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Department of Physiology, Institute of Biomedicine, University of Turku, Kiinamyllynkatu 10, FIN-20520 Turku, Finland
Address all correspondence and requests for reprints to: Ilpo T. Huhtaniemi, M.D., Ph.D., Department of Physiology, University of Turku, FIN-20520 Turku, Finland. E-mail: ilpo.huhtaniemi{at}utu.fi
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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30 µg/liter), and a slow
stimulation, between 48–72 h, in response to lower PRL doses (1–10
µg/liter). In addition, extracellular Ca2+ (1.5
mmol/liter) increased the effect of PRL on human CG (hCG)-stimulated
StAR messenger RNA expression and progesterone (P) production.
Importantly, the biphasic effects of PRL on LHR gene expression and
hCG-mediated P production were abolished in the presence of anti-PRL
antiserum, demonstrating specificity of PRL action. The PRL effects on
StAR expression, and steroid and cAMP production, apparently reflect
its effects on LHR function. The relevance of the PRL effects observed
in mLTC-1 cells was supported by demonstration of similar PRL responses
in hCG-stimulated testosterone production of isolated mouse Leydig
cells. Collectively, these findings clearly demonstrate the biphasic
regulatory actions of PRL, and clarify some facets of the controversial
role of this hormone in Leydig cell function. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The steroidogenic acute regulatory (StAR) protein, a novel 30-kDa mitochondrial factor, has recently been purified, cloned, and characterized in MA-10 cells (17). The rate-limiting and regulated step in steroid hormone biosynthesis is the transport of cholesterol from the outer to the mitochondrial inner membrane (18, 19), which has recently been found to be mediated by StAR protein. It has also been demonstrated that StAR protein is intimately associated with the acute regulation of steroidogenesis in steroidogenic cells (20, 21, 22, 23). Recently, we demonstrated that human CG (hCG) markedly enhanced the StAR protein and StAR messenger RNA (mRNA) levels, which were consistent with progesterone (P) production in mLTC-1 cells (22). The possible connections of PRL and StAR protein actions are not known in the regulation of steroidogenesis.
The PRL action is mediated through its binding to a plasma membrane receptor, a member of the cytokine/PRL/GH receptor family, including those of GH, and a large number of lymphokines and related growth factors (24, 25, 26). This receptor family is characterized by a single transmembrane domain and conserved homology in the extracellular domain (25, 26). In the testis, PRL receptors are expressed in Leydig (27, 28, 29), Sertoli, and spermatogenic cells (30). Several receptors of this family, including those of PRL, were found to induce tyrosine phosphorylation and activation of receptor-associated tyrosine kinases of the Janus kinase (JAK) family (31, 32). In addition, PRL is known to activate the cytoplasmic signal transducers and activators of transcription (STATs), possibly through direct phosphorylation by the JAK2 tyrosine kinase (33). An important pathway of PRL actions is also its ability to regulate intracellular free calcium levels ([Ca2+]i) via increased Ca2+ entry (34, 35), as well as mobilization from intracellular stores (36). The findings on elevated [Ca2+]i may lie in the fact that it is involved in activation of the protein kinase C signaling pathway (37). However, crucial information is still lacking regarding the involvement of Ca2+ in PRL effects on testicular function.
The aim of the present study was to investigate the mechanisms of PRL action in a murine Leydig tumor cell line mLTC-1 (38), as monitored by responses of LHR, PRLR, StAR protein, Ca2+, steroidogenesis and various signal transduction pathways. These findings provide novel insights into the complexity of effects of PRL on testicular function.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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RNA extraction, and RT-PCR analysis of StAR and PRLR mRNA
expression
Total RNA was isolated from different treatment groups using the
single step method as described elsewhere (42).
The amplification and assessment of the StAR gene expression in mLTC-1 cells were investigated by using a quantitative RT-PCR assay method as described previously (23). Briefly, the primers used were: the StAR sense, 5'-GACCTTGAAAGGCTCAGGAAGAAC-3' and the antisense, 5'-TAGCTGAAGATGGACAGACTTGC-3' spanning bases -51 to -27 and 931 to 908, respectively, from the mouse StAR complementary DNA (cDNA) sequence (17). The variation in RT-PCR efficiency was evaluated by L19 ribosomal protein gene coamplified in each sample, using the sense primer 5'-GAAATCGCCAATGCCAACTC-3' and the antisense primer 5'-TCTTAGACCTGCGAGCCTCA-3'. RT and PCR of the target genes were run sequentially in the same assay tube using 2 µg of total RNA from different groups, as previously described (23). A 25-µl aliquot of each reaction was analyzed by electrophoresis on a 1.2% agarose gel. Gels were then vacuum dried and exposed to Kodak x-ray films (XAR-5, Eastman Kodak Co., Rochester, NY) at 4 C for 1 to 3 h. The relative levels of different signals were quantified by phosphorimagery and densitometry (Tina 2.0 Package, Straubenhardt, Germany).
The primers used in RT-PCR assay for PRLR mRNA were designed from the published rat PRLR cDNA sequence (43). The sense primer, 5'-GACAAGGAAACATTCACCTGCTGGTG-3', and the antisense primer, 5'-GGAACTGGTGGAAAGATGCAGGTCATC-3,’ spanned bases 128 to 103 and 826 to 800 bp, respectively. Briefly, 2 µg of total RNA from mLTC-1 cells were analyzed in RT-PCR assay as described previously (29). The RT-PCR product was analyzed in 1.2% agarose gels, which displayed an approximately 700-bp fragment of the expected size of PRLR mRNA. This fragment was subcloned into the T-vector (Promega Corp.), and used to determine the PRLR mRNA expression by Northern hybridization analysis (see below).
Northern hybridization analysis of StAR, LHR, PRLR, and JAK2 mRNA
expression in relation to PRL
The mLTC-1 cells were cultured in the absence or presence of
increasing concentrations of PRL (0–300 µg/liter) for 12, 24 or
48–60 h. For Northern hybridization, 15–20 µg of the total RNA were
resolved on 1.2% denaturing agarose gel and transferred onto nylon
membrane (Hybond, Amersham Pharmacia Biotech, Aylesbury,
UK). Prehybridization and hybridization were carried out as previously
described (44, 22). Briefly, an antisense cRNA probe, a
NotI fragment (980 bp) of the mouse StAR cDNA, a
BglII fragment (410 bp) of the extracellular domain of the
rat LHR, a BamHI fragment (700 bp) of the rat PRLR cDNA, and
a BglII fragment (670 bp) of the mouse JAK2 cDNA, were
produced by in vitro transcription (Promega Corp.), dNTPs and [
-32P]-UTP
(Amersham Pharmacia Biotech). The membranes were exposed
to x-ray films (Kodak XAR-5) for 1–3 days at -70 C or to
phosphorimage plates (Fujifilm BAS-5000, Fujifilm I&I, Japan) for
12 h, and quantified as above.
Isolation and purification of mouse Leydig cells
Preparation of mouse testicular Leydig cells was carried out
according to the procedure adopted earlier (23). In brief,
interstitial cells were dispersed by collagenase treatment (0.2%, 20
min, 34 C) in DMEM/F12 (1, 1) (Life Technologies, Inc.) containing 25 mmol/liter HEPES, pH 7.4, 18 mmol/liter
sodium bicarbonate and 0.2% BSA (Sigma, St. Louis, MO).
After removing collagenase by washing, the cells were purified by a
continuous Percoll (Amersham Pharmacia Biotech)
gradient (density range 1.01–1.126 kg/liter) centrifugation. The cell
types gathered at the zone to 1.07 kg/liter of Percoll were collected
and washed, and the Leydig cell purity was approximately 80%, as
determined by 3ß-hydroxysteroid dehydrogenase staining (23, 45). The cells were further subcultured in growth medium
supplemented with 10000 U/liter penicillin and 50 mg/liter
streptomycin, and evaluated for their functional activity as specified
below.
Effects of PRL on hCG-induced cAMP, progesterone (P) and
testosterone (T) production
The mLTC-1 cells (8 x 105
cells/well) were cultured for 24, 48, and 72 h with increasing
doses of PRL (0–300 µg/liter) in the presence of 0.1 mmol/liter
1-methyl-3-isobutylxanthine (MIX, Aldrich, Steinheim, Germany). The
cells were washed and restimulated for 3 h without or with 50
µg/liter hCG. Specificity of PRL actions was assessed in the absence
or presence of varying dilutions (1:1000 to 1:10000) of mouse anti-PRL
antiserum (NIDDK, AFP-1310178), and followed by hCG stimulation. This
antiserum has been demonstrated to be highly potent (1:400,000) in
mouse PRL RIA. The physiological relevance of PRL in testicular
function was assessed further through its effects on isolated mouse
Leydig cell steroid production. The media collected from subsequent
experiments were determined for cAMP, P and T levels by specific RIAs
(46, 47, 48). In certain experiments, the cells were lysed
using 0.3 N NaOH containing 1% SDS and subjected to
determination of total protein.
Measurement of DNA synthesis in mLTC-1 cells
To determine the mitogenic effect of PRL, mLTC-1 cells were
plated at a density of 6 x 104 cells/well
in 24-well plates. Following 12–24 h culture in the absence or
presence of PRL (1–100 µg/liter),
[3H]-thymidine (2 Mbq/well, Amersham Pharmacia Biotech) was added to the cells, and the culture was
continued for an additional 12 h. The cells were washed with PBS,
treated with 10% trichloroacetic acid on ice, lysed using 0.3
N NaOH containing 1% SDS and counted in a ß-spectrometer
(Rack-ß, Wallac OY, Turku, Finland).
45Ca2+ uptake, and involvement of
Ca2+ in hCG-mediated PRL response
To study the PRL-mediated Ca2+ uptake,
mLTC-1 cells were plated on 24-well plates at a density of 6 x
104 cells/well. Twenty-four hours later, the
cells were washed, cultured in calcium-free medium (S-MEM, Life Technologies, Inc.), and incubated with 5 µCi/m of
45Ca2+ (Ca-45, NEN Life Science Products, Boston, MA) in the absence or presence of
PRL (3–300 µg/liter). The cells were incubated for 15 min at 37 C,
followed by washing with ice-cold PBS, lysed in 500 µl 1% SDS
containing 0.3 N NaOH and the lysates were counted in a
ß-counter as above.
To further understand the involvement of Ca2+ in
PRL function, 5 x 105 cells/well in 6-well
plates were plated on the day preceding an experiment. The role of
Ca2+ (1.5 mmol/liter) in PRL action was assessed
on hCG-mediated StAR mRNA expression and P production. The cells were
stimulated for 48 h without or with 10 µg/liter of PRL, followed
by 3 h treatment with hCG in the presence or absence of
extracellular Ca2+. The additives, EGTA and
verapamil 123
-isopropyl-[(N-methyl-N-homoveratryl)-
-aminopropyl]
3,4-dimethoxyphenylacetonitril hydrochloride 125 were constituted
fresh, and applied to the cells during 3-h incubation periods, and
subjected to the analysis of StAR mRNA and P levels as above. The
concentrations of Ca2+, EGTA and verapamil used
were based on our previous finding (22).
Plasmids and transfections
The murine LHR promoter constructs were prepared as previously
described (49). Briefly, the LHR promoter constructs
contained the first 950- or 2040-bp of the 5'-flanking region of the
murine LHR gene (50) in front of the firefly luciferase
coding sequence. A cytomegalovirus (CMV) promoter/ß-galactosidase
plasmid (Promega Corp.) was used to control the
transfection efficiency.
Transfection studies were carried out by electroporation as previously described (49). In brief, 8 x 106 mLTC-1 cells were electro-shocked in a gene pulser cuvette (0.4-cm gap, Bio-Rad Laboratories, Inc. Richmond, CA). The cells were cotransfected with 40 µg of one of the LHR promoter-luciferase constructs and 2 µg of the control plasmid (CMV-ß-galactosidase) and plated on 6-well culture plates. After changing the media, the culture was continued for another 24 h in the presence and absence of 100 µg/liter PRL. The cells were collected thereafter, lysed by three cycles of freezing and thawing, and the cell lysates were assayed for luciferase activity as described previously (49).
Data analysis
The data were analyzed by one-way ANOVA, followed by Duncan’s
new multiple range test. The results are presented as mean ±
SEM, and a P value less than 0.05 was considered
statistically significant.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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demonstrate a clear time- and dose-dependent biphasic response pattern
of [125I]iodo-hCG binding to stimulation with
10–100 µg/liter of PRL. The only effect seen at 24 h was a
decrease of [125I]iodo-hCG binding at the
highest PRL level. A significant increase in binding (up to 45%)
occurred at 48 h with 10 and 30 µg/liter of PRL. At 72 h,
the increase at 10 µg/liter PRL reached 75%, whereas smaller effects
were found with the higher doses. The mLTC-1 cells treated with varying
doses of PRL (0–300 µg/liter) for 24–48 h did not display
significant changes in total protein contents or in thymidine
incorporation with different treatment groups in comparison to controls
(data not shown).
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). In
accordance with the effects of PRL on hCG binding, a 24-h pretreatment
with this hormone only showed, at doses 10 µg/liter and greater,
inhibition of hCG-stimulated cAMP production, when measured after a
subsequent 3 h stimulation with 50 µg/liter of hCG. Stimulation
of cells for 48–72 h with the higher PRL concentrations (30–300
µg/liter), still showed inhibition of hCG-stimulated cAMP production
which, however, declined as the time of culture increased. The lower
PRL doses (3–10 µg/liter) displayed stimulation of cAMP production
at these later time-points. The results of hCG-stimulated cAMP
production were in line with those of hCG binding, although the
negative effect of PRL on hCG signal transduction appeared at lower
concentrations than the respective effect on hCG binding. The biphasic
effect of PRL on cAMP production following 48 h pretreatment was
confined to the hCG-stimulated pathway, because no similar changes were
observed in basal cAMP levels, or in forskolin (FK)- or cholera toxin
(CT)- stimulated cAMP production (data not shown).
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). Interestingly, cells
stimulated for 60 h with increasing doses of PRL (0–300
µg/liter) demonstrated a biphasic response pattern, i.e.
increase up to 50% at lower concentrations, and declined (or lack of
effect) thereafter with higher PRL concentrations (Fig. 3B). The
results summarized in Fig. 3C demonstrate that PRL stimulation (12–24
h) decreased LHR mRNA expression in a dose-dependent manner, whereas
cells treated for 60 h demonstrated a biphasic dose-response to
PRL.
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Specificity of PRL actions on LHR gene regulation, and on
hCG-mediated P production in mLTC-1 cells
The biphasic effects of PRL on LHR mRNA expression were further
assessed in the presence of anti-PRL antiserum. The data presented in
Fig. 4A show that cells stimulated for
48 h with low dose of PRL (5 µg/liter) demonstrated a clear
increase in LHR mRNA expression, whereas the following increase showed
significant reduction with 50 µg/liter of PRL. Interestingly, both of
these responses of LHR mRNA were lost by incubating the cells in the
presence of mouse anti-PRL antiserum (1:6000), indicating specific
effect of PRL on the LHR gene expression. The anti-PRL antiserum was
tested at dilutions from 1:1000 to 1:10000, and 1:6000 was chosen for
the experiments (data not shown).
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), in line with the
biphasic PRL effect. Inclusion of anti-PRL antiserum into the media for
48 h completely abolished the PRL-dependent increase in P
production, compared with hCG stimulation. These data strengthen the
specificity of the PRL effects in the regulatory functions
observed.
Effects of PRL on hCG-stimulated StAR mRNA expression and P
production
Because 48 h PRL treatment demonstrated a biphasic response
on hCG-mediated cAMP accumulation; we examined next the StAR mRNA and P
levels in a similar experimental paradigm. As illustrated in Fig. 5, a dose-dependent enhancement of
hCG-stimulated StAR mRNA expression was observed up to 10 µg/liter of
PRL (194 ± 12% of control), whereas this response was remarkably
diminished at higher PRL concentrations (above 10 µg/liter).
Stimulation with PRL (10 µg/liter) alone did not show significant
increase on StAR mRNA. Practically identical responses were observed
when P production, instead of StAR mRNA expression, was measured
following the second incubation of 3 h in the presence of 50
µg/liter hCG (Fig. 5C), in accordance with the close correlation
between StAR expression and steroid production.
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show that the full-length mouse StAR hybridized with three transcripts
at 3.4, 2.8, and 1.6 kb with coordinate regulation, where the 1.6 kb
band corresponds to functional StAR protein. The data demonstrate that
PRL (10 µg/liter) increased the hCG-induced StAR mRNA expression 2-
to 4-fold following 3 h hCG (50 µg/liter) restimulation,
compared with hCG stimulation alone. On the other hand, the stimulated
levels of StAR mRNA were attenuated with higher doses of PRL (100–300
µg/liter), which further confirmed the previous results.
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shows that
stimulation with 10 µg/liter of PRL significantly increased the
subsequent hCG-stimulated T production (47 ± 3.2%), whereas at
the higher 100 µg/liter PRL dose it decreased T production by
approximately 36%, in comparison to preincubation in the absence of
PRL. The similarity of these findings with those observed with mLTC-1
cells indicates that the latter are a relevant model for studying
effects of PRL on murine testicular function.
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). These
data suggest that PRL increased the intracellular
Ca2+ through its mobilization from the
extracellular source.
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show that extracellular
Ca2+ (1.5 mmol/liter; 3 h) modestly but
consistently augmented (P < 0.01) the effects of PRL
pretreatment (10 µg/liter; 48 h) on hCG-stimulated (3 h) StAR
mRNA levels (Fig. 9A). Effect of Ca2+ on
hCG-stimulated StAR expression further increased significantly
following PRL pretreatment. The stimulatory effect of
Ca2+ on hCG- or PRL-augmented hCG response was
markedly attenuated by the Ca2+-chelating agent
EGTA (4 mmol/liter) and the calcium channel blocker verapamil (10
µmol/liter), suggesting the involvement of extracellular
Ca2+ in the stimulatory responses observed.
Ca2+ had no effects on basal and PRL pretreated
StAR mRNA levels, whereas it only potentiated the hCG-dependent
responses in the absence and presence of PRL. Subsequently, P
production in the media demonstrated similar responses as
StAR expression (data not shown), suggesting further an
intimate correlation between StAR expression and
steroidogenesis.
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).
Maximum elevation (282 ± 11%) of both PRLR transcripts was
observed at 10 µg/liter of PRL, followed by attenuation of these
levels at higher PRL concentrations.
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, Northern analysis
demonstrated qualitatively similar biphasic response to PRL (increase
up to 300% followed by decrease down to 76%) in JAK2 mRNA levels (4.8
kb) as was found in PRLR mRNA, again suggesting an association between
these two parameters.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The function of PRL is mediated through activation of the PRL receptor-associated JAK2 protein tyrosine kinase (51). The activated tyrosine kinases phosphorylate existing cytoplasmic factors known as the signal transducers and activators of transcription, STAT (52), and may indirectly bring about activation of the mitogen-activated protein kinase (53). Induction of genes by PRL has been demonstrated through activation of c-fos, c-myc, ornithine decarboxylase, and the interferon regulatory factor-1, IRF-1 (54). Studies on PRL have also demonstrated that it exerts both mitogenic and differentiating actions on its target cells. In the present study, the effects of PRL appeared to be dose- and time-dependent regarding the parameters studied in mLTC-1 cells. In addition, a dose-dependent, biphasic, short-term effect of PRL on Ca2+ entry was also demonstrated. The inhibitory action of PRL at higher concentrations on LHR mRNA expression may be due to the descending limb of a biphasic PRL effects on hCG binding and possibly other signaling pathways.
Nonspecific effects of PRL on LHR function can be ruled out as the PRL preparation used was negligibly (0.003%) contaminated by LH. Nevertheless, the specificity of the PRL effects was further confirmed by using anti-PRL antiserum, which completely inhibited the PRL responses. The biphasic actions of PRL on cAMP and P productions, as well as StAR expression were confined to the hCG-signaling pathway, because similar responses in these parameters were not observed in basal conditions or following CT/FK treatments. In the latter event, LHR is bypassed and stimulation of these responses occurs through direct activation of adenylyl cyclase followed by subsequent elevation of intracellular cAMP signaling pathways. Therefore, the effects of PRL on hCG-stimulated cAMP, P and StAR responses are mostly mediated through the LHR, a step appears to be present between the LHR and activation of GTP-binding protein (Gs). It is possible that PRL action can be attributed to alternative pathway(s), as also LHR in Leydig cells is known to be coupled with different second messengers, i.e. cAMP, Ca2+, chloride channels and arachidonic acid (55, 56, 57). The PRL-mediated stimulatory and inhibitory effects were specific, as evidenced by unaltered total protein contents and DNA synthesis of the different treatment groups. These results are in agreement with other studies, which demonstrated that PRL has no effects on Leydig cell proliferation (16, 58). However, it should be taken in to account that effects of PRL on Leydig cells vary between species, and even within the same species.
In mouse Leydig tumor cells, we (22, 23) and others (20) have demonstrated that hormonal stimulation of the StAR protein expression is intimately associated with steroid hormone biosynthesis. In accordance, a 48-h PRL treatment in the present study significantly increased hCG-stimulated StAR mRNA expression with 1–10 µg/liter of PRL, after which a subsequent decline of StAR expression occurred following treatments with higher PRL levels, which changes were concomitant with similar responses of P production. In addition, Northern blot analysis revealed three StAR transcripts (3.4, 2.8, and 1.6 kb) with coordinate regulation, all of them long enough to encode the full-length StAR protein (22).
The physiologic relevance of the PRL actions observed in mLTC-1 cells was corroborated by a similar biphasic response observed in conjunction with hCG-stimulated testosterone production of primary mouse Leydig cells. Consistent with our finding, biphasic responses of hCG-stimulated steroidogenesis to PRL have been reported in cultured adult rat Leydig cells (14, 15). The concentrations of PRL used in the above in vitro studies corresponded to circulating PRL levels attained when demonstrating PRL effects on rodent testicular function in vivo (59, 60). Hence, the in vitro findings of the present and earlier studies can be considered to reflect PRL effects in vivo. However, extrapolation of the present findings to the human are not warranted because there is no clear evidence for the presence of functionally meaningful levels of PRL receptors in the human testis (61). The hypogonadal effects of hyperprolactinemia in men are generally considered to be indirect, through suppression of gonadotropin secretion (62).
The biphasic PRL effect on levels of hCG-mediated cAMP, P, and StAR expression showed close correlation with the responses observed in LHR, PRLR and JAK2 mRNA levels. The 4.8 kb JAK2 mRNA transcript observed in the mLTC-1 cells by Northern analysis was consistent with an earlier report (63). The stimulatory effects observed with PRL at lower concentrations comply with the physiological ranges of PRL in vivo in maintaining steroid hormone biosynthesis, whereas conversely, the inhibitory doses largely corresponded to the pathological serum PRL levels occurring in hyperprolactinemia (59, 60, 62). Moreover, short-term effect of PRL at lower concentrations clearly modulated the uptake of 45Ca2+ through transmembrane Ca2+ mobilization, whereas the increase in 45Ca2+ uptake was lost at higher concentrations. In accordance, increase in [Ca2+]i has been demonstrated in CHO cells expressing functional PRLR following PRL stimulation (34, 36).
The role of Ca2+ as a second messenger has been amply reported in the regulation of diverse cellular functions. Recent data document that Ca2+ plays a crucial role in potentiating the levels of StAR expression and steroidogenesis in gonadal and adrenal cells (22, 64). Our results show that extracellular Ca2+ increased the long-term PRL-modulated hCG response of StAR gene expression and steroidogenesis in agreement with our previous finding (22). Specificity of the Ca2+ action was evaluated by using its chelator or ion-channel blocker, and both abolished the Ca2+ effect on hCG-stimulated StAR expression and steroid production. The most plausible interpretation of these results is that Ca2+ enhances similarly both the StAR response to hCG and the additional increase in this response brought about by PRL. No clear evidence for specific Ca2+ enhanced PRL response could be observed in these experiments.
The mLTC-1 cells treated for 48 h with increasing doses of PRL demonstrated a biphasic response pattern on [125I]iodo-hGH binding (data not shown). Other studies have shown by cross-linking of [125I]iodo-hGH two forms of PRLR, 39 and 101 kDa, in MA-10 cells (65). Down-regulation of PRLR expression was evident by treating these cells for 24 h with increasing concentrations of PRL. In accordance, we observed in mLTC-1 cells a significant amounts of PRLR mRNA by Northern analysis, and the levels were up-regulated by exposure to low, and down regulated to higher PRL concentrations.
Collectively, our data clarify certain mechanisms responsible for the inhibition of rodent testicular functions by hyperprolactinemia, because high PRL concentrations were found to inhibit LHR expression and other specific functions of mLTC-1 cells. Longer exposure of mLTC-1 cells to lower doses of PRL (1–10 µg/liter) demonstrated up-regulation of the signaling pathways investigated, in keeping with another, stimulatory, effect of this hormone. Over and above, PRL appears to exert its pleomorphic regulatory effects on mouse Leydig cells in a time- and dose-dependent fashion. The mechanisms behind the complexity of PRL actions on different signal pathways need further exploration.
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Acknowledgments |
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Footnotes |
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2 Equal contributors to this study.
Received April 5, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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T. Hamalainen, J. Kero, M. Poutanen, and I. Huhtaniemi Transgenic Mice Harboring Murine Luteinizing Hormone Receptor Promoter/{beta}-Galactosidase Fusion Genes: Different Structural and Hormonal Requirements of Expression in the Testis, Ovary, and Adrenal Gland Endocrinology, October 1, 2002; 143(10): 4096 - 4103. [Abstract] [Full Text] [PDF]
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