Faculty of Science
School of Anatomical Sciences
9901061h
karrenthomson@yahoo.co.uk / Clomiphene citrate (CC), a synthetic estrogen, is an efficient superovulator used in
infertility treatment. However pregnancy rates resulting from CC treatment are
low. Research has suggested that this may be due to an aberrant effect on
implantation; CC binds to estrogen receptors (ER) and may affect estrogen
responsive gene expression and thus implantation. This study investigates the
effect of CC on ERa, 90kDa heat shock protein (Hsp90) and Hoxa10 expression in
the rat uterus. Hsp90 binds to ERa in the absence of ligand and is involved in
inducing a high affinity ligand binding conformation in the ER and in
transactivation of the ER. Hoxa10 has been shown to be essential for uterine
receptivity to implantation. CC (0.25mg) was given to ovariectomized rats, either
alone or prior to a hormonal regime known to induce uterine receptivity for
implantation. Expression of ERa, Hsp90 and Hoxa10 was determined by Western
blotting, fluorescence immunocytochemistry and reverse transcription polymerase
chain reaction. The single dose CC treated rats were compared to the controls as
well as to ovariectomized rats treated with 0.5mg 17b estradiol (E2). The CC treated
pseudopregnant rats (CCPPPE treated) were compared to 5½ day pregnant and
pseudopregnant rats without CC (PPPE treated), to determine CCs effect at
implantation. E2 upregulated ERa and Hsp90 expression in the rat uterus
compared to controls (p<0.05). The finding for ERa was unexpected as other
studies have shown that E2 decreases ERa levels a few hours after administration
in the uterus. The present study therefore suggests a biphasic effect of E2 on ERa
expression in the rat uterus. The effect of E2 on Hsp90 and ERa also proposes a
balance between the levels of these two proteins in the uterus, to keep ERa in its
optimal state and suggests that too high and too low a concentration of Hsp90 may
both be inhibitory to ERa functioning. No significant difference was found in
ERa and Hsp90 expression between the non-receptive (vehicle treated) and the
receptive (PPPE treated) rat uteri, suggesting that these two genes are not markers
for receptivity. However E2 is known to induce implantation of donor blastocysts
in progesterone (P4) primed uteri. Therefore it is still essential for ERa to be
present at implantation. It is of interest that CC downregulated ERa levels both in
ii
the absence of ovarian hormones and at implantation in the rat uterus. It is
therefore proposed that this antiestrogenic effect would render the uterus less
sensitive to the E2 required to induce implantation, thus accounting for low
pregnancy rates with CC use. Although CC did not alter the expression of Hsp90
in this study, the reduction in ERa levels in response to CC may also upset the
balance in the expression of these two genes, which may affect the transcriptional
activity of ERa, and further prevent implantation. No clear results were obtained for
Hoxa10 expression with the Western blots. However based on the ICC results, CC
did not appear to affect Hoxa10 expression. Since P4 and not E2 is known to have
the predominant effect on Hoxa10 expression, it is likely that E2 analogs, such as
CC, would also not affect Hoxa10 expression to a significant degree. Future work
will aim to separate the different uterine compartments and to determine the effects
of CC on the expression of other implantation specific genes in the uterus.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:wits/oai:wiredspace.wits.ac.za:10539/1455 |
| Date | 26 October 2006 |
| Creators | Thomson, Karren Judith |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 204538 bytes, 167382 bytes, 261483 bytes, 2766105 bytes, 777721 bytes, 204434 bytes, 153606 bytes, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf, application/pdf |
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