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Regulates Plasminogen Activator Inhibitor-1 in Rat Testicular Peritubular Cells1
INSERM U-407, Bâtiment 3B, Centre Hospitalier Lyon-Sud, Pierre-Benite; and Laboratoire d’ Hématologie, Centre Hospitalo-Universitaire La Tronche (G.P., L.K.), Grenoble, France
Address all correspondence and requests for reprints to: Dr. Brigitte Le Magueresse-Battistoni, INSERM U-407, Bâtiment 3B, Centre Hospitalier Lyon-Sud, 69495 Pierre-Bénite Cedex, France.
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Abstract |
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(TNF) of
plasminogen activator inhibitor-1 (PAI-1) in cultured peritubular cells
recovered from 20-day-old rat testes. We demonstrated that TNF in a
nanomolar dose range stimulated PAI-1 messenger RNA (mRNA; Northern
blots) as well as immunoreactive (Western blots) and bioactive
(Stachrom) PAI-1 protein. Induction of PAI-1 mRNA started 4 h
after the addition of TNF (2.5-fold increase) and peaked (7-fold
increase) after 24 h of treatment. Actinomycin D and cycloheximide
inhibited the effects of TNF on PAI-1 mRNA, suggesting that ongoing
RNA and protein syntheses were required. The combined actions of
transforming growth factor- (TGF), a potent inducer of PAI-1, and
TNF on PAI-1 were less than additive, suggesting the activation of
some common pathway. TNF action on PAI-1, like that of TGF
demonstrated previously, was masked by a preexposure to phorbol
myristate acetate (a stimulator of protein kinase C) and strongly
reduced by staurosporine (an inhibitor of the protein kinase C).
Furthermore, using genistein to inhibit tyrosine kinase activity, we
not only blocked the action of TGF on PAI-1 [initiated upon binding
to the tyrosine kinase epidermal growth factor/TGF receptor
(EGFR)], but also markedly reduced that of TNF. Finally, TNF, at
a dose range that stimulated PAI-1, enhanced EGFR mRNA levels and EGF
binding. Together, the present findings suggest that some of the
biological effects of TNF on PAI-1 might be secondary to de
novo synthesis of EGFR. Because TNF probably originates from
testicular macrophages, such a regulation of PAI-1 by TNF may occur
in the context of physiological interactions between the testis and the
immune system. |
Introduction |
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The seminiferous tubule is a dynamic structure. It is characterized by a turnover of the extracellular matrix components (4), the migration of germ cells from the basal to the adluminal compartment with their subsequent entry into meiosis, and finally, the spermiation step (2). To allow these movements without perturbating the structural integrity of the seminiferous tubules and disturbing the blood-testis barrier, there must be a fine tuning of the different proteases and antiproteases present in the seminiferous tubules (see review in Ref.5). As the testis is under hormonal and local regulation (4, 6, 7, 8, 9), it is likely that the protease/antiprotease system is also under this interconnected control.
In this report, we focused our interest on plasminogen activator
inhibitor-1 (PAI-1). It is secreted by peritubular cells (10) and
inhibits the activity of the two plasminogen activators [namely the
urokinase (u-PA) and the tissue (t-PA) types] (11, 12), which are
produced by Sertoli cells (5, 13, 14). PAI-1 gene expression is induced
by growth factors and cytokines (15). In testis, previous studies have
reported that PAI-1 is up-regulated by transforming growth factor-ß1
(TGF-ß1) (16), basic fibroblast growth factor (bFGF) (17), and TGF
(17), factors for which peritubular cells exhibit the corresponding
high affinity receptors (18, 19, 20). However, and although cytokines have
been deeply implicated in testicular growth, development, and
differentiation (8, 9), no report has yet identified whether cytokines
could alter the plasminogen activation system.
As peritubular cells face the interstitium, and macrophages are a
potential source of tumor necrosis factor (TNF) within the
testis (21), we sought to determine whether regulatory interactions
could take place between the immune system and the peritubular cells
that will modify the antiprotease activity of the tubule.
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Materials and Methods |
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was purchased from Prepro Tech (Canton, MA). Human
recombinant TGF was obtained from Sigma Chemical Co. (St. Louis,
MO). Actinomycin D, cycloheximide, and staurosporine were of culture
grade and purchased from Sigma. Genistein was purchased from Upstate
Biotechnology (Lake Placid, NY).
Cell preparation
Peritubular and Sertoli cells were isolated from 20-day-old rats
and cultured at 32 C in a humidified atmosphere of 5% CO2
as previously described (22), with certain modifications (19). A
hyaluronidase treatment (0.1 mg/ml; 30 min; 34 C) was introduced after
the first collagenase-dispase digestion, followed by collection of
peritubular cells in the supernatant. After several washes in culture
medium, cells were filtered through a 20-µm nylon screen. Cells were
seeded in 60-cm2 petri dishes (Northern blots) or in
24-multiwell plates (binding studies) and cultured in 10% FCS until
confluent. Four days later, cells were cultured in the absence of serum
for 24–48 h. Alkaline phosphatase is a marker for peritubular myoid
cells (23) and has been used routinely to detect peritubular myoid
cells in the cell cultures. Alkaline phosphatase-positive cells were
less than 5% of the total cell population in Sertoli cell cultures and
greater than 85% in peritubular cell cultures.
Spermatogenic cells were isolated from 60- to 90-day-old rat testes by trypsinization. The resulting crude germ cell population (containing germ cells from all developmental steps) was submitted to a centrifugal elutriation using a rotor Beckman JE-6 (Fullerton, CA), as described previously (24, 25). Two fractions enriched at 80–85% with pachytene spermatocytes and early spermatids were harvested. After collection, the different cell populations were processed for RNA.
Treatment of peritubular cells in culture
On day 5 or 6 of culture, peritubular cells were treated for
various lengths of time with different reagents (growth factors or
drugs) as described in Results. Thereafter, culture media
were collected, and cells were either scraped from the dishes and
processed for RNA or used for binding studies. None of the treatments
performed affected cell number or viability in our experimental
conditions.
Isolation of RNA and Northern blot hybridization
Total RNA was isolated using acid-guanidinium
thiocyanate-phenol-chloroform extraction in a single step procedure as
reported previously (26). The polyadenylated [poly(A)+]
RNA was isolated using the Promega (Charbonnieres, France) PolyA tract
messenger RNA (mRNA) isolation system. The probes used for the
hybridization were a 1.2-kilobases (kb)
EcoRI-BglII fragment of the human PAI-1
complementary DNA (cDNA), a 2.2-kb EcoRI fragment
encompassing the entire extracellular domain of the rat epidermal
growth factor (EGF) receptor (ER1), a 0.8-kb EcoRI fragment
encoding only part of the intracellular domain but not the
extracellular domain (ER3), and a 1.3-kb rat glyceraldehyde-3-phosphate
dehydrogenase cDNA (GAPDH). These cDNA probes were provided by Dr. L.
R. Lund, Drs. S. H. Earp and J. M. Blanchard, respectively. Probes were
labeled using the Promega random primed DNA labeling kit (SA,
109 dpm/µg DNA). Northern blot analysis was performed as
previously described (19). A 0.24- to 9.5-kb Promega RNA ladder was
used to determine the size of the transcripts.
Western blots
SDS-PAGE and Western blotting were carried out as described
previously (27, 28). Primary antibody was a rabbit antirat PAI-1 IgG
(dilution, 1:200) from American Diagnostics (1062, Greenwich, CT).
Secondary antibody was a goat antirabbit IgG (dilution, 1:500)
conjugated to alkaline phosphatase and was obtained from Dako (Trappes,
France). Conditioned media from peritubular cells were concentrated 10
times using Centriprep (Mr cut-off, 10 kDa; Amicon,
Beverly, MA). Proteins were separated on a 10% SDS-PAGE
electrophoresis. A rat PAI-1 positive control (1060, American
Diagnostics) and prestained mol wt standards were loaded in each
SDS-PAGE gel. Under these conditions, PAI-1 migrated as a doublet band,
a major band of 49 and a minor band of 46 kDa, as reported previously
(28).
Colorimetric assay of PAI-1
Stachrom PAI-1 (Diagnostica Stago, Asnieres-Sur-Seine, France)
is a colorimetric assay based on the ability of PAI-1 to form a complex
with urokinase and inhibit the generation of plasmin (29). The reaction
mixture contains either PAI-1 calibrators or the test sample plus a
cocktail of urokinase, plasminogen, and inhibitors of
2-antiplasmin and 2-macroglobulin. The
amount of PAI-1 is inversely proportional to the amount of plasmin
generated. Results are expressed as amidolytic units (AU) per ml and
correspond to the amount of bioactive PAI-1 present in the concentrated
samples. The sensitivity of the assay ranged from 1–2 AU/ml. Inter-
and intravariations were 8.6% and 7.9%, respectively.
125I labeling of EGF and binding assays
Purified EGF (1 µg) was labeled using 125I
(Amersham, Les Ulis, France) by the chloramine-T procedure (30).
Preparations of [125I]EGF with a specific activity of
200–300 µCi/µg were obtained by this method. For measurements of
[125I]EGF binding, cells were seeded to a confluence of
500,000 cells in 24-multiwell plates. Cells were washed once with the
culture medium and then incubated with [125I]EGF (0.13
nM) at 4 C in the absence of unlabeled EGF (total binding)
or in the presence of an excess (300-fold) of unlabeled EGF
(nonspecific binding). The cells were then washed with PBS containing
BSA (1 mg/ml) at 4 C and solubilized in 0.5 N sodium
hydroxide-0.4% deoxycholate. The radioactivity was measured in a
-counter. For all experiments, specific binding is the difference
between total binding and nonspecific binding. Binding data were
evaluated by the method of Scatchard with the aid of EBDA and Ligand
computer programs (31), which were converted to BIOSOFT by G. A.
McPherson and distributed by Elsevier Biosoft (Cambridge, UK).
Data analysis
The band densities were determined by scanning densitometric
analysis using the Bioprofil scanner (Vilber Lourmat, Marnes-la-Vallee,
France). The amount of RNA in each lane of each blot was internally
standardized within a blot by assessing the amount of GAPDH mRNA per
lane. All experiments were repeated at least three times, with
independent cell preparations. A representative experiment of each
series is presented. The significance of the results was determined by
ANOVA, linear regression analysis, or Student’s t test when
comparing data from two groups. Differences are accepted as significant
at P < 0.05.
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Results |
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in doses ranging from 0.125–25 ng/ml
induced in a dose-dependent manner (P < 0.01) the 3-kb
signal, corresponding to the PAI-1 mRNA, after 8 h of treatment
(Fig. 1
). A maximal increase (3.7-fold) occurred with
12.5 ng/ml (0.7 nM) of TNF, and the ED50 was
1 ng/ml (0.06 nM).
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, the effect of TNF was time
dependent. Results are expressed as a percentage of the time-matched
controls (from 2–48 h). The first significant effects were noted after
4-h exposure to TNF (a 2.46-fold increase; P <
0.01). The stimulation reached a plateau at 24 h (a 6.72-fold
increase; P < 0.001) and thereafter was maintained for
additional 24 h.
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(12.5 ng/ml)
increased approximately 2.7-fold (P < 0.02; n =
4) the levels of immunoreactive PAI-1 after 24 h (Fig. 3). In addition, we found that TNF increased 2.7-fold
(P < 0.01) the secretion of bioactive PAI-1 measured
after 48 h of treatment. Indeed, control cells secreted 10.42
± 2.52 AU/ml (n = 3), and TNF-treated cells secreted
27.60 ± 2.22 AU/ml (n = 3).
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action on PAI-1 mRNA levels, peritubular cells were
treated with actinomycin D (5 µg/ml) or cycloheximide (50 µg/ml).
After 30 min and without changing the culture medium, cells were
treated with or without TNF (12.5 ng/ml). Incubations were stopped
8 h later. The results presented in Fig. 4 indicate
that actinomycin D inhibited significantly by 4-fold (P
< 0.001), and cycloheximide by 65% (P < 0.05) the
TNF-induced enhancement of PAI-1 mRNA levels. Of note and as
previously described (12, 17), cycloheximide by itself enhanced PAI-1
mRNA levels.
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and TGF is synthesized by peritubular cells (20). It is a potent
and fast acting inducer of PAI-1 in our culture model (17). We thus
examined the combined actions of TGF and TNF on PAI-1 mRNA
levels. Peritubular cells were treated for 8 h with TNF and/or
TGF at doses eliciting the maximal increase in PAI-1 mRNA levels.
The data presented in Fig. 5 show that PAI-1 mRNA levels
in TGF- and TNF-treated cells were not significantly different
(P = 0.2, using ANOVA) from PAI-1 mRNA levels measured
in TGF-treated cells, suggesting that the two factors may activate
some common pathway.
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-induced PAI-1 may
involve the protein kinase C (PKC) pathway (17), we analyzed the PKC
dependency of TNF regulation of PAI-1 mRNA levels. Addition of
either TNF or PMA (100 nM), a stimulator of PKC and
PAI-1 mRNA levels (17) (Fig. 6), did not further
significantly increase PAI-1 mRNA levels in cells pretreated for
24 h with 100 nM PMA (Fig. 6). We also employed
staurosporine to inhibit PKC. The dose of 100 nM was
selected because this dose did not affect PAI-1 mRNA levels in control
cells (Fig. 7), whereas it inhibited 100% of the
PMA-induced response (not shown). We observed that the stimulation
exerted by TNF was +339% (P < 0.01) in cells not
exposed to staurosporine and +102% (P < 0.04) in
cells exposed to staurosporine. Thus, exposure to staurosporine
resulted in a 70% loss of responsiveness to TNF (Fig. 7).
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acts upon binding to its high affinity tyrosine kinase
receptor, the EGF/TGF receptor (EGFR). To further investigate the
interference in the action of TNF and of TGF on PAI-1, we
analyzed the action of genistein, an inhibitor of tyrosine kinases
(32). The protocol included pretreatment with 10 µg/ml genistein,
followed by 8-h exposure to either TGF (10 ng/ml) or TNF (25
ng/ml). Doses higher than 10 µg/ml were found to affect the basal
steady state levels of PAI-1 mRNA levels (not shown). The results
presented in Fig. 8 indicate that genistein abrogated
the TGF-induced enhancement of PAI-1 mRNA levels. The stimulation
exerted by TNF was +125% (P < 0.05) in cells
exposed to genistein and +767% (P < 0.001) in cells
not exposed to genistein. Thus, exposure to genistein resulted in an
84% loss of responsiveness to TNF.
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up-regulated EGF binding and EGFR mRNA could regulate EGFR. As
previously described (20), we found that peritubular cells exhibited
high affinity receptors for EGF, with a Kd value
of 100 pM and 15,000 binding sites/cell (not shown).
Addition of TNF (20 ng/ml) to cultured peritubular cells resulted in
a time-dependent increase in EGF binding (Fig. 9). The
increase first became apparent by 6 h after the addition of TNF
20 ng/ml (+19%; P < 0.02). A plateau was observed
after 26 h (+55%; P < 0.001).
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using the ER3 cDNA probe, three major bands,
migrating at 9.6, 6.5, and 5 kb, and a diffuse and minor band of 1.3 kb
were observed in cultured peritubular cells (lane 4) and, to a lesser
extent, in 20-day-old rat cultured Sertoli cells (lane 3). A very weak
hybridization pattern was detected in early spermatids (lane 2), but
not in pachytene spermatocytes (lane 1). We next stripped this blot and
hybridized it with the ER1 cDNA probe, which extends 5' to the ER3 cDNA
probe and encompasses the extracellular domain. A hybridization pattern
similar to that yielded with the ER3 cDNA probe, was observed except
that a 2.7-kb band appeared with an intensity similar to the other
transcripts (Fig. 10b).
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(12.5 ng/ml) for 6 h resulted in a significant
stimulation of about 2.2-fold (P < 0.01; n = 3)
of each transcript (Fig. 11). In routine analysis, we
worked with total RNA rather than poly(A+). Under these
conditions, we could only quantify the 9.6-kb transcript; the other
hybridizing bands were too diffuse (Fig. 11). Maximal and half-maximal
increases in the 9.6-kb transcript were obtained with 25 ng/ml (1.4
nM) and 2 ng/ml (0.12 nM) TNF (Fig. 11). The
first significant effects (1.63-fold increase; P <
0.02) were noted after 2-h exposure to TNF (25 ng/ml). The plateau
of stimulation (a 7.6-fold increase; P < 0.001) was
reached after 24 h of treatment (Fig. 12).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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. We also presented evidence suggesting that part
of the TNF-induced enhancement of PAI-1 might be secondary to a
de novo synthesis of EGFR.
The central location of peritubular cells within the testis, between
the interstitial tissue and the inner of the seminiferous tubules,
suggests that these cells may not only establish a dialogue with
Sertoli cells and Leydig cells as reviewed previously (4), but also
with testicular macrophages through diffusible factors. Recently,
TNF has been shown to be produced by testicular macrophages (21). In
addition, increasing evidence accumulated over the past years suggests
that TNF may have a profound influence on Sertoli cell and Leydig
cell function and differentiation (see review in Refs. 8 and 9).
Therefore, in the present study we investigated whether TNF could
also regulate peritubular cells by assessing the action of this
cytokine on PAI-1, a marker for peritubular cells (10). Our interest in
studying PAI-1 stems from the fact that net protease activity within
the gonad is likely to insure the structural cohesion of the tubule and
to be involved in the formation of seminiferous cord structures during
fetal life, in the remarkable restructuring occurring at specific
stages of spermatogenesis, and in the turnover of the extracellular
matrix (see review in Ref.5).
We first showed that TNF enhanced both PAI-1 mRNA and protein
levels. The concentrations of TNF used in this study are well within
the Kd of the two high TNF affinity receptors
described previously (33). Our data are also consistent with previous
studies showing induced PAI-1 activity in cultured human endothelial,
carcinoma, or fibrosarcoma cells treated with TNF (see review in
Ref.12). We observed that the action of TNF on PAI-1 required
ongoing RNA synthesis. Additionally, posttranscriptional events such as
TNF-induced stabilization of PAI-1 mRNA might explain the difference
in the amplitude of the effects exerted by TNF on PAI-1 mRNA and on
PAI-1 protein (approximately a 7-fold vs. a 3-fold
increase). Interestingly, induction of PAI-1 mRNA levels by TNF
started 4 h after the addition of the cytokine, and mRNA levels
continued to increase with time. The stimulation reached a plateau at
24 h and thereafter was maintained for an additional 24 h.
These data contrast with the kinetics of enhancement of PAI-1 mRNA
induced with either bFGF or TGF (17). Indeed, the effects of bFGF
and TGF on PAI-1 mRNA peaked by 4 h and reached a nadir by
24 h. In addition, cycloheximide abrogated TNF action on PAI-1,
suggesting a role of protein mediators in the signal transduction
pathway. By contrast, the inhibitor of protein synthesis augmented in
an additive manner bFGF- and TGF-induced enhancement of PAI-1 (17).
Thus, our results suggested that upon stimulation by TNF, a cascade
of events was involved, leading to a sustained stimulation of PAI-1. A
similar delayed action of TNF has been previously demonstrated on
Sertoli cell inhibin mRNA levels (34).
In an attempt to elucidate the cascade of events by which TNF could
enhance PAI-1, we examined the combined actions of TGF and TNF.
Indeed, it is known that TNF can enhance TGF and EGFR in human
fibroblasts and pancreatic cancer cells (35, 36, 37). EGFR is the high
affinity tyrosine kinase receptor for TGF. It is up-regulated by
TGF (38). In addition, TGF is synthesized by peritubular cells
(20), and it stimulates PAI-1 (17). We found that TNF did not
significantly enhance PAI-1 mRNA levels in cells treated with a maximal
dose of TGF. This finding suggested the activation of some common
pathway in response to TGF and TNF.
It is known that PMA augments PAI-1 gene expression (17, 39), that
TGF-induced enhancement of PAI-1 may involve PKC activation (17),
and that TNF activates multiple intracellular signaling mechanisms,
including PKC (40). We thus examined the PKC dependency of TNF
regulation of PAI-1. We observed that down-regulation of PKC (provoked
by long term pretreatment with PMA) rendered cells refractory to
further TNF treatment, and that staurosporine significantly
decreased the effects of TNF. However, the actions of TNF could
simply be masked by the baseline increase in PAI-1 mRNA levels observed
in PMA-pretreated cells. In addition, staurosporine is not a specific
inhibitor of PKC. Therefore, more experiments will be required to
firmly establish that activation of PKC may be a common mechanism
through which TGF and TNF induce PAI-1.
We also challenged cells with genistein, an inhibitor of the tyrosine
kinases (32). As expected, TGF action on PAI-1 was totally inhibited
by genistein. In addition, we found that genistein markedly reduced
TNF-induced enhancement of PAI-1 mRNA levels. However, it cannot be
excluded that TNF-induced PAI-1 activates tyrosine kinases different
from the EGFR in our culture model. For example, TNF has been shown
in different human cell lines to activate the JNK protein kinase signal
transduction pathway, and JNK protein kinases are activated by dual
phosphorylation on tyrosine and threonine (41, 42). In addition, TNF
has been shown to induce the expression of bFGF (and bFGF induces
PAI-1) (17) and nerve growth factor (42). These factors act upon
binding to high affinity tyrosine kinase receptors, which have been
identified in peritubular cells (19, 43).
We finally investigated the action of TNF on EGFR mRNA levels and
EGF binding. We found that TNF at a dose range that stimulated PAI-1
enhanced EGFR mRNA levels and EGF binding, following a kinetics and
with an amplitude comparable to those reported earlier for human
fibroblasts (35, 36, 37). Together with our previous findings, this study
raised the possibility that some of the TNF effects on PAI-1 may
result from de novo synthesis of EGFR. However, the kinetics
of the TNF enhancement of PAI-1 mRNA levels (first significant
effects at 4 h) and of EGF binding (first significant effects at
6 h) suggest that other mechanisms are triggered by TNF to
enhance PAI-1. Previous studies have reported that TNF could
activate nuclear factor-
B and stimulate fos and
jun genes in the first hour after the addition of the
cytokine (44, 45, 46). As the products of the fos and
jun genes interact to enhance transcription of the
AP-1-responsive genes, it would be of interest to determine in our
culture model whether TNF action on PAI-1, which is an
AP-1-responsive gene (39), also involved an induction of the
fos and jun genes.
Our study also revealed that EGFR mRNA was present as four major transcripts of 9.6, 6.5, 5, and 2.7 kb in peritubular cells, to a lesser extent in Sertoli cells, and very faintly in early spermatids. These data extend the findings of a previous study using freshly isolated testicular cells and a S1 nuclease procedure (20). All transcripts except the 2.7-kb one are sufficient in size to code for the full-length EGFR whose coding region alone spans 3.6 kb (47). It would be of interest to determine whether the 2.7-kb transcript in testicular cells, like that in liver cells (47), is generated by alternative splicing of the primary receptor mRNA and encodes a truncated and secreted form of the EGFR.
Whether the in vitro data presented here reflect a potential
physiological (or physiopathological) role of TNF in the male gonad
function requires further studies. In this context, it should be
remembered that whereas TNF produced by germ cells (the other
testicular source for TNF identified to date) (48) cannot cross the
blood-testis barrier and thus cannot reach peritubular cells, TNF
may also originate apart from testicular macrophages, from the
circulatory system. This would be the case particularly in patients
with critical illness, burns, and sepsis. Interestingly, those patients
have depressed gonadal function. Consistently, administration of TNF
to rats induced a dramatic decline in testosterone levels and severe
epithelium damage. Furthermore, TNF has been shown in
vitro to reduce testosterone production by Leydig cells and to
antagonize the actions of FSH and insulin-like growth factor I in
Sertoli cells (9). As male infertility is associated in some cases with
abnormal thickening of the seminiferous tubule lamina propria or
boundary tissue (49, 50), it would be of interest to determine whether
an excess of TNF after an infection or an inflammation leads to an
increase in PAI-1, thickening the tubule basement membrane and reducing
fertility. Nonetheless, it should be noted that mice with single
deficiencies of lymphotoxin (51), 55-kDa (52) or 75-kDa (53) TNF
receptors, or PAI-1 (54) were apparently healthy and fertile. Thus,
this would suggest that other cytokines or growth factors (8, 9) and
other antiproteases (5) present in the testis exert redundant or
overlapping functions, bypassing the knocked out gene.
In summary, by use of a model of cultured peritubular cells, this study
has demonstrated that besides being regulated by cAMP analogs (16),
TGFß (16), bFGF (17), and TGF (17), PAI-1 gene expression is also
under the control of TNF. Given that PAI-1 inhibits Sertoli cell
plasminogen activator activity, TNF might be involved in the
dialogue between Sertoli cells and peritubular cells by participating
in the regulation of proteolytic activity within the seminiferous
tubules.
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Acknowledgments |
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Footnotes |
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Received July 22, 1996.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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stimulates AP-1 activity through
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spermatogenic cells. Endocrinology 133:389–396This article has been cited by other articles:
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 C. Banfi, L. Mussoni, P. Rise, M. G. Cattaneo, L. Vicentini, F. Battaini, C. Galli, and E. Tremoli Very Low Density Lipoprotein–Mediated Signal Transduction and Plasminogen Activator Inhibitor Type 1 in Cultured HepG2 Cells Circ. Res., July 23, 1999; 85(2): 208 - 217. [Abstract] [Full Text] [PDF]
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 F. Boussouar, R. Grataroli, J. Ji, and M. Benahmed Tumor Necrosis Factor-{alpha} Stimulates Lactate Dehydrogenase A Expression in Porcine Cultured Sertoli Cells: Mechanisms of Action Endocrinology, July 1, 1999; 140(7): 3054 - 3062. [Abstract] [Full Text]
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 B. Le Magueresse-Battistoni, G. Pernod, F. Sigillo, L. Kolodié, and M. Benahmed Plasminogen Activator Inhibitor-1 Is Expressed in Cultured Rat Sertoli Cells Biol Reprod, July 1, 1998; 59(3): 591 - 598. [Abstract] [Full Text]
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