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and IFN-
Regulate Expression and Function of the Fas System in the Seminiferous Epithelium
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Department of Histology and Medical Embryology, Istituto Pasteur-Fondazione Cenci Bolognetti, University of Rome "La Sapienza," Rome, Italy;
Institute of Experimental Medicine, Consiglio Nazionale delle Ricerche, Rome, Italy; and
Department of Experimental Medicine, University of L’Aquila, Aquila, Italy
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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and IFN-
markedly increase the
expression of both soluble and membrane-bound Fas in a dose-dependent
manner. The up-regulated membrane-bound Fas protein is functionally
active because it induces a significant level of Sertoli cell death in
the presence of Neuro-2a FasL+ effector cells.
Interestingly, the soluble form of Fas, which is induced by the same
cytokines but has an antiapoptotic effect, is also functional. In fact,
conditioned media from TNF--stimulated Sertoli cell cultures inhibit
Neuro-2a FasL+-induced cell death. Taken together, our data
suggest a possible regulatory role of TNF- and IFN- on
Fas-mediated apoptosis in the testis through disruption of the balance
between different forms of Fas. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The expression of Fas ligand (FasL)2 in Sertoli cells has been reported in recent studies (5). The interaction of Sertoli cells with Fas-bearing autoreactive lymphocytes, leading to the death of the latter by apoptosis, has been proposed as the mechanism responsible for the maintenance of immune tolerance in the testis (5).
The Fas/Apo-1/CD95 molecule is a cell-surface receptor that mediates apoptotic signals upon its FasL engagement (6). In recent years, the Fas/FasL system has been considered to be one of the central mechanisms in the homeostasis of immune response (7). Fas is expressed on a variety of different cell types (8), though the physiological role of its wide constitutive expression has not yet been clarified. In contrast, FasL is more restricted in its expression and is inducibly expressed on T, B, and NK cells during activation of the immune system (9, 10). Sequentially, Fas/FasL interaction down-regulates the immune response by inducing apoptosis, as activated lymphocytes express both Fas and FasL (11, 12). The fact that dysregulation of the Fas system leads to uncontrolled lymphoproliferation and severe autoimmune disorders (13, 14) means that the Fas system plays a role in the control of immune reactions in several organs in physiological and pathological conditions (9, 15, 16).
Screening of adult mouse tissues by Northern blotting analysis has shown that the testis represents the major nonlymphoid source of FasL in the body (17), whereas mRNA for its counterreceptor Fas has not been detected in the testis (8). More recently, RNase protection analysis has been used to demonstrate that the Fas gene is expressed at low levels in the mouse testis (18) and appears to be restricted to some germ cells (19). Although direct evidence by Northern blotting analysis has never been produced, it is generally assumed that FasL is constitutively expressed by Sertoli cells (5, 18). As regards the function of the Fas system in the testis, it has been postulated that this system plays a role both in maintaining the immune-privileged nature of the testis in mouse (5) and in the regulation of physiological testicular germ cell apoptosis in mouse, rat, and human (19, 20). Both roles are in keeping with the main functions performed by the Sertoli cell: the barrier function against interstitial immune attack and the regulatory function on spermatogenesis. However, regardless of whether its role in the testis is immunological or physiological, the Fas system activation requires strict regulation to avoid indiscriminate cell death.
It has been demonstrated that some cytokines are involved in the local
control of functional activities of the testis. TNF- is a testicular
paracrine factor, known to be produced by germ cells (21),
that induces transferrin release from Sertoli cells (22)
and is an important proinflammatory cytokine that induces IL-6
production and ICAM-1 and VCAM-1 expression in Sertoli cells
(4). The transduction mechanisms by which these biological
effects are obtained in Sertoli cells have been studied extensively
(23, 24). Several reports have indicated that cytokines,
particularly TNF- and IFN-, are also regulators of Fas and FasL
expression in other cellular systems (25, 26).
In the present study, we investigated the effects of TNF- and/or
IFN- on Fas/FasL expression on Sertoli cells and demonstrate that
both cytokines increase Fas, but not FasL, expression and that the
up-regulated membrane-bound Fas (mFas) is functionally active and
induces death of Sertoli cells when cocultured with Neuro-2a
FasL+ (N2a-FasL+) effector
cells.
Moreover, soluble isoforms of cell-surface receptors regulate receptor
function in a number of biological systems (27, 28). Fas
and FasL proteins can also occur both as cell-surface and soluble
proteins, with the soluble form of Fas (sFas) being generated by
alternative mRNA splicing (29, 30), whereas the soluble
form of FasL (sFasL) is produced through the proteolytic cleavage of
the membrane-bound receptor (31). Both sFas and sFasL can
protect against Fas-mediated apoptosis by neutralizing their respective
membrane receptors (32, 33). Thus, we also studied whether
TNF- and/or IFN- affect the production of sFas by Sertoli cell
cultures and found that these cytokines induce bioactive sFas. This is,
to our knowledge, the first evidence of the induction by cytokines of
both Fas transcripts, a finding that suggests that the ratio between
sFas and mFas receptors may be critical in the regulation of
Fas-mediated cell death in the testis.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Mock (neomycin-resistance cDNA)-transfected neuroblastoma Neuro-2a (N2a-Neo) and murine FasL cDNA-transfected Neuro-2a (N2a-FasL+) cell lines (kindly provided by Dr. A. Fontana, University Hospital of Zurich, Zurich, Switzerland), murine Fas cDNA-transfected murine B lymphoma (A20) (kindly provided by Dr. M. Hahne University of Lausanne, Lausanne, Switzerland), and murine plasmacytoma (P815 Fas) (kindly provided by Dr. P Vassalli, University of Geneva, Geneva, Switzerland) were maintained in DMEM (Life Technologies, Paisley, U.K.) containing 10% heat-inactivated FBS, 4.5 g/L glucose, 100 U/ml penicillin, and 100 µg/ml streptomycin (Life Technologies) at 37°C under 5% CO2.
The probes used were: a plasmid pMF1 that carries the full-length cDNA for mouse Fas cDNA, a 1.5-kb EcoRI fragment (8); a plasmid pBL-MFLW4 that carries the full-length cDNA for mouse FasL cDNA, a 0.94-kb XbaI fragment (34), both kindly given by Dr. S. Nagata (Osaka University, Osaka, Japan). Mouse c-kit probe, a 4.3-kb EcoRI fragment subcloned in pUC18, was kindly given by Dr. P. Rossi (Tor Vergata University of Rome, Rome, Italy).
DNase, collagenase, recombinant murine TNF-, and IFN- were
purchased from Boehringer Mannheim (Mannheim, Germany); trypsin was
obtained from Difco (Detroit, MI).
Sertoli cell cultures
Sertoli cells were prepared from CD1 mice as previously described (35). Briefly, testes from 15-day-old animals were sequentially digested for 20 min, first with HBSS containing 0.25% trypsin plus 10 µg/ml DNase and then with HBSS supplemented with 0.1% collagenase plus 10 µg/ml DNase to remove interstitial tissue and peritubular cells. Fragments of seminiferous epithelium, mainly composed of Sertoli cells, were cultured at 32°C in 95% air and 5% CO2 in serum-free MEM (Life Technologies). After 3 days, Sertoli cell monolayers were incubated at room temperature with 20 mM Tris-HCl buffer, pH 7.4, for 2.5 min to remove residual germ cells present in the culture (36). Sertoli cell cultures were routinely checked for possible contamination by macrophages and peritubular myoid cells by means of indirect immunofluorescence with anti-macrophage mAb (Mac-1 Ag CD11/b; Boehringer Mannheim) and by histochemical detection of alkaline phosphatase activity (37).
On the fourth day of culture, Sertoli cell monolayers were treated with
recombinant murine TNF- and/or IFN-. At the times indicated, the
cells were analyzed for Fas expression by flow cytometric analysis or
were lysed for Northern or Western blotting experiments.
Western blotting
Total Sertoli cell lysates were prepared by lysing and scraping the cells off the culture plate with 10 mM Tris-HCl, pH 6.8, 0.4 mM EDTA, 2% SDS, leupeptin, aprotinin, antipain (10 µg/ml each), and 1 mM PMSF (Sigma, St. Louis, MO).
The protein concentration of each sample was determined using the micro
bicinchoninic acid method (Pierce, Rockford, IL). Equal amounts of
proteins (50 µg) were subjected to SDS-PAGE and then transferred onto
a Hybond-C nitrocellulose membrane (Amersham, Braunschweig, Germany).
Subsequently, unspecific binding sites were blocked by treatment with
5% nonfat dry milk in TBS and 0.05% Tween-20 (TTBS) at room
temperature for 1 h. The blots were probed with polyclonal
anti-Fas Ab (0.125 µg/ml; M-20; Santa Cruz Biotechnology, Santa
Cruz, CA) or with monoclonal anti--tubulin (1:500; Sigma) in 5%
milk/TTBS for 1 h followed by a goat anti-rabbit or rabbit
anti-mouse HRP-conjugated secondary Ab (Zymed, San Francisco, CA).
After the first and second Abs, the membranes were washed three times
for 15 min with TTBS and detection was performed by the enhanced
chemiluminescence system (Amersham).
Flow cytometry
Control and treated Sertoli cells were detached with 0.02% EDTA and washed with cold PBS plus 1% BSA. For detection of Fas expression on the Sertoli cell surface, we used the FITC-conjugated hamster IgG anti-mouse CD95 mAb (Jo-2) (PharMingen, San Diego, CA). Specific mAb or the appropriate isotypic control mAb were used at 1 µg/106 cells for 30 min on ice. Cells were then washed twice with PBS plus 1% BSA and analyzed with a FACStar flow cytometer (Becton Dickinson Labware, Mountain View, CA). Cells were gated using forward vs side scatter to exclude dead cells and debris. Fluorescence of 104 cells/sample was acquired in logarithmic mode for visual inspection of the distributions and in linear mode for quantitating the expression of the relevant molecules by calculating the mean fluorescence intensity.
Cytotoxicity assay
The cytotoxicity was assayed by 51Cr release from labeled Sertoli cells. This assay was conducted in U-shaped, 96-well microtiter plates with a confluent monolayer of mouse Sertoli cells cultured as described above. After treatments with various cytokines for 24 h, Sertoli cells were washed twice with PBS and were labeled with 5 µCi 51Cr/well in 50 µl FCS (1 h at 37°C). After being washed three times with medium, these cells were used as the target and were mixed with the N2a-FasL+ effector cells (7 h at 37°C), at the ratios indicated, in a total volume of 100 µl. The plates were then centrifuged at 1000 rpm for 5 min, and 50-µl aliquots of the supernatants were assayed for radioactivity using a gamma counter. The percentage of specific lysis at various E:T ratios was calculated as follows: % cytotoxicity = [(experimental release cpm - spontaneous release cpm)/(total release cpm - spontaneous release cpm)] x 100, where experimental release cpm is the mean cpm released in the presence of effector cells (Neuro-2a cell line); spontaneous release cpm is the mean cpm released from targets (mouse Sertoli cells) cultured in medium alone or in the presence of different stimuli; and total release cpm is the mean cpm obtained by lysing target cells with 0.5% Triton X-100. The spontaneous release of 51Cr was routinely 10–15% of the total release.
A cytotoxicity inhibition assay was performed to determine bioactivity
of soluble Fas contained in medium conditioned by Sertoli cells (SCCM)
treated with TNF-. Sertoli cells were cultured in a 60-mm dish, and,
after 24 h of treatment with medium with or without 0.5 ng/ml
TNF-, SCCM were collected, filtered through 0.22-µm
low-protein-binding membranes (Millex-GV by Millipore, Molsheim,
France), and concentrated and desalted by Centriprep ultrafiltration
devices (Amicon, Danvers, MA). The SCCM were fractionated on
Centriprep-3; the retentate, concentrated 10-fold and containing
molecules with molecular masses over 3 kDa, was tested for
51Cr release inhibition in cytotoxicity assay.
The protocol of this assay was similar to previous cytotoxicity assay
with the following difference. The two different SCCM were tested in
the cococulture of 51Cr-labeled Sertoli cells
(target cells) with Neuro-2a FasL+ (effector
cells) at an E:T ratio of 10:1. The percentage of cytotoxicity was
subsequently calculated as 51Cr release as
described above.
Northern blotting
Total RNA was extracted from TNF-- and/or IFN--treated or
untreated murine Sertoli cells and from cell lines using TRIZOL Reagent
(Life Technologies). Samples were processed following the
manufacturer’s protocol. RNA was subjected to electrophoresis, through
a 1% agarose gel containing formaldehyde, and was transferred onto a
neutral nylon transfer membrane (Schleicher & Schuell, Keene, NH). The
filters were prehybridized in hybridization buffer containing 50%
formammide (Sigma), 10% dextran sulfate, 1% SDS, 1.2 M NaCl (Sigma),
and sonicated salmon sperm (Stratagene, La Jolla, CA) at 42°C for
4 h. Hybridization was performed overnight in prehybridization
buffer containing 1 x 106 cpm/ml of
32P-labeled murine Fas, FasL, or c-kit
cDNA probes. After washes, the blots were exposed to Kodak BIOMAX-MS
film (Eastman Kodak, Rochester, NY).
RT-PCR
First-strand complementary DNA was made in 20 µl reverse transcriptase buffer (Life Technologies) using 5 µg total RNA in the presence of Superscript II (200 U) and random primers (250 ng) (Life Technologies). The reaction quality was checked by PCR with specific primers for ß-actin mRNA amplification. Two microliters cDNA from the reverse transcription reaction were used as a template for the PCR. Each 50-µl PCR mixture also contained 30 pmol specific primers, nucleoside 5'-triphosphate (0.2 mM), MgCl2 (1.5 mM), 10x PCR buffer (5 µl), and 2.5 U Taq DNA polymerase (Life Technologies). PCR product of mouse Fas-ß (157 bp) was amplified using the primers 5'-GTCTCTCAGCTATCCAGTCTGA-3' and 5'-TGTATCCTGCCTGCAAGATGTGC-3'. Each sample was mixed, briefly centrifuged, overlaid with two drops of mineral oil, and placed in the DNA Thermal Cycler 480 (Perkin-Elmer, Norwalk, CT). Conditions for amplification of mouse Fasß were 94°C for 1 min, 53°C for 1 min, and 72°C for 1 min for 30 cycles. Mouse thymus was used as a positive control for Fasß detection (30).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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To study the presence of Fas in mouse Sertoli cells, we performed
Western blotting analysis by using polyclonal Abs specific for murine
Fas (M-20; Santa Cruz Biotechnology). We found that Fas is
constitutively expressed at a low level in mouse Sertoli cells lysates,
as shown by the detection of two bands at the expected molecular mass
(
45 kDa) (Fig. 1
a). The
smaller-sized band may represent the nonglycosylated primary translated
Fas protein (38). However, both bands disappeared when the
primary Ab was incubated with the blocking peptide (not shown).
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a, addition of
TNF- for 24 h to Sertoli cell cultures stimulated Fas
expression in a dose-dependent manner. The mean values of the
densitometric analysis of three Western blots demonstrated a strong
up-regulation, up to a 12-fold increase of the protein, at a maximal
dose of 100 ng/ml (Fig. 1b). Since it has been reported that
IFN- increases Fas expression in some cell types (8, 40, 41), we examined the effect of this cytokine alone and in
combination with TNF- on Sertoli cell Fas expression by Western
blotting analysis. As shown in Fig. 2,
IFN-, used at the maximal effective dose, induced a slightly lower
increase in Fas expression than that obtained with TNF-, but a
combined effect on the stimulation of Fas was obtained when Sertoli
cells were coincubated with both TNF- and IFN-.
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and
IFN- alone or in combination leads to significantly increased Fas
fluorescence on the cell surface (Fig. 3). These results are in agreement with
data obtained by Western blotting analysis.
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and IFN- increase Sertoli cell sensitivity to
FasL-mediated cytotoxicity
To ascertain the functional significance of Fas expression, we
tested whether FasL could trigger cytotoxicity in Sertoli cells.
Cultures were incubated with TNF- or IFN-, with both or with
neither, for 24 h and then labeled with
51Cr. FasL-transfected
N2a-FasL+ were used as effector cells at a
different E:T ratio with labeled Sertoli cells for an additional 7
h. The percentage of specific lysis of Sertoli cells was measured by
chromium release. As shown in Fig. 4,
lysis was observed in 5–8% of control Sertoli cells and was markedly
enhanced after treatment with TNF- or IFN-, occurring in 25–30%
of the cells. Moreover, the combination of both cytokines led to 70%
cell death, which points to a synergistic effect of the two
cytokines.
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). An annexin
V apoptosis detection assay was used to confirm that none of the
treatments induced apoptosis in Sertoli cells unless FasL-bearing cells
were added (data not shown).
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and/or IFN- increase levels of Fas mRNA, but not of FasL
mRNA, in mouse Sertoli cells
We examined the effects of these two cytokines on Fas mRNA
expression. Total RNA from Sertoli cell cultures stimulated with
TNF- (20 ng/ml) or IFN- (500 U/ml), with both cytokines or with
neither, for 24 h was examined by Northern blotting analysis using
a mouse Fas probe. Both cytokines induced a strong up-regulation of Fas
mRNA when added either separately or in combination (Fig. 6a). Since it has recently
been demonstrated that the proinflammatory cytokines IL-1ß, TNF-,
and IFN- up-regulate FasL expression on primary human keratinocytes
(26), we rehybridized the same membranes, used for the
detection of Fas mRNA, with mouse FasL probe, but we did not observe
the constitutive presence of FasL mRNA nor its induction by these
cytokines (Fig. 6b). Other authors have previously localized
Fas in germ cells, mostly spermatocytes (19). To exclude
the possibility that the Fas signal on untreated Sertoli cells was due
to germ cells contaminating Sertoli cell cultures, we next examined the
levels of c-kit, a germ cell-specific mRNA
(43). No signal was detected in Sertoli cell cultures by
Northern blotting analysis (Fig. 6c), indicating a high
grade of purity of the Sertoli cell cultures.
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and/or IFN- induce an alternative Fas transcript
encoding a soluble isoform of Fas (FasB)
A soluble, inhibitory isoform of Fas (FasB) generated by
alternative splicing of Fas mRNA (Fasß) has been described
(30). We analyzed the expression of Fasß in unstimulated
and cytokine-stimulated Sertoli cells by semiquantitative RT-PCR. Fig. 7 shows a significant induction of the
alternative transcript Fasß by both cytokines. To determine a
possible different pattern of induction for the two transcripts of Fas
by TNF-, we examined dose-dependent up-regulation of both membrane
forms of Fas mRNA and Fasß by Northern blotting analysis (Fig. 8a) and by RT-PCR (Fig. 8b), respectively. The plateau of Fasß enhancement was
evident when the concentration of TNF- was 10-fold lower than that
of the membrane Fas (0.5 ng/ml vs 5 ng/ml).
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Conditioned media from Sertoli cells (SCCM) either untreated or
treated with TNF- 0.5 ng/ml for 24 h were tested for bioactive
sFas by using cytotoxicity inhibition assay as described in
Materials and Methods. Briefly, to analyze inhibiting
activity of SCCM on FasL-mediated cytotoxicity, we added the two
different SCCM to the coculture of 51Cr-labeled
Sertoli cells (target cells) with N2a-FasL+
(effector cells) in the cytotoxicity assay. As shown in Fig. 9, only medium conditioned by
TNF--treated Sertoli cells significantly interfered with
FasL-mediated cytotoxicity. A nearly complete inhibition of
cytotoxicity was observed in control Sertoli cells expressing low
amounts of mFas, probably because the sFas present in TNF--treated
SCCM was sufficient to compete totally with mFas expressed by control
Sertoli cells. In contrast, when the membrane Fas on Sertoli cells was
up-regulated by TNF-, the inhibition of cytotoxicity obtained with
sFas contained in TNF--treated SCCM was 
40%.
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, we have incubated fresh medium plus 0.5 ng/ml
TNF-, without cells, for 24 h at 32°C, then we concentrated
it 10-fold, as for SCCM, and tested for 51Cr
release inhibition in cytotoxicity assay. This medium did not produce
any effect on FasL-mediated Sertoli cell death (data not shown). |
Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and/or IFN- increase their
expression of Fas Ag both in membrane-bound and soluble form. Moreover,
following cytokine stimulation, Sertoli cells become susceptible to
FasL-mediated cytotoxicity, which suggests a role of inflammatory
cytokines on Fas system regulation in the seminiferous epithelium. Our results suggest that inflammatory cytokines create a proapoptotic environment by inducing Fas up-regulation on the Sertoli cell surface, thus becoming effectively functional when massive contact occurs with FasL-bearing cells, such as activated T lymphocytes. Previously, we have demonstrated that inflammatory mediators enhance adhesion between Sertoli cells and lymphocytes by up-regulation of adhesion molecules specific for lymphocytes (4); this increased binding may favor Fas-mediated apoptosis of Sertoli cells, and a localized loss of Sertoli cells may cause a leakage of the blood-tubular barrier, hence leading to autoimmune orchitis. Previous reports have demonstrated the crucial regulatory role of the Fas system in the induction of autoimmune diseases such as Hashimoto’s thyroiditis (44), rheumatic disease (45), and multiple sclerosis (46). In Hashimoto’s thyroiditis, the IL-1ß-mediated induction of Fas on thyrocytes results in the immune response by lymphocytes and tissue homeostasis destruction by the thyrocytes themselves, which constitutively express FasL.
Recently, new insight into the role of the Fas system in autoimmune
diseases has been provided by the demonstration of a new role for FasL
in inflammation (47). FasL was discovered to induce in
peritoneal exudate cells the processing and release of IL-1 and
IL-1ß that are responsible for neutrophil infiltration. FasL induces
apoptosis in vanguard neutrophils, which in turn simultaneously release
active IL-1, leading to massive neutrophil infiltration. Classically,
apoptosis is defined as a type of cell death that does not provoke
inflammation. This is probably true in most programmed cell death in
which dying cells do not express IL-1. Because Sertoli cells in the
testis produce IL-1 (48), it may be possible that their
FasL-mediated apoptosis induces the release of IL-1, thus amplifying
the inflammation and enhancing the pathological effects in autoimmune
disease.
Since the first cloning of FasL cDNA, the testis came out as the organ expressing the highest level of FasL RNA. In the last few years, a number of studies have reported the localization of this molecule in Sertoli and/or in germ cells (18, 49). The role of this molecule in the physiology of normal testis is also controversial (5, 19). Furthermore, the expression of FasL during testis development and its localization during the cycle of the seminiferous epithelium have not been studied. Similarly the presence and distribution of FasL in experimental or spontaneous autoimmune disorders of the testis have not been yet investigated. Although it has been reported that Sertoli cells constitutively express FasL (5, 18), we observed neither FasL mRNA in cultured Sertoli cells, nor apoptosis in Sertoli cell cultures expressing Fas, regardless of cytokine treatment. Apoptosis took place only when FasL-transfected effector cells were added to the cultures. Thus, our data could be explained either by postulating that the presence of FasL on Sertoli cell is too low to be detected by Northern blotting analysis and to induce fratricide or that FasL expression in Sertoli cells is age dependent and is absent in prepuberal mice, from which cell cultures were derived. By contrast, in this study we demonstrate that the low basal level of Fas expressed on Sertoli cell surface is functional, leading to 2–8% of apoptosis in Sertoli cells cocultured with N2a-FasL+.
Moreover, it is noteworthy that the same cytokines that up-regulate the
apoptotic membrane-form of Fas in Sertoli cells also induce the
expression of an alternative Fas transcript encoding a soluble
antiapoptotic isoform of Fas, suggesting a more complex role of
cytokines in the regulation of Fas system-dependent apoptosis in the
testis. Although the two detection methods used to analyze mFas and
sFas RNAs expression cannot be directly compared, it is remarkable that
the plateau of sFas mRNA expression is reached at a low dose of TNF-
(0.5 ng/ml) (Fig. 8). The SCCM collected from Sertoli cells treated
with 0.5 ng/ml TNF- totally inhibits the basal apoptosis of control
Sertoli cells cocultured with N2a-FasL+, but only
partially inhibits the increased apoptosis in TNF--treated Sertoli
cells. It is likely that the amount of functional sFas may be too low
to completely block Fas-mediated apoptosis of TNF--treated Sertoli
cells, at least under our in vitro conditions. Furthermore, this result
may also suggest that in vivo, at the physiologically low
concentrations of TNF- produced by germ cells, the soluble
antiapoptotic form of Fas is secreted in the seminiferous tubule as a
survival factor. TNF- and IFN- are cytokines present in the
seminiferous tubule (21, 50) and are strongly up-regulated
in inflammatory disease; they may control apoptosis in the testis in
both physiological and pathological conditions by modulating
Fas-mediated cell death. Finally, because FasL is abundantly expressed
in the testis (17), whereas Fas protein expression is low
(18), the modulation of Fas levels may be important in the
control of Fas-mediated apoptosis in this organ.
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Footnotes |
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2 Abbreviations used in this paper: FasL, Fas ligand; sFas, soluble Fas; mFas, membrane Fas; N2a-FasL+, murine FasL cDNA-transfected Neuro-2a tumor; N2a-Neo, neomycin-resistance cDNA-transfected Neuro-2a tumor; SCCM, Sertoli cell-conditioned medium.
Received for publication January 27, 2000. Accepted for publication April 26, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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in mouse spermatogenic cells. Endocrinology 133:389. on transferrin release from Sertoli cells in culture. Endocrinology 129:256. induces interleukin-6 production and integrin ligand expression by distinct transduction pathways. J. Biol. Chem. 273:7566. leads to intercellular adhesion molecule-1 expression. J. Biol. Chem. 274:28978. and IFN- render microglia sensitive to Fas ligand-induced apoptosis by induction of Fas expression and down-regulation of Bcl-2 and Bcl-xL. Eur. J. Immunol. 28:4398.[Medline]
synthesis in rat testis. Mol. Cell Endocrinol. 82:R13.[Medline]
and - expression in the rat testis. Endocrinology 136:4925.[Abstract]
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V. Pentikainen, K. Erkkila, L. Suomalainen, M. Otala, M. O. Pentikainen, M. Parvinen, and L. Dunkel TNF{alpha} Down-Regulates the Fas Ligand and Inhibits Germ Cell Apoptosis in the Human Testis J. Clin. Endocrinol. Metab., September 1, 2001; 86(9): 4480 - 4488. [Abstract] [Full Text] [PDF]
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 A. D'Alessio, A. Riccioli, P. Lauretti, F. Padula, B. Muciaccia, P. De Cesaris, A. Filippini, S. Nagata, and E. Ziparo Testicular FasL is expressed by sperm cells PNAS, February 22, 2001; (2001) 51566098. [Abstract] [Full Text]
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A. D'Alessio, A. Riccioli, P. Lauretti, F. Padula, B. Muciaccia, P. De Cesaris, A. Filippini, S. Nagata, and E. Ziparo Testicular FasL is expressed by sperm cells PNAS, March 13, 2001; 98(6): 3316 - 3321. [Abstract] [Full Text] [PDF]
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