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in Rat Testis: In Vivo Response to Depletion and Repletion of Vitamin A
Department of Genetics and Cell Biology, Department of Biochemistry and Biophysics, Center for Reproductive Biology, Washington State University, Pullman, Washington 99164
Address all correspondence and requests for reprints to: Kwan Hee Kim, Department of Genetics and Cell Biology, Washington State University, Pullman, Washington 99164-4234. E-mail: khkim{at}wsu.edu
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Abstract |
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(RAR). This degeneration can be reversed by injecting retinol, a
precursor of RA, into vitamin A-deficient (VAD) rats. To determine the
relationship between this ligand-dependent testicular degeneration and
regeneration and the expression levels of RAR messenger RNA and
protein, testes were depleted and then replenished with retinol
in vivo. Results showed that RAR messenger RNA and
protein levels declined to VAD amounts after 7 weeks on a VAD diet.
This decline was due to decreased RAR levels in early meiotic
spermatocytes and the loss of advanced germ cells. Interestingly, the
advanced germ cells still contained RAR, but the protein was
primarily cytoplasmic instead of nuclear, indicating inactivity as a
transcription factor. In VAD testis, RAR levels were low and then
increased primarily in Sertoli cells after retinol replenishment. TUNEL
analyses showed that most germ cells at the basal aspect of
seminiferous tubules were undergoing apoptosis during degeneration.
These results indicate that RAR is either down-regulated or
inactivated in RA-deficient testis and coincident with that, testes
degenerate by apoptosis or selective loss of germ cells. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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A single injection of retinol, a precursor of RA, into VAD rats reinitiates spermatogenesis in a synchronous manner such that the testis contains only three or four spermatogenic stages instead of the fourteen that are found in an adult rat (4). Repopulation of the testis occurs primarily from the remaining type A1 spermatogonia (8, 10) as most of the preleptotene spermatocytes that remain in the VAD testis degenerate by an unknown mechanism and do not contribute to the repopulation of the testis (8, 10).
Nuclear retinoid receptors are thought to mediate the action of RA (for
review, see Ref.11). Two families of retinoid receptors have been
identified: the retinoic acid receptors (RAR) that recognize
all-trans RA as their ligand and the retinoid X receptors
(RXR) that recognize 9-cis RA as their ligand. Each family consists of
three subtypes of receptors, , ß, and 
. These receptors are
transcription factors that regulate the transcription of genes
containing appropriate RAR response elements (RARE) or RXR response
elements (RXRE).
One of the six receptors, RAR, has been shown to play a critical
role in the vitamin A regulation of spermatogenesis in the testis.
RAR knockout mice were observed to have a testicular phenotype
similar to that seen in VAD rats (2, 12). There are two major
transcripts for RAR expressed in the testis, 3.4 kb and 2.7 kb (13).
The 3.4-kb transcript is expressed in both Sertoli cells and germ
cells, whereas the 2.7-kb transcript is predominantly expressed in
Sertoli cells. In situ hybridization and
immunohistochemistry have been used to localize RAR messenger RNA
(mRNA) and protein to Sertoli cells, early meiotic spermatocytes, and
round spermatids undergoing elongation processes (14).
In addition, the relative amounts of the 2.7-kb and 3.4-kb transcripts
were shown by Northern blot analysis to increase 2- to 3-fold in
Sertoli cells within 30 min of retinol repletion of VAD rats (13, 15).
However, it has not been determined whether relative amounts of RAR
mRNA and protein change during degeneration of the testis in animals on
a VAD diet. If relative amounts of RAR decline during the
development of the VAD condition, then this would explain why the
testes of the VAD rats and the RAR knockout mice have nearly
identical phenotypes.
In this study, we have examined how retinol depletion and repletion
affect the expression and localization of RAR mRNA and proteins in
rat testis. We have also examined whether testicular degeneration
arises in part from germ cell apoptosis during retinol depletion. The
results indicate that there is a correlation between germ cell
apoptosis and RAR mRNA and protein expression.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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RNA isolation and Northern blot analyses
RNA was isolated from the degenerating testes of at least two
animals per time point using the standard method of homogenization
followed by extraction with phenol and guanidium isothiocyanate
(16).
For Northern blot analyses, 30 µg of RNA per sample were separated by
electrophoresis through an 1.0% formaldehyde-agarose gel, transferred
to nylon filters (Micron Separations, Inc., Westboro, MA) and
cross-linked with UV irradiation. Escherichia coli and rat
ribosomal RNAs were also subjected to electrophoresis in the same gels
as markers. To prepare the complementary DNA (cDNA) probe, the 1389-bp
RAR cDNA was excised with EcoRI restriction endonuclease,
purified, and radiolabeled (17). Hybridization and washes were
conducted as previously described (17). A phosphorimaging screen
(Molecular Dynamics, Sunnyvale, CA) was exposed to filters overnight
and then scanned by a fast-scan image analyzer to determine the levels
of RNA hybridization (Image Quant, Software Version 4.2, Molecular
Dynamics, Sunnyvale, CA). Multiple blots using RNA from different
animals were prepared and comparable results were obtained.
In situ hybridization
Testes and epididymides from at least two male rats per
treatment were fixed in 4% paraformaldehyde/0.25% glutaraldehyde for
6 h at room temperature and then processed as previously described
(17). The antisense and sense rat RAR complementary RNA (cRNA), and
the antisense SGP-2 cRNA probes for in situ hybridization
were prepared as previously described (14, 17). Before hybridization,
the cRNA probes were cleaved to about 200 bp by alkaline hydrolysis, so
that they had similar access to cells in each tissue section. Serial
tissue sections were hybridized with the cRNA probes at 50°C for
20 h. After hybridization, slides were washed, treated with RNAse
A, and dehydrated (17). Slides were dipped in Kodak NTB-2 nuclear
tracking emulsion (Eastman Kodak, Rochester, NY), and exposed at 4°C
for 18 days.
Statistics and quantitation of tubules and silver grains
To quantify the frequency of tubules with silver grains in their
basal aspect (see Fig. 3
), at least 400 tubules per testicular section
were counted in testes from two rats on a VAD diet for 6 weeks and from
two normal adult rats.
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). At least 200 cells were counted over
a testicular cross-section or epididymal cross-sections from rats that
had been on a VAD diet for 7 weeks. Counts were performed on tissue
from two different animals. For comparison, silver grains were counted
over round spermatids from stages I, IV-V, and VIII in the
spermatogenic cycle of normal adult testis. At least 150 cells were
counted per circular-shaped seminiferous tubule section and three
tubule sections were counted per stage.
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), an area covering a
circular-shaped seminiferous tubule section was measured by the
Bioquant program. All of the silver grains within the area were counted
and counts per area calculated. Three tubule sections were counted per
animal and counts were performed for two animals.
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= 0.05 (Tukey’s HSD test,
Minitab, Inc., State College, PA).
Immunohistochemistry
Testes and epididymides from two rats for each time point were
fixed in Bouin’s and prepared as described previously (14, 17). The
immunohistochemical reactions were also as described previously (14, 17). The primary antibody was a polyclonal IgG raised in rabbits
against amino acids 443–462 (SCSPSLSPSSNRSSPATHSP) of the C-terminus
of human RAR (Santa Cruz Biotechnology, Santa Cruz, CA). The amino
acid sequence in the rat RAR was 100% homologous to the
corresponding human RAR peptide (our unpublished data). This
antibody was shown to recognize a 55-kDa protein from a rat testicular
extract on a Western blot (14). As a negative control, serial sections
were put through the same procedure without any primary antibody.
Additional negative control sections were incubated with an excess of
synthetic immunizing peptide (33 µg/ml, Santa Cruz Biotechnology,
Santa Cruz, CA) along with the anti-RAR antibody.
TUNEL (terminal deoxynucleotidyl transferase (TdT)-mediated
dUTP nick-end labeling) assay
The TUNEL assay was performed essentially as described by the
manufacturer of the Apoptosis Detection System (Promega Biotech
Corporation, Madison, WI). This assay involves the detection of
apoptotic cells by incorporation of fluorescein 12-dUTP using TdT at
the 3' OH end of fragmented DNA. Testicular and epididymal tissue from
at least two animals were prepared as described for in situ
hybridization. Tissue sections were further fixed in 4% methanol-free
formaldehyde before and after proteinase K digestion. Then, TdT
incubation buffer was added and the reaction carried out for 60 min at
37 C. The sections were washed in 2 x SSC (300 mM
NaCl, 30 mM sodium citrate), followed by PBS, and lightly
counterstained with hematoxylin. A positive reaction was characterized
by bright green fluorescence at the site of the fluorescein 12-dUTP
incorporation and was visualized using a fluorescence microscope. As a
negative control, sections were put through the same procedure without
TdT enzyme. A positive control was prepared by pretreating the sections
with DNase I, which resulted in multiple DNA fragments with 3'-OH ends
where fluorescein-dUTP could be incorporated.
Photography
Digital images were acquired using a Kodak DCS 420 digital
camera (Eastman Kodak, Rochester, NY) or a Pixera Studio 1.0 digital
camera (Pixera Corporation, Cupertino, CA). Figures were assembled and
labeled using Adobe Photoshop (Adobe Systems Inc., Mountain View, CA).
Figures were printed on a Tektronix Phaser 440 dye sublimation printer
(Tektronix, Inc., Wilsonville, OR).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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mRNA in testes from vitamin
A-depleting rats mRNA were observed to be low in
testes from rats on a VAD diet for 9 weeks compared with those in
testes from normal adult rats (13). Therefore, Northern blot analyses
(Fig. 1) were performed to determine when
RAR transcript numbers declined during the development of the VAD
condition. A significant decline in RAR mRNA occurred in testes from
rats on a VAD diet after 7 weeks (P 
0.05), and the
amounts were down to that seen in VAD animals by 7 weeks.
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transcripts was due
to a decrease in RAR mRNA in individual cells or to a loss of germ
cells from the testis, in situ hybridization was performed
on sections of testes from rats on a VAD diet for 6, 7, 8, or 9 weeks
(Fig. 2). After 6 weeks on a VAD diet,
the stage-specific pattern of RAR mRNA expression in normal adult
rat testes was still apparent (Fig. 2), and the frequency of the
spermatogenic stages was not significantly different from that in
normal adult testes (P 0.05; data not shown). In
contrast, there was a decrease in the number of silver grains over germ
cells delineating the basal aspect of seminiferous tubules in the
testes from the rats on a VAD diet for 6 and 7 weeks (compare Fig. 2A
with 2B and 2D; see arrowheads in A). After 7 weeks on a VAD
diet, the overall number of silver grains in the seminiferous
epithelium had declined further (Fig. 2D) compared with that seen in
testes from normal adult rats (Fig. 2A). This was partly due to the
sloughing off of advanced germ cells from the seminiferous epithelium
as silver grains were clearly seen over germ cells in the lumen of some
tubules (Fig. 2, D and F, see arrows). By week 9, the number
of silver grains in the seminiferous tubules was similar to background
(Fig. 2H).
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mRNA in cells delineating the basal aspect of the
seminiferous tubule (Fig. 2A
, see arrows), the proportion of
tubules demonstrating this pattern of expression was quantified in
normal adult testes and in testes from rats on a VAD diet for 6 weeks
(Fig. 3). There was a 2-fold decline in
the number of tubules with silver grains located at the basal aspect in
the testes from rats on a VAD diet for 6 weeks as compared with normal
adult testes (Fig. 3) (P 0.05).
Cellular localization of RAR mRNA in degenerating testis and
epididymis
To identify the relationship between expression of RAR
transcripts and the types of cells residing in the degenerating testis,
the testicular sections were examined at a higher magnification. Testes
from rats on a VAD diet for 6 weeks had no visible loss of germ cells
(compare Fig. 4A and E); however, the
presence of immature round germ cells as opposed to mature spermatozoa
in the epididymis indicated that some germ cells were being lost (data
not shown). After 7 weeks on a VAD diet, the loss of germ cells
resulted in an abnormal testicular morphology (Fig. 4B), although the
remaining germ cells continued to express RAR mRNA. Furthermore,
immature round germ cells that had sloughed from the testis and
accumulated in the epididymis still contained RAR mRNA (caput region
shown; Fig. 4F, arrows). By 8 weeks of vitamin A depletion,
there was even greater loss of germ cells as well as the presence of
multinucleated cells (Fig. 4C, arrow). By 9 weeks, the
testicular sections exhibited the classic VAD morphology with no
detectable lumen, and RAR mRNA was dispersed all over the tubule
sections (Fig. 4D).
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mRNA in the germ cells remaining
in the testes of rats on a VAD diet for 7 weeks was similar to that
found in the normal adult testis, silver grains were counted over round
spermatids that had sloughed off and were in the lumen of seminiferous
tubules or the lumen of the epididymis (Table 1). The amount of RAR
transcripts present in the round spermatids in the lumen of
seminiferous tubules and epididymides were not significantly different
than the average of silver grains counted for round spermatids at stage
I, stages IV-V, and stage VIII of the spermatogenic cycle in normal
adult testis (P 0.05).
In situ hybridization analysis of RAR transcripts in
retinol-replenished testis
The expression of RAR mRNA in testis has been shown to be
increased by retinol treatment of VAD rats (13, 15). For more extensive
analysis, in situ hybridization was performed on sections of
VAD testes and those collected at various times after treatment with
retinol (Fig. 5), and the number of
transcripts was quantified (Fig. 6). By
2 h after retinol treatment, the amount of RAR transcripts had
increased (Fig. 5C) and by 4 h, the amounts of RAR mRNA were
significantly higher (about 4-fold) than those found in the VAD testes
(P 0.05) (Figs. 5E and 6). Transcript numbers
declined significantly by 12 h postretinol replenishment
(P 0.05) (Fig. 5G, Fig. 6), and the number of
transcripts was similar to that in the VAD testes by 24 h (Figs. 5I and 6) (P 0.05).
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transcripts were seen scattered
throughout the tubule in VAD rats (Fig. 7A) indicating the presence of
transcripts primarily in the cytoplasm of Sertoli cells. This
expression of RAR mRNA in Sertoli cells after retinol replenishment
is further substantiated by the similarity of hybridization patterns
seen for the RAR mRNA 4 h after retinol treatment (Fig. 7C) and
SGP2 mRNA (Fig. 7E), which is known to be expressed primarily in
Sertoli cells (18). However, some expression in germ cells could not be
completely ruled out; although silver grains clustered over the
remaining germ cells should be apparent, if expression in germ cells
was occurring.
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protein in testis and
epididymis during vitamin A depletion
protein, immunohistochemistry was performed on testicular sections from
rats that were depleting and depleted of vitamin A. In degenerating
testes from rats on a VAD diet for 6 weeks, only a few early meiotic
germ cells contained RAR in the cytoplasm (Fig. 8, A and B, large arrow in
8B), and after 7 weeks, all early meiotic germ cells did not express
RAR (Fig. 8, C and D, large arrow in 8D). This was in
contrast to the presence of relatively high levels of RAR in the
nuclei of these cells lining the basal aspect of the normal adult
seminiferous tubules (Fig. 8, J and K, arrows).
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was observed in round (Fig. 8B)
and elongating spermatids (Fig. 8A) on sections from animals on the VAD
diet for 6 weeks. Following 7 weeks on the VAD diet, RAR protein was
present primarily in the remaining pachytene spermatocytes (Fig. 8, C
and D). However, in both spermatocytes and spermatids, the protein was
no longer localized to the nucleus, as in the normal adult testis, but
was cytoplasmic (Fig. 8, compare A–D with J and K). After 8 weeks on
the VAD diet, the RAR immunohistochemical staining was seen over
multinucleated giant cells (Fig. 8, E and F) and by 9 weeks, very
little RAR immunostaining was present in the tubule cross-sections
(Fig. 8G).
Immunohistochemistry of epididymides from animals on a VAD diet for 7
weeks revealed immunostaining for RAR in the cytoplasm of immature
germ cells that had sloughed from the seminiferous epithelium. These
cells contained RAR in the cytoplasm similar to that seen in germ
cells still residing in the testis (Fig. 8H). By 8 weeks on the VAD
diet, the only germ cells positive for RAR were in the cauda of the
epididymis, and at 9 weeks (VAD) virtually no germ cells in the
epididymis stained for RAR (data not shown).
Immunohistochemical analysis of RAR protein in
retinol-replenished testes
To determine the cellular localization of RAR protein during
retinol repletion, immunohistochemical analysis was performed on
sections of VAD testes and testes collected at 4, 8, and 24 h
after retinol treatment. In the VAD testis, immunostaining for RAR
protein was detectable, but low, with occasional tubules showing faint
cytoplasmic staining of Sertoli cells (Fig. 9A). However, by 4 h after retinol
injection, the intensity of the staining for RAR protein increased
(Fig. 9, B and E) and remained relatively high in Sertoli cells by
8 h following retinol injection (Fig. 9C). By 24 h
postretinol treatment, the immunostaining for RAR decreased to
amounts comparable with that seen in the VAD testes (Fig. 9D).
Localization of RAR in Sertoli cells was cytoplasmic, with more
intense staining seen near the nucleus (Fig. 9E). Only occasional
Sertoli cells showed nuclear staining for RAR. There was no
detectable staining for RAR in type A spermatogonia and preleptotene
spermatocytes in sections from retinol-treated animals.
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with
B–F). After 6 weeks on a VAD diet, a slight increase was seen in the
number of apoptotic cells at the basal aspect of some seminiferous
tubules (Fig. 10B). The relative number of apoptotic cells in the
seminiferous tubules was highest in the degenerating testis from
animals on a VAD diet for 7 weeks (Fig. 10, C and D), followed by a
decrease at week 8 (Fig. 10E) and was at its lowest in the testis from
VAD animals (Fig. 10F).
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). In the early degenerating testis
(6 and 7 weeks on a VAD diet), the apoptotic cells were mostly located
basally and appeared to be early meiotic germ cells (Fig. 11, A–D,
arrows). As degeneration progressed (7.5 to 8 weeks on a VAD
diet), pachytene spermatocytes situated closer to the lumen than the
early meiotic germ cells (Fig. 11E) and multinucleated giant cells
(Fig. 11H) became apoptotic.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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mRNA and
protein in Sertoli cells and early meiotic germ cells. For Sertoli
cells, the amounts of RAR mRNA and protein were seen to increase by
4 h after retinol injection during reinitiation of spermatogenesis
of VAD rats. For early meiotic germ cells, the amounts of RAR mRNA
and protein were high in the nucleus of germ cells in normal adults,
but decreased 2-fold by 6 weeks in animals on a VAD diet.
In addition, the results imply that the RAR isoform expressed in
Sertoli cells and early meiotic cells may be RAR2, which is induced
by RA. Alternative splicing of the RAR gene has been shown to
generate two major isoforms, RAR1 and RAR2, of which RAR2 has
been shown to contain a RARE inducible by RA, whereas RAR1 does not
contain a RARE (19). Whether or not RAR2 is the isoform expressed in
Sertoli cells and early meiotic germ cells requires further
investigation.
For late meiotic and haploid cells, the number of RAR transcripts
and protein in each cell in the seminiferous tubules or epididymides
was not decreasing during vitamin A depletion in animals on a VAD diet.
Instead, there was a change in the subcellular localization of RAR
from the nucleus to cytoplasm. Thus, it appears that RA regulates the
subcellular localization of the receptors in late meiotic and haploid
cells.
Changes in the subcellular localization of RAR have been reported
previously (20, 21, 22, 23). RA has been shown to cause translocation of the
promyelocyte-RAR fusion oncoprotein, PML-RAR, from the cytoplasmic
compartment to the nucleus (20). In addition, many phosphorylation
sites have been reported for RAR protein (21, 22), and modification
by phosphorylation has been shown to enhance nuclear localization of
the receptor (23). In this context, the changes in subcellular
localization seen for RAR during the development of the VAD
condition in testis could be regulated by phosphorylation. In any case,
it is clear that the changes in subcellular localization induced by RA
reflect the complexity of the functional organization within the cell,
and an understanding of RAR activity requires a precise knowledge
about how receptor localization is regulated within the cell.
The testis is thought to degenerate during vitamin A deficiency by two methods: germ cell death or premature loss of germ cells into the lumen of seminiferous tubules (6). We have demonstrated here that both germ cell death and premature loss of germ cells occur during vitamin A deficiency, but also that apoptosis mainly occurs in early meiotic germ cells, whereas some pachytene spermatocytes and most round spermatids detach from Sertoli cells and are lost.
Furthermore, a relationship between apoptosis and expression and
activity of RAR is noteworthy. The amounts of RAR transcripts and
protein declined in early meiotic germ cells in degenerating testis in
rats on a VAD diet for 6–7 weeks, and these cells were seen by the
TUNEL assay to be apoptotic. RAR may be important for survival of
early meiotic germ cells. This is consistent with a previously reported
finding that RAR is inhibitory to the RAR-specific apoptotic
pathway in thymocytes (24).
Alternatively, the induction of an apoptotic pathway in germ cells may
not involve RAR directly in the testis because the role of RAR is
thought to be in cell differentiation (25, 26, 27). A plausible mechanism
for germ cell apoptosis may involve direct stimulation by RXRs. The
RXR homodimer or the ligand for RXR, 9-cis RA, has been shown to
directly induce apoptosis in cells regardless of their state of
differentiation (28), and RXR and RXRß have been shown to be
expressed in testicular cells (29, 30); however, it is not known
whether RXRs are induced during testicular degeneration.
Not all germ cells were undergoing apoptosis during testicular degeneration in rats on the VAD diet for 6–9 weeks. Instead, pachytene spermatocytes and round spermatids were lost into the lumen of seminiferous tubules and accumulated in the epididymis. This is consistent with reports that in normal testis, spermatogonia and meiotic spermatocytes are the main cells that undergo apoptosis and apoptotic haploid germ cells are rarely detected (31, 32, 33, 34).
Interestingly, the pachytene spermatocytes and round spermatids
remaining in the testis during vitamin A depletion were expressing
RAR mRNA and protein. In fact, these same germ cell types that had
prematurely sloughed off and accumulated in the epididymis still
contained RAR transcripts and protein; however, the RAR protein
in these cells was no longer located in the nucleus but was seen
instead in the cytoplasm. The loss of RAR activity in the nucleus
may have induced the premature detachment of pachytene spermatocytes
and round spermatids, as RA and RAR have been implicated in the
maintenance of cell-cell adhesion and the deposition of extracellular
matrix proteins (35, 36, 37, 38).
In conclusion, the studies here demonstrate that rats depleted of
vitamin A have a decreased amount of RAR in early meiotic germ cells
or inactive RAR in advanced germ cells. This could explain why rats
depleted of vitamin A and the RAR knockout mice have a similar
testicular phenotype. The testicular degeneration that occurs during
the development of the VAD condition is due to apoptosis selective for
the early meiotic germ cells, and a loss of the advanced germ cells
into the lumen of seminiferous tubules and the epididymis.
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Acknowledgments |
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Footnotes |
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Received September 2, 1997.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Sci USA 90:7225–7229 gene expression in the rat testis: potential role during
the prophase of meiosis and in the transition from round to elongating
spermatids. Biol Reprod 56:549–556[Abstract]
expression in the rat
epididymis. Biol Reprod 54:1111–1119[Abstract]
are generated by alternative splicing
and differential induction by retinoic acid. EMBO J 10:59–69[Medline]
in acute promeylocytic
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by protein kinase A. Mol Endocrinol 9:860–871-dependent signaling pathway in the induction of tissue
transglutaminase and apoptosis by retinoids. J Biol Chem 270:6022–6029 isoforms from mouse testis. Biochem Biophys Res Commun 229:211–218[CrossRef][Medline]
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Z. Fu, T. Kubo, K. Sugahara, T. Noguchi, and H. Kato Cloning of Complementary Deoxyribonucleic Acids Encoding Quail (Coturnix coturnix japonica) Retinoic Acid Receptor {beta} Isoforms and Changes in Their Gene Expression During Gonadotropic Growth Biol Reprod, January 1, 2001; 64(1): 231 - 241. [Abstract] [Full Text]
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Z. Fu, H. Kato, K. Sugahara, and T. Kubo Retinoic Acid Accelerates the Development of Reproductive Organs and Egg Production in Japanese Quail (Coturnix coturnix japonica) Biol Reprod, December 1, 2000; 63(6): 1795 - 1800. [Abstract] [Full Text]
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 M. Komada, D. J. McLean, M. D. Griswold, L. D. Russell, and P. Soriano E-MAP-115, encoding a microtubule-associated protein, is a retinoic acid-inducible gene required for spermatogenesis Genes & Dev., June 1, 2000; 14(11): 1332 - 1342. [Abstract] [Full Text]
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 K. W. Braun, W. A. Tribley, M. D. Griswold, and K. H. Kim Follicle-stimulating Hormone Inhibits All-trans-retinoic Acid-induced Retinoic Acid Receptor alpha Nuclear Localization and Transcriptional Activation in Mouse Sertoli Cell Lines J. Biol. Chem., February 11, 2000; 275(6): 4145 - 4151. [Abstract] [Full Text] [PDF]
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B. Boulogne, C. Levacher, P. Durand, and R. Habert Retinoic Acid Receptors and Retinoid X Receptors in the Rat Testis During Fetal and Postnatal Development: Immunolocalization and Implication in the Control of the Number of Gonocytes Biol Reprod, December 1, 1999; 61(6): 1548 - 1557. [Abstract] [Full Text]
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J. M. Dufour and K. H. Kim Cellular and Subcellular Localization of Six Retinoid Receptors in Rat Testis During Postnatal Development: Identification of Potential Heterodimeric Receptors Biol Reprod, November 1, 1999; 61(5): 1300 - 1308. [Abstract] [Full Text]
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Y. Morita and J. L. Tilly Segregation of Retinoic Acid Effects on Fetal Ovarian Germ Cell Mitosis Versus Apoptosis by Requirement for New Macromolecular Synthesis Endocrinology, June 1, 1999; 140(6): 2696 - 2703. [Abstract] [Full Text]
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A. S. Cupp, J. M. Dufour, G. Kim, M. K. Skinner, and K. H. Kim Action of Retinoids on Embryonic and Early Postnatal Testis Development Endocrinology, May 1, 1999; 140(5): 2343 - 2352. [Abstract] [Full Text]
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