fig. 2. Structures of phthalate diesters: Twelve phthalate diesters with length of the side-chains from 1 to 10 are listed. They are dimethyl (DMP, C1), diethyl (DEP, C2),
dipropyl (DPrP, C3), di-n-butyl (DBP, C4), butylbenzyl (BBzP, C4, 6), di-n-pentyl (DNPP, C5), di-cyclohexyl (DCHP, C6), di-n-heptyl (DNHP, C7), di-ethylhexyl (DEHP, C8),
di-n-ocytl (DNOP, C8), diisononyl (DINP, C9), and diisodecyl (DIDP, C10) phthalates.

B. Zhao et al. / Chemico-Biological Interactions 183 (2010) 79–84
81
late monoesters and incidence of anomalies such as cryptorchidism
2.2. Preparation of microsomal protein
and shortened anogenital distance of newborn males and increased
insulin resistance [1,3], and thus provide evidence of endocrine
system disruption.
Rat kidney microsomes were prepared as described previously
[17]. In brief, rat kidney cortex was homogenized in 0.01 mM PBS
buffer containing 0.25 M sucrose (pH 7.4), and cleared of large cell
debris by centrifugation at 750 × g for 30 min. The homogenates
were centrifuged at 12,000 × g for 30 min, and the postmito-
chondrial supernatants were obtained by ultracentrifugation at
105,000 × g for 60 min, twice, to collect the microsomal pellets.
The postnuclear supernatants were centrifuged at 105,000 × g, the
resultant microsomal pellets were resuspended. Protein contents
were measured using Bio-Rad Protein Assay Dye Reagent Concen-
trate. Microsomes were used for the measurement of 11␤-HSD2
activities.
Elevation of serum glucocorticoid levels has been linked to
suppression of the reproductive system and to the metabolic syn-
drome [6]. Access of glucocorticoids to the classic glucocorticoid
receptor (GR) is controlled by 11␤-hydroxysteroid dehydrogenase
(11␤-HSD) that catalyzes interconversion of active glucocorti-
coid (cortisol in human or corticosterone, CORT, in rodents) to
inert steroids (cortisone in human or 11-dehydroxcorticosterone,
11DHC, in rodents). It is now known that there are two isoforms.
The type 1 isoform (11␤-HSD1), an oxidoreductase, first puri-
fied from rat liver [7] and then cloned [8], requires the pyridine
nucleotide cofactor NADP(H): NADP+ for dehydrogenase activity
to catalyze the conversion of CORT into 11DHC, and NADPH for
reductase activity to catalyze 11DHC to CORT [9]. The type 2 iso-
form (11␤-HSD2) was identified later, and cloned after purification
from the kidney [10,11]. 11␤-HSD2 has no detectable reductase
activity and is known to be an exclusively oxidative enzyme acti-
vated by a different cofactor, NAD+ [10]. 11␤-HSD2 is primarily
localized in classic mineralocorticoid receptor targeted tissues such
as kidney and colon but is also found in non-mineralocorticoid
receptor targeted tissues such as placentum and testis [12]. In
kidney, 11␤-HSD2 eliminates the natural glucocorticoid cortisol
or CORT and thereby protects mineralocorticoid receptors from
occupation by active glucocorticoids. In human, mutation of the
11␤-HSD2 gene causes a syndrome named apparent mineralocorti-
coid excess (AME) characterized by hypertension and hypokalemia
in which circulatory aldosterone levels are subnormal [12]. The
AME can also be caused by the inhibition of the enzyme using chem-
icals such as glycyrrhentic acid, a component of licorice [12]. The
inhibition or mutation of placental 11␤-HSD2 may be related to
the restriction of fetal growth and prematurity in rat models and
humans [13,14].
2.3. 11ˇ-HSD2 assay
11␤-HSD2 activity assay tubes contained 10 nM (within the
range of physiological levels) of CORT (containing 30,000 cpm [3H]-
CORT). CORT was used as substrate to measure 11␤-HSD2 oxidase
activity. The kidney microsomes were incubated with substrates,
0.2 mM NAD+, 5 mM DTT and various concentrations of phthalates.
The reactions were stopped by adding 2 ml ice-cold ether. The
steroids were extracted, and the organic layer was dried under
nitrogen. The steroids were separated chromatographically on
thin layer plates in chloroform and methanol (90:10, v/v), and the
radioactivity was measured using a scanning radiometer (System
AR2000, Bioscan Inc., Washington, DC). The percentage conversion
of CORT to 11DHC was calculated by dividing the radioactive
counts identified as 11DHC by the total counts associated with
CORT plus 11DHC.
2.4. Determination of half maximum inhibitory concentrations
(IC50) and inhibitory mode
The IC50 was determined by adding 10 nM substrate and 0.2 mM
NAD+ and various concentrations of phthalates to 250 reaction
buffer (0.1 mM PBS) containing 40 ␮g microsomal protein. The
mode of inhibition was assayed by adding various concentrations
of either CORT or NAD+.
It has been observed that dicyclohexyl phthalate (DCHP) can
inhibit human 11␤-HSD2 activities [15]. However, there are over
20 phthalates commonly present in the environment for which
the relationship between structure and inhibitory potency is not
known. In the present study, we compared 12 phthalate diesters
and 4 monoesters for the ability to inhibit human or rat 11␤-HSD2
and also examined the mode of inhibition.
2.5. Statistics
Each experiment was repeated three to four times. Data were
subjected to nonlinear analysis by GraphPad (Version 3, GraphPad
Software Inc., San Diego, CA) for IC50. Data were subjected to analy-
sis by one-way ANOVA followed by DUNCAN multiple comparison
testing to identify significant differences between groups when
three and more groups were calculated. All data are expressed as
means SEM. Differences were regarded as significant at P < 0.05.
2. Materials and methods
2.1. Chemical and animals
[1,2-3H] Corticosterone (3H-CORT), specific activity, 40 Ci/
mmol) was purchased from Dupont-New England Nuclear (Boston,
MA). [1,2-3H]11dehydrocorticosterone (3H-11DHC) was prepared
from labeled 3H-CORT as described earlier [16]. Cold CORT and
11DHC were purchased from Steraloids (Newport, RI). The fol-
lowing phthalates were purchased from Sigma–Aldrich Company
(St. Louis, MO, USA): dimethyl phthalate (DMP), diethyl phtha-
late (DEP), dipropyl phthalate (DPrP), di-n-butyl phthalate (DBP),
butyl benzyl phthalate (BBzP), di-n-pentyl phthalate (DNPP), DCHP,
DEHP, di-n-heptyl phthalate (DNHP), di(n-octyl) phthalate (DNOP),
diisononyl phthalate (DINP) and diisodecyl phthalate (DIDP) as
well as mono-methyl phthalate (MMP) and mono-benzyl phtha-
late (MBzP). MEHP and mono-n-butyl phthalate (MBP) were from
TCI America (Portland, OR, USA). Long-Evans rats (250–300 g)
were purchased from Charles River Laboratories (Wilmington,
MA). Human kidney microsomes were purchased from BD Gentest
(Woburn, MA).
3. Results
3.1. Effects of phthalates on 11ˇ-HSD2 activities in human and
rat kidney microsomes
The structures of the 12 phthalate diesters are shown in Fig. 2. Of
the 12 phthalate diesters tested at 1 mM (a concentration that may
be achieved during extracorporeal membrane oxygenation expo-
sure to DEHP in infants [18]), DPrP and DBP significantly inhibited
rat 11␤-HSD2, the IC50s were 85.59 ␮M (DPrP) and 13.69 ␮M (DBP).
DPrP and DBP were less potent for the inhibition of human 11␤-
HSD2, the IC50s were >1 mM. The other 9 phthalate diesters had
alcohol moieties consisting of either 1–2 or 5–10 carbons in non-
cyclic chains and had no effects on 11␤-HSD2 activity at the 1 mM
concentration (Fig. 3). Dicyclohexyl phthalate (DCHP) had signif-

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B. Zhao et al. / Chemico-Biological Interactions 183 (2010) 79–84
Fig. 3. Effects of phthalate diesters on kidney 11␤-HSD2 activities. (A) Human kid-
ney microsomes. (B) Rat kidney microsomes. The measurement of 11␤-HSD2 was
performed in the presence of 1 mM phthalates according to Section 2. Data are pre-
sented as mean SEM (n = 4). A significant difference compared to control (DMSO
solvent) was shown at *P < 0.05, ***P < 0.001.
icant inhibitory potency for 11␤-HSD2 (IC50 of 32.64 ␮M for rat
enzyme). BBzP is different from dibutyl phthalate only by the sub-
stitution of one butyl group for the benzyl group, and did not inhibit
the enzyme.
Fig. 4. Effects of phthalate diesters and monoesters on 11␤-HSD2 activities in
human and rat kidney microsome. The measurement of 11␤-HSD2 was performed
in the presence of 1 mM phthalates according to Section 2. Mean SEM (n = 4), *,
*** indicates significant differences compared to their phthalate diesters at P < 0.05,
P < 0.001.
Because phthalate diesters are metabolically converted into
monoesters, we examined the potencies of monoester metabolites
of three diesters, DMP, BBzP and DEHP, that did not inhibit 11␤-
HSD2 activity. At equimolar concentrations relative to their diesters
DMP and DEHP, MMP and MEHP were more potent inhibitors of
both human and rat 11␤-HSD2. MBzP was an inhibitor of rat 11␤-
HSD2 (Fig. 4). We measured human and rat 11␤-HSD2 activities
at various concentrations of MEHP. The IC50s were 110.8 10.9
(human) and 121.8 8.5 ␮M (rat) (Fig. 5). MBP, the metabolite of
DBP, did not inhibit 11␤-HSD2 (Fig. 4). Given that BBzP, which also
has only one butyl group, was not inhibitory, we infer that two butyl
groups in the alcohol moiety confer more potency than a single
butyl group.
either 3 or 4 carbons in unbranched chains, or a cyclohexyl group.
The 9 phthalate diesters that did not inhibit 11␤-HSD2 at 1 mM
concentrations had non-cyclic alcohol moieties consisting of 1–2
or 5–10 carbons in branched or unbranched chains. We infer that
the size of the alcohol moiety in the diester is an important crite-
rion for potency. The two cyclohexyl groups in DCHP might confer
a more compact shape to the compound than the two 6-carbon
unbranched chains in DNHP because the three dimensional struc-
ture of cyclohexane is the chair conformation.
Although DEHP, DMP and BBzP did not inhibit 11␤-HSD2, the
monoester metabolites of these compounds, MEHP, MMP and MBzP
did (Fig. 4). This result is consistent with the notion that the metabo-
lites of the commonly used phthalate DEHP cause greater harm than
the original compounds [4]. BBzP, a non-inhibitor, differs from DBP,
a potent inhibitor in that one of the alcohol moieties is a benzyl
group instead of a butyl group. The metabolite MBzP lacks the butyl
group, and has increased the potency for the inhibition of rat 11␤-
HSD2 relative to the parent compound. Similarly, the removal of
one ethylhexyl group in DEHP as by metabolic converstion to MEHP,
significantly increased the potency for the inhibition of 11␤-HSD2
(Fig. 4).
Fetal testosterone production of rats is inhibited by exposures
of rats to phthalates in vivo. BBzP, DBP, DEHP, DCHP and DPrP
were among the most potent inhibitors while DMP, DEP and DIOP
had no effects [19–21]. Thus the structural characterization that
distinguishes the inhibitors and non-inhibitors of testosterone
production may overlap with the pattern we observed for the inhi-
bition of rat 11␤-HSD2 as DPrP, DBP and DCHP were most potent
(Fig. 3). However, whether the same relationship between structure
and activity is relevant to the inhibition of testosterone is worthy
of further investigation. Although DEHP and BBzP did not directly
inhibit 11␤-HSD2, each of these diesters would be metabolically
activated after conversion to MEHP and MBzP, respectively. This
would be consistent with ability of these two diesters to inhibit
fetal testis function in vivo and absence of inhibition in vitro.
3.2. The mode of inhibition of 11ˇ-HSD2 by MEHP
We examined the mode of inhibition of MEHP for rat and human
kidney 11␤-HSD2. Dixon plots showed that MEHP was competi-
tive with CORT for the inhibition of 11␤-HSD2 activities in both
human (Fig. 6) and rat (data not shown) microsomes. When various
NAD+ concentrations were used to measure the inhibition of MEHP
at fixed substrate concentration, we found that MEHP competed
with NAD+ (Fig. 7). Therefore MEHP was a competitive inhibitor of
human and rat 11␤-HSD2, possibly by competing with the cofactor
NAD+.
4. Discussion
The present study showed that the phthalate diesters DPrP,
DBP and DCHP were inhibitors of both human and rat 11␤-HSD2
enzymes (Fig. 3). DBP was the most potent, with an IC50 = 13.69 ␮M
measured for the rat enzyme. The potency of DCHP for the inhibi-
tion 11␤-HSD2 was comparable to what has been reported [15]. The
distinguishing characteristic of these inhibitory diesters compared
to nine other phthalate diesters, was the approximate dimension
of the alcohol moieties. The three phthalates that were potent
inhibitors of 11␤-HSD2 had alcohol moieties that consisted of

B. Zhao et al. / Chemico-Biological Interactions 183 (2010) 79–84
83
Fig. 5. Dose–response curve for the inhibition of human and rat 11␤-HSD2 activities by MEHP. The measurement of 11␤-HSD2 was performed in the presence of 0.1–1 mM
MEHP according to Section 2. IC50s for human and rat enzymes are 110.8 10.9 and 121.8 8.5 ␮M, respectively. Mean SEM (n = 4).
We analyzed the kinetics of the inhibition by MEHP against
either CORT, the substrate, or NAD+, the pyridine nucleotide
cofactor. The inhibition by MEHP in the presence of increasing con-
centrations of CORT was non-competitive (Fig. 6). The inhibition
in the presence of increasing concentrations of NAD+ was com-
petitive for both human and rat 11␤-HSD2 (Fig. 7). We infer that
MEHP competes for the NAD+ binding site, because of a structural
similarity between MEHP and NAD+. Whether MEHP inhibits other
NAD+-dependent enzymes is currently under investigation in our
laboratory.
The IC50s of some phthalates for the inhibition of 11␤-HSD2
are around 10–100 ␮M. Although current environmental levels of
DEHP may not lead to accumulations of MEHP of this magnitude,
phthalate exposure can reach up to 167.9 mg/day during medi-
cal infusion [5]. It has been reported that infants were exposed to
very high levels of DEHP (42–142 mg/kg/day) during extracorporeal
membrane oxygenation [18]. The amount of phthalates transferred
through intravenous tubing could constitute a considerable body
burden to neonates. Furthermore, as phthalate monoesters have
been detected in most human samples [22], cumulative effects of
phthalates are possible. It has been found that in in vivo animal
models, phthalates have dose-additive cumulative effects on fetal
testis function [22].
An additional area for concern is the danger phthalates pose to
fetal development. In a recent case-control study, we showed that
low birth weight was associated with high levels of phthalates in
the mothers’ placenta [23]. Although direct inhibition of placental
11␤-HSD2 by phthalates has not yet been shown, circumstantial
evidence from other studies leads us to infer that this mechanism
may account for low birth weight in populations exposed to high
levels of these environmental pollutants. For example, high cortisol
levels in the presence of a compromised 11␤-HSD2 adversely affect
fetal development in several animal models and human conditions
[24,25]. Complete loss of, or even reduced 11␤-HSD2 activity causes
low birth weight and intrauterine growth retardation [26,27]. In
mothers with pregnancy-induced hypertension (preeclampsia) the
expression of angiotensin II receptor subtype I in the placenta
is increased compared to normal pregnancies [28], because the
downregulation of 11␤-HSD2 leads to increases in local cortisol
that upregulate angiotensin II subtype I receptor promoter via glu-
cocorticoid receptor element [29]. Diminished 11␤-HSD2 activity
that occurs in intrauterine growth retardation or in preeclampsia
Fig. 6. Dixon plots of human 11␤-HSD2 in the presence of MEHP. The measurement
of 11␤-HSD2 was performed in the presence of a range of CORT concentrations
according to Section 2.
Fig. 7. Lineweaver–Burk plots of rat 11␤-HSD2 in the presence of MEHP. The
measurement of 11␤-HSD2 was performed in the presence of a range of NAD+
concentrations according to Section 2, [CORT] = 10 nM.

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B. Zhao et al. / Chemico-Biological Interactions 183 (2010) 79–84
and that allows for more than 10–20% diffusion of maternal glu-
cocorticoids leads to increased fetal blood cortisol levels [30,31].
The normal role of 11␤-HSD2 in the placenta may be to con-
trol the availability of glucocorticoids, as 11␤-HSD2 is upregulated
during the development of the cytotrophoblast into the syncy-
tiotrophoblast [32,33], and compromised 11␤-HSD2 activity and/or
high cortisol is detrimental to both mother and fetus.
Phthalates also have been found to interfere with the devel-
opment of testis and male reproduction [34,35]. 11␤-HSD2 has
been demonstrated in fetal and adult mammalian (human, rat
and pig) Leydig cells in testes [36–38]. Therefore, 11␤-HSD2 plays
important roles in cortisol inactivation Leydig cells to prevent from
glucocorticoid-mediated suppression of testosterone production.
Suppression of 11␤-HSD2 in Leydig cells could lead to the suppres-
sion of testosterone production in these cells.
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In conclusion, we tested phthalates that varied in size and
shape, and found that phthalates that inhibited 11␤-HSD2 had
alcohol moieties that were intermediate in size to those on the
non-inhibitory phthalates. Inhibition was not strictly dependent
on the number of carbons in the moieties, but on some aspect of
the space-filling conformation of the moiety.
Conflict of interest statement
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Acknowledgement
The authors thank Chantal M. Sottas for technical support.
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