| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
ARTICLES |
Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot 76100, Israel
Address all correspondence and requests for reprints to: Dr. Abraham Amsterdam, Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot 76100, Israel. E-mail: abraham.amsterdam{at}weizmann.ac.il
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
. Forskolin did not stimulate apoptosis when incubated with
these cells. However, it augmented by 1.2-fold the p53-induced
apoptosis in cells shifted from 37 to 32 C. Dex further enhanced
apoptosis by 1.9-fold in p53-activated cultures (32 C). Incubation of
the cells with Dex dramatically reduced Bcl-2 levels to 15% of control
at 37 C (P < 0.01) or 32 C in the presence or
absence of forskolin (P < 0.01). Our data suggest
that glucocorticoids exert a protective effect against induced
apoptosis in immortalized granulosa cells and a stimulatory
effect on apoptosis in myeloid leukemia cells. Moreover, modulation of
Bcl-2 levels plays an important role in mediating the glucocorticoid
effect on cell survival. The opposite effect of glucocorticoids on
Bcl-2 levels in the two cell lines may be due to the different
ontogeneses of the two cell types: epithelial for granulosa cells
vs. mesenchymal for myeloid cells studied in the present
work. |
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Highly steroidogenic human granulosa cell lines were recently established by triple transfection with simian virus 40 DNA, Ha-ras oncogene, and the temperature-sensitive (TS) mutant of p53 (p53 Val135) (12). Although these immortalized cells (HO-23) lose their ability to respond to gonadotropins (LH/hCG/and FSH), they secrete progesterone at levels comparable to primary human granulosa cells in response to forskolin (FK) or 8-bromo-cAMP (12). These cells preserve their high steroidogenic capacity by expressing the Ad4 binding protein/steroidogenic factor-1 (Ad4BP/SF-1), which is characteristic of a steroidogenic tissue (13, 14), and by enhancement of expression, in a cAMP-dependent manner, of the steroidogenic acute regulatory protein (StAR), which is an essential and rate limiting factor in steroidogenesis that is responsible for the transport of cholesterol into mitochondria (15, 16), and the cytochrome P450 side-chain cleavage enzyme system (17, 18, 19). HO-23 cells can undergo rapid and massive apoptosis after shifting the temperature of growth of cAMP-stimulated cells from 37 to 32 C, which allows manifestation of the wild-type p53 activity (20). Therefore, establishment of immortalized human granulosa cell lines that keep their high steroidogenic potential allow systemic studies of the cellular and molecular mechanisms that control steroidogenesis and apoptosis in the most abundant somatic follicular cells, the granulosa cells (21).
Early observation demonstrated that glucocorticoids augment the
gonadotropin-induced production of progesterone in MA-10 Leydig tumor
cells (22) and immature preantral rat granulosa cells
(23). Recently, dexamethasone (Dex) was demonstrated to
enhance progesterone, pregnenolone, and 20-hydroxy-4-pregnen-3-one
in primary rat granulosa cells prepared from preovulatory follicles and
in immortalized rat granulosa cells (24). However, to date
no work has been reported on the effect of glucocorticoids on
cAMP-induced steroidogenesis in primary or immortalized human granulosa
cells. We therefore decided to test in detail the effect of
glucocorticoids on steroidogenesis in HO-23 immortalized human
granulosa cells.
Ovarian cell death is an essential process for the homeostasis of ovarian function in human and other mammalian species (reviewed in Refs. 21, 25 , and 26). Recent observations suggest that degeneration of follicular and luteal cells is mediated via programmed cell death (27, 28, 29). Typical morphological and biochemical events characterizing apoptosis have been observed in primary human and rat granulosa cells obtained from either preantral or preovulatory follicles. These include chromatin condensation and fragmentation of DNA (11, 29, 30). Estradiol was found to act as a survival factor in both the rabbit corpus luteum and granulosa cells, and it is associated with a shift in the expression of Bcl-2 gene family members (31). In contrast to estrogen, androgens enhance ovarian granulosa cell apoptosis from rats (32), and progesterone was recently suggested to suppress apoptosis in human granulosa cell (33). Androgens were reported to antagonize the protective effect of diethylstilbestrol (DES), which exerts an estrogen-like activity, in attenuating rat granulosa cells apoptosis (32).
Glucocorticoids induced apoptosis in most cells of the vascular system, such as rat thymocytes, myeloma cells, and peripheral blood monocytes (34, 35, 36, 37). In contrast, in the mouse mammary gland, glucocorticoid and progesterone inhibited involution and programmed cell death (38). The effect of glucocorticoids on apoptosis in the female reproductive organs is not well understood. Dex did not stimulate apoptosis in cultured porcine granulosa cells (39). However, no data were presented in that work showing whether Dex could protect against apoptotic stimuli such as serum deprivation. Thus, the effect of Dex on apoptosis in human ovary is not completely understood. In the present report we demonstrate that Dex and hydrocortisone protect against serum deprivation-, cAMP-, and p53-induced apoptosis in immortalized HO-23 human granulosa cells. The protective effect involved an increase in Bcl-2 levels. Moreover, glucocorticoid protection against apoptosis does not involve stimulation of the cell cycle.
As glucocorticoids induce apoptosis in most cells of the vascular system, such as rat thymocytes, myeloma cells, and peripheral blood cells (34, 35, 36, 37), it seemed essential to compare the effect of glucocorticoid on a representative cell type that is sensitive to glucocorticoid-induced apoptosis. We chose LTR-6 myeloma cells because they express the same TS mutant of p53 (Val135) as the HO-23 cells and discovered that the glucocorticoid-induced apoptosis in myeloid cells involved down-regulation of the intracellular level of Bcl-2.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
(GR), and monoclonal mouse
antihuman Mdm2 (Marine Double Minute 2(20) were purchased from
Santa Cruz Biotechnology, Inc. (Heidelberg, Germany). Goat
antirabbit and antimouse IgG coupled to horseradish peroxidase were
obtained from Biomakor (Rehovot, Israel). Antihuman StAR antibodies
were provided by Dr. J. F. Strauss III (University of Pennsylvania
Medical Center, Philadelphia, PA). Antihuman adrenodoxin polyclonal
antibodies were provided by Dr. W. L. Miller (University of
California, San Francisco, CA).
Reagents
Reagents for terminal deoxynucleotidyltransferase-mediated dUTP
nick end labeling (TUNEL) were purchased from Roche Molecular Biochemicals, (Mannheim, Germany). FK (a potent activator of
adenylate cyclase), Dex, and 4',6-diamido-2-phenylindole hydrochloride
(DAPI; for DNA staining) were purchased from Sigma. RU-486
was a gift from Dr. F. Kohen (Department of Biological Regulation,
Weizmann Institute of Science).
Cells and treatments
The human granulosa cell line HO-23 was established by triple
transfection of primary human granulosa cells with simian virus 40 DNA,
Ha-ras oncogen, and a TS mutant of p53 (p53
Val135) as described previously
(12). Shifting the temperature from 37 to 32 C leads to
manifestation of wild-type p53 activity (28, 40). The
LTR-6 cell line was derived from mouse M1 myeloid leukemia
cells, which contain a stable, transfected TS mutant of p53
(Val135 p53). At 32 C wild-type p53 function was
exerted by its mutant in LTR6 cells, resulting in programmed cell death
(41).
Biochemical assays
Progesterone was determined by RIA at the end of stimulation
(42, 43). Protein was quantified by the Bradford method
(44).
Western blot analysis
Cells were washed with cold PBS and harvested with a rubber
policeman using lysis buffer containing 50 mM HEPES (pH
7.2), 150 mM NaCl, 1.5 mM
MgCl2, 1 mM EGTA, 1 mM
phenylmethylsulfonylfluoride, 1% Triton X-100, 10 µg/ml leupeptin,
10% glycerol, 30 mM NaF, 30 mM sodium
pyrophosphate, 1 mM orthovanadate, and 5 µg/ml aprotinin.
Lysates were boiled in sample buffer for 10 min. Samples containing
equal amounts of protein (30 µg) were separated by 12% (to detect
Bcl-2) or 10% (to detect GR and Mdm2) SDS-PAGE. The blots were
blocked using 5% milk powder in PBS plus 0.05% Tween-20 and reacted
(overnight, 4 C) with antihuman Bcl-2 antibody or antihuman Mdm2
antihuman GR, followed by 1-h incubation at room temperature with
goat antirabbit or antimouse IgG conjugated to horseradish peroxidase.
The detection was carried out using enhanced chemiluminescence
(12, 20).
Phase contrast and immunofluorescent microscopy
Staining for DAPI and TUNEL reaction were performed on cells
grown on glass coverslips. At the end of culture period, medium was
aspirated and stored at -20 C for progesterone measurement. Cells were
washed with PBS, fixed with 3% paraformaldehyde for 30 min in 24 C,
and permeabilized for 4 min with 1% Triton X-100 in PBS at 4 C,
incubated with TUNEL reaction mixture (fluorescein-deoxy-UTP and
terminal deoxynucleotidyl transferase) for 60 min at 37 C in a
humidified atmosphere in the dark. Cells were washed with PBS and
stained with DAPI for 30 min at 24 C (20). Cells were
washed intensively with PBS, mounted in Mowiol, and analyzed by
Carl Zeiss Axioskop Microscope (Carl Zeiss,
Oberkochen, Germany) in both phase and fluorescent modes.
Fluorescence-activated cell sorting (FACS) analysis of DNA
content
Floating cells and trypsinized cells, which were originally
attached to the bottom of the culture dishes from each treatment, were
collected and combined to ensure a complete recovery of the cell
population. Cells were washed with cold PBS and fixed in cold methanol
(-20 C) for 1 h. Subsequently, cells were centrifuged,
resuspended in 0.5 ml cold PBS, and stained for 15 min with 50 µg/ml
propidium iodide in the presence of ribonuclease A (100 µg/ml). Cells
were then analyzed in the fluorescence-activated cell sorter. Five
thousand events from the gated subpopulation were recorded separately
(20).
Statistical analysis
Analysis of progesterone production and densitometer tracing on
Western blots autoradiograms (mean ± SD) were
performed using ANOVA followed by Fisher’s least significant
difference test. Differences between treatment groups were considered
statistically significant at P < 0.05.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
). Production
of progesterone in nonstimulated cells was 10
pg/106 cells·24 h. FK elevated progesterone
production by 240- and 180-fold (P < 0. 05) at 37 C
for 29 h and at 24 h at 37 C, followed by 5 h at 32 C,
respectively. Dex had no effect on progesterone production by itself,
but enhanced the level of FK-stimulated progesterone production by 4-
and 8-fold at 37 and 32 C respectively (P < 0.01; Fig. 1A). To verify whether this synergistic effect of Dex and FK on
progesterone production involves increased expression of steroidogenic
enzymes, we analyzed cell lysates by Western blot. Although there was a
clear induction of StAR and ADX by forskolin (which is a specific
marker for the cytochrome P450 side chain cleavage enzyme system), no
further induction of expression of these gene products was revealed in
the presence of Dex (Fig. 1B), suggesting that enhancement of
steroidogenesis by Dex in the absence of serum does not involve
regulation of StAR and ADX.
|
) and FACS analysis of propidium
iodide-treated cells (Figs. 3 and 4) in which the
Go/G1 subfraction of cells
(peak I) was considered the apoptotic population, and peaks II and III
represent cells with 2N and 4N chromosomes characteristic of this cell
line (20). At 37 C it was clearly demonstrated by all the
methods that Dex protects against serum deprivation- and FK-induced
apoptosis (Figs. 2 and 3). Shifting the temperature to 32 C, which
activates p53, led to a 4.3-fold increase in the rate of apoptosis. FK
increased apoptosis by 2-fold at 37 C and by 1.7-fold at 32 C compared
with that in nonstimulated cells at the same temperature (Fig. 4)
|
|
|
, A and B). Hydrocortisone (100 nM)
blocked serum deprivation-induced apoptosis to 50% at 37 C and to
94% at 32 C. Hydrocortisone (100 nM) also attenuated
cAMP-induced apoptosis to 50% at 37 C and to 80% at 32 C.
Glucocorticoids are therefore able to attenuate apoptosis induced by
serum deprivation, p53 activation, or forskolin stimulation. As shown
in Fig. 3, the protection by Dex against apoptosis did not involve
changes in the incidence of cells in the G2/M
phase (peak IV) (20), suggesting that this protection does
not involve stimulation of cell proliferation.
Does Dex protect against apoptosis via interaction with the
GR?
To examine the sensitivity of the cells to the protection against
apoptosis by Dex, cells starved for 24 h were incubated during
this period with increasing concentrations of Dex. Dex protected
against apoptosis in a dose-dependent manner with an
ED50 of 7 nM, and 80% of protection
was achieved already at 10 nM (Fig 5).
|
).
Protective effect of glucocorticoids against granulosa cell
apoptosis involves increase in Bcl-2 levels
To examine whether such a protection involves modulation of the
expression of tumor suppressor genes or survival genes, we tested the
intracellular levels of Bcl-2 and Mdm2. Cells treated at 37 C or 32 C
with 100 nM Dex showed significant elevation of Bcl-2
expression compared with nontreated cells (3.1-fold at 37 C and 3-fold
at 32 C; P < 0.01; Fig 6). Thus, an increase in Bcl-2 levels
stimulated by Dex may be involved in the protective effect of Dex on
apoptosis induced by p53 activation (Fig. 6) and serum deprivation (not
shown in Fig. 6). Stimulation of cells with 20
µM forskolin for 29 h at 37 C or for
24 h at 37 C followed by 2 or 5 h at 32 C reduced the level
of Bcl-2 by 50% (P < 0.05). This decrease in Bcl-2
level exerted by forskolin was completely abolished by 100
nM Dex. Mdm2 expression was not affected by Dex
treatment, although its expression was reduced by forskolin
(20).
|
(Fig. 7). LTR-6 cells were treated with 100
nM Dex at 37 and 32 C with or without 20 µM
forskolin, a concentration similar to that used for treatment of HO-23
human granulosa cells. Cells incubated at 32 C were stimulated for
apoptosis due to activation of the mutant p53 (2-fold), but FK was
without effect (Fig. 8B). Interestingly
100 nM Dex clearly enhanced apoptosis by 3.5-fold at 37 C
and by 1.9-fold at 32 C compared with cells incubated either at 37 or
32 C without Dex (Fig. 8, A and B). FK did not augment apoptosis in the
presence of Dex (Fig. 8B). The apoptotic effect of Dex was mainly due
to the shift of the G1/G0
population to the G0 subpopulation, which is
characteristic for apoptotic cells (Fig. 8A, peak I). These data
suggest that Dex exerts an opposite apoptotic effect in myeloid
leukemia cells, in contrast to its effect in HO-23 cells.
|
|
). Forskolin (20
µM) only slightly reduced Bcl-2 levels at 37 C.
However, it did not affect Bcl-2 levels at 32 C, but reduced Bcl-2
levels to 15% (P < 0.01) if cells were coincubated
with Dex and forskolin. Thus, Bcl-2 levels are involved at least in
part in the apoptotic signal exerted by Dex in LTR6 myeloid leukemia
cells.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
As glucocorticoids are implicated in induction of apoptosis in many
cell types, such as immature thymocytes (48), mature
activated peripheral T lymphocytes (49, 50), several
leukemic cell lines (51), and multiple myeloma-derived
cell lines (36), our finding of the protection against
granulosa cell apoptosis is somewhat unusual, although it was recently
demonstrated that glucocorticoids attenuate apoptosis induced by serum
deprivation in human peripheral blood neutrophils (52) and
suppress apoptosis induced by tumor necrosis factor in glomerular
endothelial cells (53) and by interferon-
and
transforming growth factor-ß1 in lung epithelial cells and in
hepatoma cells, respectively (54, 55, 56).
Nevertheless, in the present work we demonstrate for the first time that glucocorticoid hormones such as dexamethasone and hydrocortisone can inhibit p53- and cAMP-induced apoptosis, which are among the most common effectors of apoptosis in normal and immortalized granulosa cells (11, 28, 30, 57).
p53 levels are enhanced dramatically after x-ray and ionic radiation as well as by chemotherapy with cisplatin, hypoxia, nitric oxide, and other stresses that may lead to massive apoptosis in target cells (reviewed in Ref. 58). Elevation of nuclear p53 protein in apoptotic granulosa cells was clearly demonstrated in atretic follicles, and activation of p53 was correlated with apoptosis in steroidogenic immortalized rat and human granulosa cells (20, 28, 47). We suggest that glucocorticoids, in particular in epithelial cells such as ovarian granulosa, mammary gland, and liver parenchymal, may negate the p53 signals for apoptosis and therefore may prevent excessive damage of essential glandular cells and tissue after different kinds of stress stimuli implicated with p53 up-regulation.
Dex exerts its antiapoptotic effect in a dose-dependant manner with an ED50 of 7 nM, which is in line with the Kd found in human osteosarcoma cell lines (59), and can be blocked by RU486, which has been shown to inhibit the release of GR from cytoplasmic complexes containing heat shock proteins, thus suppressing the subsequent translocation of receptors into the nucleus (60, 61). Taken together, our observation of the low ED50 for inhibition of apoptosis and the blocking of this effect by RU486 strongly suggest that the antiapoptotic effect of glucocorticoid in granulosa cells is exerted through interaction with its specific receptor. Indeed, we identified GRs in HO-23 cells by Western blot.
Nonstimulated HO-23 cells produce extremely low levels of progesterone (10 pg/106·24 h). However, it is very unlikely that this amount of progesterone contributes significantly to the antiapoptotic effect of Dex exerted by serum deprivation or p53 activation, as the actual concentration of progesterone in these cell cultures is 0.01 nM, which is 150 times lower than the Kd level (1.5 nM) calculated for the affinity of progesterone to its receptor (62). It is even more unlikely that FK-induced steroidogenesis contributes to the protective effect of Dex, as the elevation in progesterone produced by cAMP stimulation coincides with an elevation rather than a decrease in apoptosis in HO-23 cells. However, we cannot exclude the possibility that progesterone may have some antiapoptotic effect in primary human granulosa cells (33).
We demonstrated that Dex increases cAMP-induced steroidogenesis in immortalized granulosa cells (24), but no information is available on specific enhancement of cAMP-induced steroidogenesis in human granulosa cells. In the present work we demonstrated that Dex can enhance cAMP-induced steroidogenesis concomitantly with attenuation of apoptosis, and in preliminary work we show that Dex enhances hCG/cAMP-induced steroidogenesis in preovulatory human granulosa cells (Sasson, R., and A. Amsterdam, in preparation), suggesting that this process can explain at least in part the enhancement of steroidogenesis by Dex. Although we cannot rule out that Dex may moderately change the expression of steroidogenic enzymes or factors as was demonstrated for the FSH-17 rat granulosa cell line (24), we did not find significant changes in the expression of StAR and ADX by Dex in cAMP-treated HO-23 cells. The main differences between the previous (24) and the present work is the use of cells of different mammalian origins and that the stimulation of steroidogenesis was performed in serum-free medium, whereas previous experiments were performed in the presence of 5% FCS, which may contain growth factors and other factors that may enhance de novo synthesis of StAR and ADX. In the absence of serum, the inhibition of apoptosis by Dex is probably the main cause of the enhanced cAMP-induced steroidogenesis, especially in cultures of HO-23 cells that were incubated at 32 C, which dramatically activated the apoptotic effect of p53.
We demonstrated that Dex significantly changes Bcl-2 levels. It was recently demonstrated that Dex suppresses apoptosis in human gastric cancer cell lines through modulation of bcl-x gene expression (63). Inhibition by Dex of transforming growth factor-induced apoptosis in rat hepatoma cells is associated with Bcl-xL induction, another survival gene of the Bcl-2 family (56). However, in this work we for the first time report the involvement of Bcl-2 in Dex protection against apoptosis through the up-regulation of its intracellular levels. Whether Dex can enhance other survival factors that may protect against ovarian cell death, such as Bcl-xL, should be explored in the future. Moreover, the possibility that glucocorticoids can enhance specific ovarian survival factors, such as inhibitor of apoptosis proteins (64) and growth differentiation factor-9 (65) secreted by the egg, should be explored either in vivo or in intact follicles in vitro.
Glucocorticoids are known to induce apoptosis in acute myeloid leukemia cell lines and in other cell types, such as mature activated peripheral T lymphocytes (66, 67). However, the mechanism is not completely understood. On the one hand, in myeloid leukemia cells lines no change in Bcl-2 level was recorded upon Dex-induced apoptosis. On the other hand, it was suggested that the Bcl-2 level in T lymphocytes can switch the effect of glucocorticoids from proapoptotic to antiapoptotic when lymphocytes expressing Bcl-2 are exposed to other apoptotic stimuli, such as serum depletion (68). In the present work we observed that in the LTR-6 myeloid leukemia cell line expressing the same Ts p53 plasmid (Val135 p53) and the same type of GR as in the granulosa cell line HO-23, glucocorticoids enhanced p53-induced apoptosis, in contrast to HO-23 cells, where glucocorticoids suppressed p53-induced apoptosis. Thus, different cell types can respond in opposite ways to stimulation by Dex, which may involve in both cases modulation of the Bcl-2 level in an opposite manner. Here we observed a situation in which Dex can display different activities in different cell types, implying some sort of communication with other factors that affect or reflect cell specificity (69). It was already shown that the cell specificity of GR action at PifG (proliferin gene) is determined by the composition of AP-1, specifically the ratio of c-Jun to c-Fos (69, 70). F9 cells, which lack AP-1 activity under appropriate culture conditions, displayed no hormonal regulation from PifG-linked CAT sequences; HeLa cells, which express AP-1 predominantly as c-Jun homodimers, enhanced reporter expression in response to the steroid hormone Dex, and CV-1 cells, in which AP-1 is comprised mostly of c-Jun and c-Fos heterodimers, repressed reporter gene expression upon hormone addition (69). Thus, the ratio of c-Jun and c-Fos can sometimes operate to determine the different glucocorticoid responses in different cells.
The roles of glucocorticoids in the direct regulation of ovarian function are poorly understood. However, it was recently hypothesized that glucocorticoids serve an antiinflammatory role during ovulation, thereby promoting rapid healing of the wound left by follicular rupture (71). Glucocorticoid interference with PG synthesis (71) and with granulosa cell apoptosis as described in the present paper in HO-23 cells may deepen our understanding of the mechanism by which glucocorticoids exert their antiinflammatory action in the ovary.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
2 Incumbent of the Joyce and Ben B. Eisenberg Chair of Molecular
Endocrinology and Cancer Research at Weizmann Institute of Science
(Rehovot, Israel).
Received May 30, 2000.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
and
lipopolysacharide-induced apoptotic cell death in bovine glomerular
endothelial cells upstream of caspase 3 activation. Br J Pharmacol 127:1633–1640[CrossRef][Medline]
cytotoxicity. J Endocrinol Invest 16:591–599[Medline]
and Fas. Am J Physiol 273:L921–L929
This article has been cited by other articles:
 |
|
 |
|
  J. Komiyama, R. Nishimura, H.-Y. Lee, R. Sakumoto, M. Tetsuka, T. J. Acosta, D. J. Skarzynski, and K. Okuda Cortisol Is a Suppressor of Apoptosis in Bovine Corpus Luteum Biol Reprod, May 1, 2008; 78(5): 888 - 895. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Myers, M. C. Lamont, S. van den Driesche, N. Mary, K. J. Thong, S. G. Hillier, and W. C. Duncan Role of Luteal Glucocorticoid Metabolism during Maternal Recognition of Pregnancy in Women Endocrinology, December 1, 2007; 148(12): 5769 - 5779. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Sunak, D. F Green, L. R Abeydeera, L. M Thurston, and A. E Michael Implication of cortisol and 11{beta}-hydroxysteroid dehydrogenase enzymes in the development of porcine (Sus scrofa domestica) ovarian follicles and cysts Reproduction, June 1, 2007; 133(6): 1149 - 1158. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Harms, K. Albrecht, U. Harms, K. Seidel, L. Hauck, T. Baldinger, D. Hubner, G. Kronenberg, J. An, K. Ruscher, et al. Phosphatidylinositol 3-Akt-Kinase-Dependent Phosphorylation of p21Waf1/Cip1 as a Novel Mechanism of Neuroprotection by Glucocorticoids J. Neurosci., April 25, 2007; 27(17): 4562 - 4571. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Engmann, R. Losel, M. Wehling, and J. J. Peluso Progesterone Regulation of Human Granulosa/Luteal Cell Viability by an RU486-Independent Mechanism J. Clin. Endocrinol. Metab., December 1, 2006; 91(12): 4962 - 4968. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. N. Fru, C. A. VandeVoort, and C. L. Chaffin Mineralocorticoid Synthesis During the Periovulatory Interval in Macaques Biol Reprod, October 1, 2006; 75(4): 568 - 574. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A Higuchi, S Shimmura, T Takeuchi, M Suematsu, and K Tsubota Elucidation of apoptosis induced by serum deprivation in cultured conjunctival epithelial cells Br. J. Ophthalmol., June 1, 2006; 90(6): 760 - 764. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Ranta, D. Avram, S. Berchtold, M. Dufer, G. Drews, F. Lang, and S. Ullrich Dexamethasone induces cell death in insulin-secreting cells, an effect reversed by exendin-4. Diabetes, May 1, 2006; 55(5): 1380 - 1390. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. M. Moore, C. J. Merkle, P. Miketova, R. K. Salyer, B. J. Torres, R. C. Schaeffer Jr, and D. W. Montgomery Cytosine arabinoside induces programmed endothelial cell death through the caspase-3 pathway.</ |
