BIOLOGY
OF
46, 586-594
REPRODUCTION
Developmental
Regulation
Production
(1992)
of Sertoli
by Hormones,
and
ROSSELLI3
Endocrinology
Reproductive
Aromatase
Retinoids
MARINELLA
Department
Cell
the
and
Center,2
Activity
Testicular
MICHAEL
Plasminogen
Paracrine
Activator
Factor,
PModS1
K. SKINNER2
University
of Ca4fornia,
San Francisco,
Ca4fornia
Vanderbilt
University
School
of Pharmacology,3
and
San
94143-0556
of Medicine,
Francisco
Nashville,
Tennessee
37232-6600
ABSTRACT
Testicular
vitro.
peritubular
The
current
plasminogen
day-old
to be high
and
cells.
production
at various
was
the
to affect
aromatase
ability
of FSH
to stimulate
hormone
activity
at any
aromatase
of Sertoli
examined,
cells,
Insulin
duction
also
addition,
during
uPa
of FSH
data
activator
indicate
stages
that
tPa
by
prepubertal
production
Results
by
imply
that
such
as plasminogen
factor
to promote
and
Sertoli
cell
hormone
activator
responsiveness
two
functionally
[3]. The production
der the
veloped
control
of androgens
that the gonadotropin
drogen
increase
production,
the secretion
the maintenance
cytoarchitectural
developing
gerseminiferous
tu-
November
‘This
NIH
study
Sertoli
12,
of Califomia,
grant
on peritubular
which
in turn
been
for
was
not
to suppress
that
the
may
PM0dS
developmental
stages
in prepubertal
Sertoli
suppressed
PM0dS
cell
of hormones
was
that
support
pubertal
pro-
to have
appear
no
in part
the proposal
development.
as the
vary
uPa
found
functions
results
during
more
to
production
by Swiss
who
National
is a PEW
K. Skinner,
Francisco,
San
Foundation
cell
and
an
Scholar.
Reproductive
Francisco,
of M.R
Endocrinology
Center,
protein
regulatory
that
In
differentiates
FSH [3,4,7].
Sertoli
cell
(ABP)
agent
cells is the
P450-associated
shown
aromatization
enzyme
to stimulate
stage
Uni-
586
in-
of development.
particularly
during
the
to early
is
pubertal
activity
in prepubertal
the ability
of FSH to regulate
in more
adult
stages
of devel-
aromatase
in the early
as the Sertoli
to be important
tissue
remodeling
and
of androgens
to estrogen
aromatase
[8-10].
FSH has
aromatase
is postulated
needed
for
investigated
activity
is high
pubertal
stages
cell differentiates
Previously,
translocation
and
but
a crude
redeto a
mix-
proteins
(PSP) was shown
in midpubertal
Sertoli
cells
activity
cell
is due
product
for the proteolytic
of the seminiferous
of spermatogenic
across
the blood-testis
barrier
[13, 14]. Through
the levels of cAMP, FSH stimulates
Pa production
CA 94143-0556.
extent
identified,
The actions
of purified
PModS
on adfunctions
at various
stages
of pubertal
[9]. Therefore,
to hormones
to low levels
adult
to a greater
previously
[121. The possibility
that this inhibitory
PModS
is addressed
in the current
study.
Plasminogen
activator
(Pa) is a Sertoli
of transfer-
support
single
ture of peritubular
cell-secreted
to decrease
aromatase
activity
for the process
of sperparacrine
factor
PModS
cell
androgen-binding
any
opment
sponsive
creases
1991.
M.K.S.
Michael
San
Sertoli
Combined
differentiation
action
only
significantly
Purified
influence
of the
by FSH
Sertoli
cells [8-11];
however,
aromatase
activity
decreases
has decell an-
cells
modulates
and
Sertoli
by the
1991.
was supported
(HD20583)
‘Correspondence:
versity
24,
cells.
alone
found
imply
35.
found
to FSH in stimulating
aromatase
during
pubertal
development.
at each
retinol
and
was
and
was
in late-pubertal
PM0dS
Results
cells.
FSH
20-,
10-,
activity
development
has not been
thoroughly
the focus
of the present
study.
A cellular
function
primarily
attributed
cells
produce
a
effects
on Sertoli
been
purified
and
[2, 5]. The hypothesis
LH stimulates
Leydig
functions
important
[2,3, 6]. The purified
July
the
cluding
ditional
related
forms,
PModS(A)
and
of PModS
appears
to be un-
which
acts
of PModS,
midpubertal
Received
not
and
rin
of local
paracrine
factors
[1]. Peritubular
factor termed
PModS
[2] that has dramatic
cell function
in vitro
[3,4].
PModS
has
Accepted
cells,
Sertoli
may
than
bule
and interact
with
Sertoli
cells
through
the complex
extracellular
matrix
between
the cells.
In addition,
peritubular cells and Sertoli
cells interact
through
the production
stimulates
by
influenced
from
in
and
development.
Sertoli
cells play a fundamental
role in
of spermatogenesis
by creating
the proper
support
and unique
microenvironment
for
minal
cells. Peritubular
cells surround
the
Sertoli
cell
matogenesis
PMOdS
was
activity
to hormones
Interestingly,
function
aromatase
Aromatase
production.
stimulated
cell
Sertoli
isolated
estrogen
Sertoli
production.
Sertoli
cell
maintain
were
(uPa)
Sertoli
PMOdS
on
cell
that is less responsive
was found
to increase
midpubertal
INTRODUCTION
exists
in
PModS(B)
was
activator
effects
Sertoli
be nonresponsive
in midpubertal
(tPa)
on
cells
and
examined.
of development
activator
production
activator
dramatic
of development).
and
activity
production
plasminogen
tPa
production.
the
stage
production
stimulate
of cellular
differentiation
act as a differentiation
pubertal
developmental
estrogen
stages
to decline
of the
plasminogen
and
to
and
aromatase
of urokinase.type
stimulated
ability
on plasminogen
independent
PMOdS
may
tissue-type
cells
to influence
and
has
hormones
Sertoli
late-pubertal
found
activity
that
and
development.
and
Sertoli
PM0dS
of PModS
of pubertal
to a more
adult
activity,
plasminogen
production
the
and
effect
cell
whereas
termed
actions
midpubertal,
in prepubertal
only
Sertoli
cell differentiation
In contrast
to aromatase
Production
factor
the
stages
to prepubertal.
hormone-responsive
FSH
a paracrine
to examine
designed
correspond
found
promote
activity.
produce
was
activator
rats (ages
Sertoli
cells
study
to
that
activity
tubule,
cells
increasing
by Sertoli
PModS
cells [13,
production
tion
AND
PUBERTAL
SERTOLI
15]. Although
the developmental
regulation
of Pa
has not been
thoroughly
examined,
Pa produc-
appears
to
be
somewhat
independent
differentiation
and primarily
Previously
a crude
preparation
was not found
to influence
[16].
duce
Peritubular
an inhibitor
fects
of PModS
In the
study
reported
rified
PModS
on
cell
requires
elevation
of cAMP.
of PSP that contained
PModS
Pa production
by Sertoli
cells
cells,
however,
have
of Pa [14] that may
in crude
of Sertoli
secreted
here,
been
have
protein
we
examined
shown
masked
to prothe ef-
preparations
[16].
the
actions
of pu-
Pa production.
CELL
DEVELOPMENT
PModS
587
Preparation
PModS
was
serum-free
purified
AND
using
a membrane
with
sulfate
medium
precipitate
was applied
monium
tioned
umn.
The
cell
transferrin
active
with
a high-salt
Cell
Preparation
Sertoli
and
cells
and
peritubular
testis of rats 10, 20,
isolated
by sequential
trvpsin,
10
and
coliagenase,
procedure
short,
very
quired
ments
cells
per
Peritubular
by low-speed
a 24-multi-well
cells
1 x
i05
plate.
Cells
with
obtained
per
were
(1.7
rats,
re-
10%
100
ng/ml;
from
and
maintained
medium
changes
in Ham’s
F-12
and peritubular
newborn
calf
insulin,
treat5 x
testicular
cells were
cm2)
experiments,
the cells were
cultured
with
ulatory
agents,
including
FSH (National
MD),
a modified
resuspended,
cells
the
10-day-old
X g) were
of the
[3]. The
centrifugation,
in medium
Baltimore,
from
with
With
(20
were
supernatant
described
5% CO2 atmosphere
with
toli cells were
maintained
mented
with 0.01%
BSA,
tained
isolated
hvaluronidase,
described
[18].
centrifugations
digestion
as previously
at approximately
were
of age. Sertoli
cells were
digestion
[17], including
Sertoli
cells between
enzymatic
Cells were
plated
at approximately
well.
collagenase
preparation
cells
35 days
enzymatic
and
previously
low-speed
to sediment
and washes.
lected
Culture
1-mi
well
of
at 32#{176}C
in a
every 48 h. Sermedium
supplecells were
mainserum.
In some
one or more
regPituitary
Agency,
5 p.g/ml;
retinol,
0.35
i.LM; and testosterone
1 p.M. Where
indicated,
cells were
treated
with PSP at a concentration
of 50 p.g protein/ml,
or
PM0dS(A)
or PM0dS(B)
at a minimal
concentration
of 25
ng protein/mi.
Treatments
were
initiated
at the time
of
plating
and maintained
until the
Histochemical
analysis
of the
age
group
populations
termined
was
performed
of the
cell
utilized.
Germinal
cell contamination
was
with the method
of Berg [19], and peritubular
decell
contamination
was
chemical
localization
isoactin
tamination
toli cell
times
stated
in Results.
cultured
cells
from
each
[21]
to establish
determined
of desmin
the
purity
with
the immunohisto[20] and a-smooth
muscle
as previously
described.
after 5 days of culture
was
preparations
from
10-day-old
The germ
cell connot detectable
in Serrats and was 4% and
9% of the cell populations
from
20- and 35-day-old
rats,
respectively.
The peritubular
cell contamination
was less than
2% in the Sertoli
cell cultures
from 20- and 35-day-old
rats
and was 40% in cell preparations
from
10-day-old
rats.
column.
stored
of
and
applied
eluted
An am-
condiHPLC col-
bioassay
Bound
and
and
were
absorbance
at 214 nm,
acid analysis.
Aromatase
proteins
to two
with
Sertoli
applied
to a
were
successive
acetonitrile.
at -70#{176}Cbefore
Puuse,
in
biochemical
protein
assays,
and
amino
Activity
Aromatase
activity
from aromatization
3H]testosterone
viously
plated
by
collected
buffer
of PModS
limit.
the presence
of 1 mg/ml
BSA. The purity
of PModS
preparations
was determined
chromatographicallv
and electrophoretically
as previously
described
[3]. The mass of PModS
used was determined
with combined
procedures
including
the
cell
col-
forms
exclusion
Mr
was
columns
[3].
cell serum-free
con100-fold
by ultrafiltraCorp.,
Lexington,
MA)
determined
affinity
cell
described
of this
concentrated
to a size-exclusion
production,
C4 reverse-phase
rified
a 3 000
peak,
peritubular
as previously
Briefly,
freshly
collected
peritubular
ditioned
medium
was concentrated
tion with an Amicon
system
(Amicon
eluted
METHODS
concentrated
medium
heparin-sepharose
MATERIALS
from
conditioned
was
described
[22]
with
p.1 medium
was
removed
day
assay
and
of the
assessed
by the
the
from
replaced
following
the
release
of 3H20
7-dione
or [1 ,2ctBoston,
MA) as pre-
of [la-3H]andro-4-ene-3,1
(New
England
Nuclear,
modification:
1-ml
with
cell
500
culture
500
on
p.1 medium
the
con-
taining
1 p.Ci/ml
radiolabel.
After a 17-h incubation
at 32#{176}C
in a 5% CO2 atmosphere,
the cultures
were
sonicated
and
an aliquot
was removed
for DNA analysis.
The remaining
sample
was
tran-coated
Chemical
A charcoal,
transferred
charcoal
to a tube and incubated
with dex(0.2%
[wt/vol]
70K dextran
[Sigma
Co., St. Louis, MO], activated
10 mM Tris, and 1 mM
2% [wt/vol]
EDTA, pH
with Norit7.5) for 2 h
at 4#{176}C,
and then centrifuged
at 13 000 X g for 15 mm. Auquots of the supematant
containing
3H2O were then counted.
Blank values
were
established
from identical
incubations
in
the absence
expressed
per
of cells.
as counts
micrograms
trogen
assay
Aromatase
per minute
of DNA.
activity
(cpm)
(3H20
released)
was
and was normalized
To confirm
by the cells, an estrogen
of Sertoli
cell-conditioned
the
RIA was
medium.
production
used
of es-
with
a direct
Pa Assay
Pa activity
tometric
tioned
M, pH
Sigma);
was
assay
measured
[23].
medium
was
7.5, containing
0.01%
(v/v)
L-Leu L-Lys, p-nitroanilide,
ml fibrinogen
fragment
tissue-type
(tPa)
form
A 100-p.l
with
an
sample
indirect
spectropho-
of Sertoli
cell-condi-
incubated
at 37#{176}C
in 400 p.! Tris 0.10
0.13 p.M plasmmnogen
(bovine
plasma,
Tween
80; and
0.30
Sigma).
When
was added
for
of the
enzyme.
The
mM
S-2251
(D-Val
required,
0.06 mg/
the activation
of the
absorbance
change
ROSELLI
588
AND
SKINNER
TABLE
1.2
a.,
a
C-’
1.0
a)
-C
0.6
-
+
-
-
IU)
NDt
tPa (0.12 IU)
SSP (10 g)
ND
ND
(106
preparations
*Standard
0
C
/
#{149}E 0.2
L
-C
U)
a)
0
+
76.4
76.4
ND
246
(lU)
±
of tPa
54.4
492
46
(Boehringer,
±
46
Indian-
U .U
C
0
10
20
30
ug
DNA
40
50
0
50
100
150
nmol substrate
200
(S-2251)
Sertoli
ious
FIG. 1. Linearity
of aromatase
activity
and Pa activity
assays.
Aromatase
activity
in Sertoli
cells isolated
from 20-day-old
rats, with freshly
isolated cells plated at increasing
cell number
(p.g DNA/mI)
in the presence
of 0.5 .Ci/ml
3H-andro-4-ene-3,17-dione
after a 17-h incubation.
Pa activity
was assayed
with increasing
concentrations
of hydrolyzed
substrate
(nmol
substrate,
S-2251)
as detected
with an absorbance
at 405 nm.
after
a 6-h
incubation
conditioned
tected
was
media.
b
The
omitting
micrograms
dard
curve
measured
blanks
protease
from
the
assay
was
mixture.
present
amount
in the
of reacted
culture
well.
substrate
was
The
fragment
protein
method
was
lyophilized
concentration
of Bradford
was
and
stored
measured
A stan-
and
then
activity.
The
animals
incubated
aromatase
activity
(release
to
erally
within
used were
the linear
of
between
10 and 20 p.g DNA/well
and are
range
of the assay. The Pa assay was based
peptide
(S-2251)
assay [23). This Pa
assay
is linear
over a broad
range
of
concentrations
with regard
to the absor-
at 405 nm generated
from
1). The absence
of plasminogen
the
hydrolyzed
in the assay
product
is used
the background
non-Pa
proteolytic
activity,
negligible.
Fibrmnogen
fragments
have
previ-
ously
been
shown
to specifically
activate
tPa activity
[23].
The presence
of plasminogen
and fibrinogen
fragment
was
used
to specifically
detect
tPa. The presence
of plasmino-
10 mg
gen and the absence
of fibrmnogen
fragments
was used
to
detect
urokinase-type
plasminogen
activator
(uPa).
The use
of tPa and uPa standards
in the assay
is shown
in Table
1
-20#{176}C.
to the
[24].
a..
U
U)
(0
-C-’
a,
a.
C-,
C
Assay
-C-,
.
-.
E
‘0
DNA was measured
mide [25] as previously
icated
cell suspension
bromide
thymus
fluorometrically
with ethidium
brodescribed
[3]. An aliquot
of the sonwas added
to an equal
volume
of
solution
temperature
for 30
with 350 nM excitation
the culture
mately
0.1
at var-
was found
to be linear
to the micrograms
of DNA
in the culture
well (Fig. 1). The cell densities
gen-
C-
calf
plated
3H-androgen
3H20)
present
-I
ethidium
were
with
aromatase
to determine
which
was
with
at
according
midpubertal
assess
bance
(Fig.
of fibrinogen
(human
plasma,
Sigma)
was treated
with
13
mg of cyanogen
bromide
in 1 ml 70% formic
acid overnight
at room
temperature.
The solution
was diluted
1:10
with
water
and dialyzed
against
water
overnight.
The fibrinogen
from
densities
spectrophotometric
substrate
(S-2251)
deThe
obtained
by hydrolyzing
the substrate
S-2251
non-enzymatically
10 M NaOH.
For the preparation
of the fibrinogen
fragment,
cells
cell
on the hydrolysis
of a chemically
reactive
that can be detected
in a colorometric
without
assay were
calculated
in regard
to the
substrate
(S-2251)
and normalized
to
of DNA
of the
against
Pa-independent
plasminogen
results
of the activity
nanomole
converted
with
+
apolis, IN) and uPa (Sigma,
St. Louis. MO) were compared to Sertoli
cell-secreted
proteins
(SSP) in the Pa
enzymatic
assay in the absence or presence
of plasminogen and fibrinogen
fragment.
tND denotes
nondetectable
levels of Pa activity.
ai’
500
0
DNA
substrate).
Fibrinogren
0.4
a-,
the
converted
4-’
U,
U,
‘U
a
(nmol
Plasminogren
uPa
E
‘-‘
1000
CU
Pa activity
.4J
1500
a.
.4)
C-,
1.
and
allowed
to incubate
wells.
This
pg DNA.
was
assay
used
has
to quantify
a Sensitivity
DNA
levels
4-)
(U
a.
a-.
-
.4)
a.
-C
C
U)
C-,
-C
in
of approxi-
For the validation
of the activity
assays,
the linearity
of
both
the aromatase
and Pa assays
are shown
in Figure
1.
The aromatase
activity
assay was based
on the release
of
l-I2O from 3H-androgen
upon aromatization
to estrogen
[22).
#{149}
a
C
(U
.0
C0
U)
DC.0
a)
C,)
‘U
E
U)
4-,
(U
(U
a
0
0.
C-C
3
5
7
Day of Culture
3
5
7
Day of Culture
of aromatase
activity and Pa production
in Sertoli
samples/cells
collected
on Days 3, 5, and 7
of culture. Cells were incubated
in the absence
(0) or presence
(S) of 100
ng/ml
FSH at the time of plating.
The results are presented
for aromatase
activity
as fold increase
of activity (cpm/pg
DNA) present
in control cells
on Day 5 of culture.
The results are presented
for Pa activity
as the level
of converted
substrate
as assessed
with absorbance
at 405 nm. The mean
± SEM
is presented
from three different
experiments
done in triplicate.
FIG.
RESULTS
Q
C-)
4-)
C-)
a-,
at room
mm. Fluorescent
emission
at 585 nm
was then monitored.
A standard
curve
DNA
o
0
U-
cells from
2.
Time
course
20-day-old
rats with
PMocIS
AND
PUBERTAL
SERTOLI
Pa activity
present
in conditioned
rations
of cell culture
are shown
60
incubated
in the
7 days
E
C-,
absence
of culture.
Basal
FSH
>
-I-I
>
30
C-)
stimulation
pubertal
periments,
C’,
CD
4-s
for
were
and
was
similar
at each
time
late-pubertal
Sertoli
cells.
data were
obtained
on Day
The basal
production
20
-.
are
and FSH-stimulated
by Sertoli
cells
shown
the basal
prepubertal,
in Figures
levels
remained
midpubertal,
3). FSH-stimulated
30
Age (days)
40
FIG. 3. Aromatase
activity
in Sertoli
cells from various
stages
of pubertal
development.
Sertoli
cells were isolated
from
10-, 20-, and 35-dayold rats and cultured
for 5 days in the absence
(0) or presence
l#{149})
of 100
ng/mI
FSH. Sertoli
cells were incubated
in the presence
of 0.5 p.Ci/ml
3Handro-4-ene-3,17-dione
for 17 h and subsequently
were
sonicated;
the
amount
of 3H20 release
(cpm) was detected
with the charcoal
assay. Results are presented
as cpm/g
DNA and are the mean ± SEM from a minimum
of three different
experiments
done in triplicate.
duwere
3, 5, and
found
to
point
examined
is shown
analysis
and for Pa
Preliminary
with pre-
For subsequent
5 of Sertoli
cell
excul-
0.
a-,
from
3 and
aromatase
activity,
aromatase
activity
was
highest
the
(Fig.
with
pre-
development,
uPa was found
to increase
slightly,
and it was
responsive
to FSH only at the prepubertal
stage
of development
Basal levels of tPa production
were lower than those
of uPa and increased
slightly
with
pubertal
development.
FSH was found
to stimulate
tPa production
by prepubertal,
midpubertal,
and late-pubertal
Sertoli
cells. As previously
of Sertoli
A more
U)
4. For
relatively
constant
between
and late-pubertal
Sertoli
cells
tPa is the form
of Pa that is hormone
responuPa is nonresponsive
to FSH in midpubertal
and late pubertal
Sertoli
cells.
tPa was found
to be independent
a.
levels of aromatase
and Pa
10-, 20-, and 35-day-old
pubertal
Sertoli
cells, reduced
with midpubertal
cells, and
absent
with late-pubertal
cells. Therefore,
as the Sertoli
cells
reached
a more
adult stage of differentiation,
aromatase
activity became
nonresponsive
to FSH. The Pa production
was
separated
into uPa and tPa activities
(Fig. 4). With pubertal
shown
[26],
sive, whereas
CU
‘U
0.
cells.
detailed
analysis
The
of the
hormone
stimulation
of the differentiated
regulation
of
stage
of Pa produc-
U)
tion is shown
in Figures
5 and 6. Both
midpubertal
and
late-pubertal
Sertoli
cells were
nonresponsive
to FSH in regard to uPa production;
interestingly,
however,
FSH signif-
0.0
a.’
CO
a.
and
ture.
rats
‘C-,
CU
a.
-
a.
u
C-,
C
C
C
,
C
U
a
DC
Co
U,
a
Co
Co
0.
of FSH
of aromatase
of aromatase
activity
was on Day 5 of culture
production
on either
Day 5 or Day 7 of culture.
analysis
indicated
that similar
data were
obtained
-I-f
DC
or presence
levels
at various
2. Cells
(Fig. 2). A more
thorough
analysis
with statistics
in subsequent
figures.
Data indicated
that optimal
-1_J
C
U,
medium
in Figure
“Pa
remain
relatively
constant,
and optimal
FSH stimulation
was
observed
on Day 5 of the culture
(Fig. 2). Basal
levels
of
Pa production
were
found
to increase
during
culture,
and
50
.
589
DEVELOPMENT
to demonstrate
the specificity
of the Pa assay. Hereafter,
activity”
will denote
tPa and uPa activity
combined.
The aromatase
activity
in midpubertal
Sertoli
cells
70
0’)
CELL
icantly stimulated
uPa production
by prepubertal
Sertoli
cells
(Fig. 5). As indicated
above,
FSH stimulated
tPa production
-
a
from
U)
10
20
Age
30
(daysl
40
#{176}-
-
10
20
Age
30
40
(days)
FIG. 4. Pa production
by Sertoli
cells from various
stages
of pubertal
development.
Sertoli
cells were isolated
from 10-. 20-, and 35-day-old
rats.
and a 72-h-medium
collection
was obtained
on Day 5 of culture
with cells
cultured
in the absence
(0) or presence
(#{149})
of 100 ng/ml
FSH. Both uPa
and tPa activity
was examined.
Results
are presented
as nmol converted
substrate/ij.g
Sertoli
DNA and are the mean of a representative
experiment.
Sertoli
cells
at all developmental
stages
examined
(Fig.
6). Insulin
at high concentrations
(5 p.g/ml/ml),
probably
acting
at the insulin-like
growth
factor-I
(IGF-I)
receptor,
stimulated
both
uPa and tPa production
from
prepubertal
Sertoli
cells.
fects observed
cells.
Retinol
This was in contrast
with
midpubertal
alone
had no effect
duction,
it
but
did
significantly
to the lack of insulin
efand late-pubertal
Sertoli
on prepubertal
uPa pro-
inhibit
basal
uPa
production
590
ROSELLI
AND
SKINNER
35 Day
20 Day
10 Day
(0
0
60
50
0)
40
Cl)
60
I
>-
50
40
T
-I-)
Co
a.
I
30
-
30jj
20
0
C
10
C
E
U,
(0
CF
a-
I
-
ATM
-_
C Fl
FIG.
5.
Effect
of regulatory
ATM
C F IA
AB
agents
on uPa production
by Sertoli
TM
AB
TREATMENT
TREATMENT
TREATMENT
cells
isolated
from
10-, 20-, and
35-day-old
rats.
Cells were cultured
in the absence
(Control,
C) or presence
of ‘tOO ng/mI
FSH (F); 5 .g/ml
insulin
(I), 0.35 M
retinol
(R); 1 M
testosterone
(T); a mixture
of FSH, insulin,
retinol,
and testosterone
(M); and 25 ng/mI
PModS(A)
(A) or
PModSIB)
(B). Medium
collected
on Day 5 of culture
was assessed
for activity
and expressed
as nmol
converted
substrate/g
Sertoli DNA. Results are presented
as the mean ± SEM from a minimum
of three different
experiments
done in triplicate.
‘Statistically
different
from control,
with p < 0.05 using Student’s
t-test.
by niidpubertal
Sertoli
duction
by late-pubertal
not statistically
significant
fect on basal tPa production,
to stimulate
6). Therefore,
tion
and
fects
and
tended
to reduce
uPa
was approximately
influence
Sertoli
pro-
cells,
although
the reduction
was
(Fig. 5). Retinol
alone
had no efbut inhibited
the ability
of FSH
opmental
FSH-stimulated
The
for
tPa
minimum
and
this inhibitory
respectively
(Fig.
of retinol
production
on
7). The
midpubertal
by
maximum
effect
IC0
Sertoli
for
the
cell
10 Day
fluence
retinol
nM and
inhibitory
250 nM. Testosterone
cell Pa production
examined
PM0dS(A)
Sertoli
cells
the actions
is shown
Pa production
found
5 and
PModS(B),
6). The
were
8. FSH
prepubertal
the
and
midpubertal
(0
0
-4-)
-4.,
0)
Cl,
.1-I
CO
a.
-
-C
r
Cells
CFIRTP4AB
TREATMENT
FIG. 6. Effect
were cultured
of regulatory
and treated
CFIRTMAB
CFIRTMA
TREATMENT
agents
on tPa production
and the data are presented
by Sertoli
cells isolated
from
as described
in the legend
TREATMENT
10-, 20-,
to Figure
and
5.
also failed
Pa production
35-day-old
rats.
to
activity
regulatory
35 Day
20 Day
to inor late-
of aromatase
only
forms
found
midpubertal,
regulation
was
purffied
not
(Figs.
5 and 6). PModS
of FSH on Sertoli
cell
of the
in Figure
to stimulate
was not found
to
at any of the devel-
by prepubertal,
(data
not shown).
A detailed
analysis
ef-
(Figs.
and
Pa production
pubertal
influence
midpubertal
effective
were
100
stages
of PModS,
tPa production
by midpubertal
Sertoli
cells (Fig.
retinol
inhibited
basal levels of uPa produc-
Sertoli
cells.
concentrations
1 p.M,
cells
Sertoli
agent
cell
PModS
AND
PUBERTAL
SERTOLI
CELL
aromatase
>
100
-I-,
591
DEVELOPMENT
activity.
Insulin,
retinol,
and
testosterone
had
effect on aromatase
activity at any of the developmental
examined
(Fig. 8). A combination
of FSH, insulin,
‘
no
stages
retinol,
-I-,
and testosterone
the regulatory
matase
activity
4-,
C-)
C0
-s-I
c
tions of regulatory
agents
used
to be optimal
for the regulation
0
functions
-I-I
.-
.0
41
#{149}1I
also
of the
shown
actions
The
II
same
tions
0)
0
the
cells
2:.
bertal
with
Neither
the
Sertoli
PM0dS(A),
0
1
2
Retinol
3
4
cells
and
50-ng/ml
To
(uM)
FIG. 7. Inhibitory
effects
of retinol
on Pa production
by Sertoli
cells
isolated
from 20-day-old
rats. Cells were incubated
in the presence
of FSH
and increasing
concentrations
of retinol
(ij.M), and a 72-h-medium
collection was obtained
on Day 5 of culture
to determine
Pa activity
for both uPa
(0) and tPa (#{149}).
Results
are presented
as the percentage
of inhibition
and
are the mean from a representative
experiment.
All treatments
were found
to be statistically
different
(p < 0.05) from control
non-retinol
treated
cells
with Students
t-test.
Day
matase
10).
A similar
was
actions
was
assessed
ditioned
treatment
medium
conditions
from
of FSH
Sertoli
not
activity
was
or
late-pubut
on midpubertal
were
observed
in the
3H-testosactivity
of FSH-stimulated
midpubertal
had
aro-
Estrogen
Sertoli
production
no
20-
PM0dS(B).
aromatase
PModS(B).
not
Ser-
with
and
of PM0dS(B),
estrogen
of
found
statistically
ac-
cell-con-
Days 2-5 of culture
with the
(Fig. 10). PModS(B)
also inhibited
PM0dS(A)
To
activity,
or presence
prepubertal
inhibition
with
pro-
shown).
of PModS
of PM0dS(A)
to stimulate
cells.
(data
of FSH
effects
in
concentra-
transferrin
PModS(B),
as a substrate
cumulation
same
absence
on
observed
20 Day
I
a,
I
-..
W
“V
*
a..
same
the
by midpubsignificant
35 Day
80
80
60
60
I
4-,
-I,
-
used
activity
ability
in the
of FSH
inhibitory
also
PModS(B)
on aromatase
forms
Similar
is
,.
8u
E
0.
--
(Fig.
cells
Interestingly,
the
9).
at the
of the
concentrations
this
was
assay
ertal
10
(Fig.
confirm
terone
actions
activity
nor
stimulate
actions
FSH
cells.
inhibited
toli
PModS
of PModS
9).
aromatase
PM0dS(A)
optimally
treated
(Fig.
to influence
0
been
shown
of Sertoli
cell
levels of aromatase
activity
or late-pubertal
Sertoli
cells.
Sertoli
analysis
were
PMOdS
I-I
to
by midpubertal
extend
c
basal
of
found
on
8. Neither
to affect
midpubertal,
preparations
were
duction
of PModS
in Figure
alone
was found
in prepubertal,
C-)
have previously
of a number
[27].
Analysis
CO
U)
CD
had similar
effects
as FSH alone.
None
of
agents
including
FSH had any effect on aroin late-pubertal
Sertoli
cells. The concentra-
40
*
nflM
#{241}#{241}#{241}#{241}#{241
#{241}#{241}
40
4-’
C.)
:
40
2:
(0
E
.
a
T
II
II
L
-C
II
20n
“
CF
IA
TM
AB
TREATMENT
FIG. 8. Effect of regulatory
agents
rats. Aromatase
activity
was determined
and presented
as cpm/p.g
Sertoli
DNA.
the legend
to Figure
5.
C
F
#{241}#{241}#{241}
I
A
T
N
AS
CF
TREATMENT
IA
TM
AS
TREATMENT
on aromatase
activity
in Sertoli
cells isolated
from
10-, 20-, and 35-day-old
on Day 5 of culture
after a 17-h incubation
with ‘H-andro-4-ene-3,17-dione
Cells were cultured
and treated
and the data are presented
as described
in
ROSELLI
592
AND
SKINNER
20 Day
10 Day
35 Day
0)
0.
C.)
a..
4-’
-‘-4
a.
--S
C.)
-C
a,
U,
(0
4-)
(0
a
C-C
F
FA
TREATMENT
C
FB
C
F
FA
TREATMENT
FB
C
F
FA
TREATMENT
FB
FIG. 9. Effect of PM0dS
on FSH regulation
of aromatase
activity
in Sertoli
cells. Cells were isolated form 10-, 20and 35-day-old
rats and cultured
for 5 days in the absence
(control,
C) or presence
of FSH (F); FSH plus PModS(A)
(FA); or FSH plus PModS(B(
(FBI. Aromatase
activity
was determined
after a 17-h incubation
with ‘H-andro-4-ene3,17-dione
and presented
as cpm/p.g
Sertoli
DNA. Results
are presented
as the mean
± SEM
from a minimum
of
three different
experiments
done in triplicate.
The letter inset within
bars denotes
the results
of a statistical
analysis
of variance,
with a difference
in letter indicating
a statistical
difference
(p < 0.05).
effect
on
production,
FSH actions
with
aromatase
activity
or
but did have slight
inhibitory
actions
and
can
lOB).
reduce
Data
the
and
estrogen
indicate
that PModS(B),
ability
of FSH to stimulate
production
estrogen
(Figs.
9
but not PM0dS(A),
aromatase
activity
by midpubertal
Sertoli
cells.
testicular
dramatic
production
Sertoli
cell
investigated
paracrine
effects
on
in vitro
functions
factor
Sertoli
[3, 4, 7). The
have
not
in the
study
reported
the potential
physiological
role
identified
Sertoli
cell functions
to aromatize
enzyme
procedures
androgen
were
scribed,
and
activities
consider,
here
to help
other
were
with
the
P450
and
culture
were
determined.
however,
is the
validated
conditions
An
purity
as
for
previously
dethese
experimental
variable
to
of the cell populations
used.
Peritubular
cells produce
PModS
and are the primary
somatic cell contaminant
of Sertoli
cell preparations.
As previously
described
[7],
both
midpubertal
and late-pubertal
Sertoli
cell
cultures
contain
a low
level
(less
peritubular
contaminant.
However,
prepubertal
from
10-day-old
rats contain
a higher
level
(approximately
40%),
presence
of peritubular
zation
and has been
tion
and
hormone
which
must
be
cells interferes
shown
to influence
responsiveness
than
effects
of extracells can also in-
of added
PModS.
In addition,
matrix
derived
from
peritubular
2%)
in subsequent
preparations.
Sertoli
of
Sertoli
cells
of contaminant
been
for
the
garded
as a requirement
in the
adult
for
spermatogenic
primary
Sertoli
both
agent
been
shown
as the
blood-testis
controlling
to be relatively
presented
that FSH
prepubertal
can
in the
significantly
Sertoli
cell
toli cell becomes
ulate uPa expression
throughout
indicated
for
Pa
is
translocation
barrier
of
[13].
production
FSH
[13, 15]; however,
insulin
can
tPa by prepubertal
actions
stimulate
preparation.
pubertal
that
study,
The
of Pa ac-
nonresponsive
current
re-
development
to FSH
however,
The
uPa
[26].
indicate
uPa production
by the
Therefore,
as the Ser-
differentiated,
the ability
of FSH to regis lost; in contrast,
tPa is regulated
by
that
These
of
pubertal
such
the
and
is FSH through
the actions
of cAMP [13, 15, 26].
of Pa activity
influenced
by FSH is tPa, whereas
Results
FSH
processes
uPa
required
expression
throughout
cell
with Pa and its
to be impor-
remodeling
the
across
regulatory
tivity
form
has
cells
tion
potential
prepubertal
to produce
tissue
Therefore,
[7]. The
[2, 28, 29]. The
on
shown
continuous
with data normaliSertoli
cell func-
considered
of data
have
spermatogenesis.
contaminant
may mask
the
hormone
responsiveness
[30]. Therelimitation
needs
to be considered
analysis
cells
the
tPa [14]. The proteolytic
activity
associated
ability
to activate
plasminogen
are postulated
and
aromatase
examining
effects
cellular
tant
elucidate
and the production
of Pa [13]. The
the levels of aromatase
and Pa ac-
established
optimal
on
and
of PModS.
Two previously
are the ability
of the cells
to estrogen
complex
[8,9)
for assessing
tivities
PModS
has been
shown
cell transferrin
and ABP
actions
of PModS
been
examined
cell
fluence
Sertoli
cell
fore,
this experimental
DISCUSSION
The
to have
production
of PModS
by this peritubular
in the prepubertal
Sertoli
cell preparation
development.
is the
primary
results
stimulate
Sertoli
of insulin,
Previous
regulator
of the
current
the
production
cells
to the
probably
reports
study
of both
same
acting
have
of Pa produc-
degree
at the
indicate
uPa
and
as FSH.
IGF-I
re-
PModS
AND
PUBERTAL
SERTOLI
B
A
140
CC
120
b
Co
0.
C-)
a..a-’
a.
CC
0.
100
60
U
U)
CU
40
C-)
‘0
0
C0.
C
U
DC
C
C-
(0
a
0
C-
20
.4-,
U)
LU
-
t,_
C
F
FA
FB
F
C
TREATMENT
FA
FB
TREATMENT
FIG. 10. Effect of PM0dS
on FSH regulation
of (A) aromatase
activity
and (B) estrogen
production
by Sertoli
cells isolated
from 20-day-old
rats.
Cells were cultured
in the absence
(Control,
C) or presence
of FSH (F); FSH
plus PM0dS(A)
(FA); or FSH plus PModS(B)
(FB). Aromatase
activity
was
determined
on Day 5 of culture after a 17-h incubation
with 3H-testosterone
and data are presented
as cpm/g
Sertoli DNA. Estrogen
production
was
assessed
with a 72-h-medium
collection
on Day 5 of culture
and is expressed
as pg/pg
Sertoli DNA. Results are presented
as the mean ± SEM
from a minimum
of three different
experiments.
The letter inset within the
bars denotes
the results of an analysis
of variance,
with a difference
in
letter indicating
a statistical
difference
(p < 0.05).
ceptor,
are
midpubertal
sults
ertal
lost
and
imply
Sertoli
as the
Sertoli
cells
late-pubertal
are
stages
differentiated
that the regulation
of Pa production
cells is distinct
from the predominant
possibility
Sertoli
cell
on peritubular
that
insulin
preparation.
cells have
may
Although
not been
mediate
to the
of development.
ulation
of tPa production
observed
at later stages
development.
A limitation
of this interpretation
however,
is the presence
of peritubular
cells
pubertal
insulin
however,
peritubular
a Pa inhibitor
[14]
of PModS
in the
study demonstrated
and
cells
have
been
may mask
the
that
crude
preparation
that the purified
PM0dS(B),
had
no
of PSP [161. The
forms of PMOdS,
effect
on
Pa production
by cultured
Sertoli
cells from any of the developmental
stages
examined.
This is the first Sertoli
cell function
observed
that is not influenced
by the actions
of PModS.
The lack of
Pa regulation
by PModS
suggests
that PModS
is not a gen-
a-’
a
593
Recently,
produce
PM0dS(A)
C
0
80
a-,
C-,
-C
actions
current
DC
b
DEVELOPMENT
cells [16].
shown
to
IOU
a
CELL
its
Re-
by prepubFSH regof pubertal
of data,
in the pre-
the actions
documented,
effects
of
the
indirectly
through
the peritubular
cell requires
further
investigation.
As previously
reported,
androgens
do not influence
Pa
production
with the culture
procedures
utilized;
however,
retinol
inhibited
basal
uPa production
and FSH-stimulated
tPa production
by midpubertal
Sertoli
cells. The actions
of
retinoids
on other
Sertoli
cell functions
have previously
been
demonstrated
[27], but this study
appears
to be the first to
report
inhibitory
activity
associated
with
retinol.
The inverse
actions
of FSH and retinol
provide
a potentially
efficient
mechanism
for regulating
Pa production.
Further
investigation
of the actions
of retinoids
on Pa production
is
required
to address
questions
such
as why the inhibitory
response
was primarily
isolated
to midpubertal
Sertoli
cells.
Pa production
was found
to increase
slightly
with the more
advanced
developmental
stages
of cultured
Sertoli
cells, and
FSH increased
tPa production
by prepubertal,
midpubertal,
and late-pubertal
Sertoli
cells. Therefore
the production
and
regulation
of Pa production
appears
somewhat
independent of the differentiation
stage of the Sertoli
cells.
Previously,
a crude
preparation
of PModS
was found
to
have no influence
on Pa production
by midpubertal
Sertoli
eral activator
be primarily
of all Sertoli
cell functions,
involved
in influencing
the
the
possibility
cell.
This
other
Sertoli
The ability
is more
directly
cell function
examined,
of freshly
isolated
[10]
but instead
differentiation
addressed
may
of
with
aromatase
and cultured
the
activity.
[8, 9] Ser-
toli cells to synthesize
estrogen
has been
shown
to be due
to the aromatase
enzyme
complex
associated
with the endoplasmic
reticulum
posed
for estrogen
lation of Leydig cell
cells
The
have
also
physiological
cells
the
remains
of the cell. The initial
function
proproduction
by Sertoli
cells was regusteroidogenesis
[8-11];
however,
Leydig
been
role
to be
primary
shown
to produce
estrogen
for estrogen
production
elucidated.
regulator
FSH
of estrogen
has
been
PM0dS(A)
fluence
stages
Therefore,
aromatase
cell differentiates.
The
and
PModS(B),
alone
on aromatase
activity
examined.
A limitation
to be
and
aroma-
is a Sertoli
cell
stages
of puberty
[9]. Aromatase
activity
was found
to be high and
responsive
in prepubertal
Sertoli
cells but declined
not responsive
to hormones
at the late-pubertal
development.
as the Sertoli
[11,31].
Sertoli
shown
production
tase activity
[8-10,
22]. Estrogen
production
function
that is predominant
at the early
by
hormone
and was
stages
of
activity
is suppressed
purified
forms of PModS,
were
found
to have
no
in-
at any of the developmental
of this interpretation
of the
data, however,
is the presence
of peritubular
cells
in the
prepubertal
Sertoli
cell preparation
that produced
PModS.
Although
the levels
of PModS
present
in unconcentrated
conditioned
media
ble, the possibility
from
these
preparations
that endogenous
PModS
fects of exogenous
inability
of PModS
PModS
remains
to be investigated.
to directly
influence
aromatase
probably
levels.
correlates
PModS
has
with
the
been
Lack of effect
shown
levels,
but instead
stimulates
tivity in several
cell types,
[10,31],
and human
adipose
to have
effects
PModS
alone
may
aromatase
activity.
of PModS
hibited
the
midpubertal
and
were
used
FSH
demonstrated
to stimulate
PModS(A)
in the
significant.
PModS(B)
analysis,
The
activity
of PMOdS
on cAMP
no
on
effect
levels
PModS
not have the
Examination
ability
of FSH
Sertoli
cells.
that was not statistically
of both PM0dS(A)
and
negligimask
ef-
cAMP
cGMP levels [4]. Aromata.se
acincluding
Sertoli
[9, 22], Leydig
cells [32], has been
shown
to
be regulated
by alterations
in cAMP
pendent
protein
kinase
activity.
Since
on cAMP,
influence
were
may
and cAMP-dehas no effect
ability
of the
that
to directly
combined
PModS(B)
aromata.se
activity
caused
a suppression
inin
Optimal
concentrations
from various
preparations
as previously
demonstrated
[3].
594
ROSELLI
This
difference
in the
the first functional
forms
of PModS.
is required
actions
to understand
the
mone
responsive.
shown
to inhibit
with midpubertal
between
of the
significance
activities
Sertoli
is less
be due
to the
presence
appear
to be primarily
current
is an early pubertal
Sertoli
cells. Both
regulated
by FSH.
Analysis
are
and
addition,
the
required
actions
to further
our
of this testicular
observations
made
understanding
paracrine
with
PModS
of the
factor.
In
in the
current
study
utilized
an in vitro
analysis
of Sertoli
function;
elucidation
of the physiological
importance
and function
of
PModS
will ultimately
require
in vivo physiological
studies.
JH, Fritz
Biol
1978;
Reprod
activity
11. Mallea
218:143-
Byron
Glenn.
Lisa
Ha!burnt,
assistance
and
Jeff
Margaret
tance
in preparation
of this
manuscript.
derbilt
Reproductive
Biology
Research
Parrott.
Peach
and
Dr. Catherine
and
Claudia
We acknowledge
the
T. Anthony
Schumann
for
assistance
of the
1. Skinner
MK.
Cell.cell
MK.
Fritz
gen
control
that
interactions
modulates
perituhular
Sertoli
cell
Rev 1991:
secrete
functions.
Proc
NatI
under
Acad
S. produced
l3iol
hy testicular
Chem
4. Norton
tions
PM. Anthony
1988;
peritubular
cells
of the paracrine
that
modulate
factor
and
Sertoli
cell
MR.
a testicular
paracrine
of Sertoli
factor
cell
P-Mod-S.
differentiation
Endocrinology
1989:
cell
l’l.
of epidermal
by calcium.
FEB
1987;
secrete
a protein
activity
under
an-
in Sertoli-cells.
En-
activator
by Sertoli
cell
9:227-236.
tB. Rat testicular
peritubular
activator
Biol
activity.
cells
Reprod
in culture
1988;
38:359-
l( Hormonal
tubal
influences
segments
lB. Identification
Fritz
cells.
JH,
Cell
Roller
of Sertoli
PS, Skinner
1986;
cell
of
paracrine
testicular
follicle
Cell
enriched
activator
of the
seminif-
factor
involved
peritubular
cells
44:85-97.
LB. Effects
Mol
cycle
61:553-560.
between
LB. Fibronectin
in Sertoli
of the
of a non-mitogenic
Fritz
Fritz
1983;
interactions
preparations.
MK,
of plasminogen
stages
Biol
Endocrinol
NF,
cell
contaminants
Cell
cell
Mol
on formation
at defined
stimulating
Endocrinol
hormone
1975;
synthesis
is a marker
cultures.
Biol
on
3:57-70.
for
Reprod
peritubular
1984;
30:199-
19. Berg
JW. Differential
1963:
20. Anthony
CT, Skinner
ticular
peritubular
21. lung
lB.
1990;
cells.
of testis.
Am J Clin
Path
muscle
isoactin
R, Heather
1989;
of rat
testicular
is a specific
hormone
tures.
1980;
Endocrinology
(en JH, Mullaart
spectrophotometric
Bradford
MM.
quantities
of protein
effects
characterization
of tes-
40:811-823.
peritubular
myoid
differentiation
cells
marker.
in
Biol
Re-
for
GTG.
and
Kiuft
extrinsic
in plasma.
A rapid
An
alternative
of aromatase
method
activity
for
the
in Sertoli
study
cell
cul-
107:464-471.
E, Chang
assay
to measurements
J.
M, Dorrington
of follicle-stimulating
23. yerhei
Reprod
42:351-365.
Gore-Langton
Thromb
sensitive
utilizing
C, Wijngaards
(tissue-type)
G. A simple,
plasminogen
Haemostatis
method
the principal
for
1982;
the
appli-
48:226.
quantitation
of protein-dye
sensitive
activator
of microgram
binding.
Anal
Biochem
72:248-252.
Karsten
U, Wollenberger
direct
Anal
in sections
and histochemical
Biol
Characterization
a-smooth
prod
25.
of spermatozoa
MK. Cvtochemical
mvoid
PS, Fritz
culture:
22.
staining
23:513-515.
fluorometric
Biochem
Hettle
A. Improvements
estimation
1977:
AJ, WaIler
MK. Griswold
is
and
RNA
in cell
bromide
method
tissue
homogenates.
and
for
77:464-470.
ER, Fritz
lB.
MD.
regulated
by
Hormonal
by Sertoli
activator produced
Skinner
in the ethidium
of DNA
Secretion
hormones
stimulation
cells.
Biol
of testicular
and
alters
Reprod
the
1986;
transferrin
retinoids.
Biol
wpe
of plasmin-
34:895-904.
by cultured
Reprod
Sertoli
1982;
27:211-
28.
Tung
PS. Fritz
1980;
23:207-217.
lB. Interaction
Hutson
JC, Stocco
binding
protein
DM.
of Sertoli
Peritubular
secretion
bs Sertoli
cells
and mvoid
cells
in vitro.
cell
influence
on
cells
in culture.
Endocrinology
Biol
the efficiency
Reprod
of androgen
1981:
108:1362-
1368.
Dvm
M, Lamsam-Casalotti
membrane
the ac-
of Sertoli
siveness
124:2711-
5, Jia
increases
MC,
G-protein
cell
Kleinman
levels
adenvl
cvclase
and
HK,
Papadopoulos
follicle-stimulating
activity.
V. Basement
hormone
Endocrinology
1991:
respon.
128:1167-
1176.
MR.
hy perituhular
6. Skinner
of stimulatorv
action
211.
27 19.
S. Skinner
PS, Fritz
MK,
Sertoli
18. lung
function.
through
1977:
of plasminogen
Can J Biochem
cultures
30.
Regulation
E, lung
testis
17. Dorrington
PMod-
263:28&i-2890.
JN, Skinner
of
CF. Purification
cells
of plasminogen
Endocrinol
in mesenchvmal-epithelial
29.
MR. Fetterolf
cells
of aromatase
lB. Secretion
Cell
epithelium.
16. Skinner
1985:
82:114-118.
3. Skinner
Modulation
221.
andro-
Sci USA
Mol
LB. Karmallv
erous
12:45-77.
a protein
aro-
123:2100-2110.
an inhibitor
cells
Endocr
synthesis.
of rat testicular
inhibitory
in Sertoli
peritubular
induction
FE. Fritz
cultures.
by cultured
27.
cells
estradiol-
1985; 116:38-46.
and
activity
J. Testicular
AJ, Balek(ian
15. Fritz
26.
in the testiv
lB. Testicular
localization
FFG,
(FSH)
aromatase
inhibits
1988;
M, Smith
ogen
2. Skinner
stimulates
72:2677-2681.
371.
Van-
REFERENCES
factor
development.
of testicular estrogen
F, Rommerts
hormone
on
that
docrinologv
13. Lacroix
assis-
Center.
1975:
Endocrinology
AJ, Navaroli
(EGF)
control
1976;
technical
paracrine
pubertal
hormone
PNAS
DT. Control
development.
G, Caillea
drogen
24.
We thank
testicular
147.
12. Verhoeven
ACKNOWLEDGMENTS
expert
cells.
DR. Dufau ML. Cellular
during
factor
Follicle-stimulating
lB. Armstrong
of follicle-stimulating
growth
of the
throughout
18:55-64.
LE, Machado
action
DT.
Sertoli
C, Aquilano
10. Tsai-Morris
Actions
secretion
129:353-360.
in cultured
9. Dorrington
MK.
trarisferrin
Armstrong
synthesis
cable
for
cell
1991;
JH,
matase
M, Skinner
Sertoli
Endocrinology
secrete
analysis
of the
and Pa produc-
Rosselli
on
8. Dorrington
14. Hettle
of PMOdS
actions
suggests
that a potential
function
of PMOdS
may be to promote
and maintain
Sertoli
cell differentiation.
This is supported
by the observation
that PModS
had no
effect
on Pa production
and suppressed
aromatase
activity
in response
to FSH. Characterization
and molecular
cloning
of PModS
functions
CT,
(PModS)
enriched
of pubertal
development.
Pa producto be independent
of Sertoli
cell dif-
whereas
aromatase
activity
that decreases
in late-pubertal
7. Anthony
cell
hor-
of PModS.
in the
SKINNER
17
difference
PModS
may promote
pubertal
state that
may
stages
in part
ferentiation
cell function
two
forms
Previously
a crude
preparation
of PSP was
the actions
of FSH on aromatase
activity
Sertoli
cells [12], and it was suggested
that
activity
at various
appears
the
two
of this
The actions
of purified
PModS(B)
observed
study
support
this previous
speculation.
The current
study
provides
a thorough
hormonal
regulation
of aromatase
activity
tion
tion
is
of PModS(B)
to suppress
the accells suggests
an alteration
in
to a more
adult
stage
of devel-
opment.
Results
imply that
differentiation
to a late-stage
inhibitory
on aromatase
difference
observed
Further
characterization
in bioactivity.
The ability
tions of FSH on midpubertal
Sertoli
cell differentiation
this
of PM0dS(B)
AND
Fritz
lB. Androgen
cells.
51K. Cell-cell
Mol
Cell
interactions
stimulation
Endocrinol
of Senoli
1985:
in the testis.
cell
function
is enhanced
31.
mone
0:1l5-122.
Ann
NY Acad
Valladares
Sci
1987;
513:158-
32.
LE. Payne
in mature
Simpson
of estrogen
ER, Merrill
Al-I. Induction
rats.
of testicular
Endocrinology
JC, Hollub
biosynthesis
b
1979;
aromatization
AJ, Graham-Lorence
human
adipose
b
luteinizing
hor-
105:431-436.
cells.
S. Mendelson
Endocr
CR. Regulation
Rev 1989;
10:136-148.