• Development and Reproduction
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• v.11 no.1
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• pp.49-54
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• 2007

Ethane 1,2-dimethane sulfonate(EDS), a toxin which specifically kills Leydig cells(LC), has been widely used to prepare the reversible testosterone(T) depletion rat model. In the present study, we monitored the gene expression profiles of pituitary gonadotropins, LH and FSH, up to 7 weeks after EDS injection. Adult male Sprague-Dawley rats($300{\sim}350\;g$ B.W.) were injected with a single dose of EDS(75 mg/kg i.p.) and sacrificed on weeks 0, 1, 2, 3, 4, 5, 6 and 7. Total RNAs were purified from each pituitary, and the message levels of common alpha subunit($C{\alpha}$) of pituitary glycoprotein hormones, LH beta subunit($LH{\beta}$), FSH beta subunit($FSH{\beta}$) and GnRH receptor(GnRH-R) were evaluated by semi-quantitative RT-PCRs. The message levels of $C{\alpha}$ increased sharply during weeks 1-4, then return to the control level on week 5. The mRNA levels of $LH{\beta}$ were elevated after week 2, reached the peak at week 4, then declined to the control level after week 5. The message levels of $FSH{\beta}$ were elevated after week 2, reached the peak at week 3, then declined to the nadir at week 5. Similarly, the mRNA levels of GnRH-R were elevated after week 2, reached the peak at week 3, then gradually declined to the control level after week 5. The present study indicated that EDS treatment could induce reversible alterations in the transcriptional activities of gonadotropin subunits and GnRH-R in the anterior pituitary from male rats. EDS injection model might be useful to understand the mechanism of hormonal regulation of hypothalamus- pituitary neuroendocrine axis in male rats.

• Asian-Australasian Journal of Animal Sciences
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• v.26 no.4
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• pp.488-500
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• 2013

The objective of this study was to compare the effectiveness of the protocols for superstimulation of follicular growth in Thai native heifers. Heifers (n = 20) were randomly divided into four groups of five heifers/group. Heifers were given a single dose by i.m. administration of 100 mg Follicle Stimulating Hormone dissolved in polyvinylpyrrolidone (FSHp) at 24 h. Ovum pick up (OPU) occurred at 72 h ($F_{24}O_{72}$ protocol; Group 1) or 96 h ($F_{24}O_{96}$ protocol; Group 2), and at 36 h and OPU at 72 h ($F_{36}O_{72}$ protocol; Group 3) or 96 h ($F_{36}O_{96}$ protocol; Group 4) after follicular ablation. The dynamics of ovarian follicular growth were monitored by twice-daily ultrasonographic examinations. Blood sample collections were performed every 12 h after initiation of treatment for assessment of FSH, E2 and P4 profiles. All heifers were subjected to eight repeated sequential sessions of OPU. The follicular deviation commenced $24{\pm}5.32$ h after follicular ablation in all groups. The circulatory FSH surged quickly from 24 to 36 h (>0.8 ng/ml) after follicular ablation and circulatory estrogen levels steadily increased from 36 h until OPU in all groups. At the end of the OPU sessions, the mean number of aspirated follicles/heifer/session in $F_{36}O_{72}$ protocol (Group 3) and $F_{36}O_{96}$ protocol (Group 4) were higher than in the two other groups (p<0.05). The number of cumulus-oocyte complexes (COCs), cleaved and day 8 blastocysts rates in the $F_{36}O_{72}$ protocol (Group 3) were higher than in the other groups (p<0.05). It can be concluded that a single dose i.m. administration of 100 mg FSHp at 36 h and OPU at 72 h after follicular ablation ($F_{36}O_{72}$ protocol; Group 3) was the most effective protocol for superstimulation of follicular growth for repeated OPU and subsequent in vitro embryo production in Thai native heifers.

• Clinical and Experimental Reproductive Medicine
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• v.39 no.1
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• pp.22-27
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• 2012

Objective: To evaluate the effectiveness of GnRH antagonist multiple dose protocol applied during early and late follicular phase (MDP-EL) in comparison with standard GnRH agonist luteal long protocol (LP) in each non-obese and obese polycystic ovary syndrome (PCOS) women undergoing IVF. Methods: Two hundred eleven infertile women with PCOS were recruited and randomized to undergo either GnRH antagonist MDP-EL (antagonist group) or standard GnRH agonist luteal LP (agonist group). IVF cycle outcomes were compared between the two groups. Results: Total dose and days of recombinant human follicle stimulating hormone (rhFSH) administered were significantly fewer in the antagonist group than in the agonist group. Incidence of severe ovarian hyperstimulation syndrome was significantly lower in the antagonist group. However, IVF and pregnancy outcomes were similar in the two groups. When all subjects were divided into non-obese and obese subgroups, in non-obese PCOS subgroup, IVF and pregnancy outcomes were comparable in the antagonist and agonist groups but total dose and days of rhFSH were also significantly fewer in the antagonist group. Similar findings were also observed in obese PCOS subgroup. Conclusion: GnRH antagonist MDP-EL is at least as effective as GnRH agonist LP and may be a more patient-friendly alternative in controlled ovarian stimulation for PCOS patients undergoing IVF, independent of body mass index.

• Clinical and Experimental Reproductive Medicine
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• v.40 no.2
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• pp.83-89
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• 2013

Objective: To investigate outcomes of stimulated IVF cycles in which GnRH antagonist was omitted on the ovulation triggering day. Methods: A total of 86 women who underwent controlled ovarian hyperstimulation with recombinant FSH and GnRH antagonist flexible multiple-dose protocols were recruited and prospectively randomized into the conventional group (group A) or cessation group (group B). The GnRH antagonist, 0.25 mg/day of cetrorelix, was started when the leading follicle reached 14 mm in diameter and was continuously administered until the hCG triggering day (group A, 43 cycles) or until the day before hCG administration (group B, 43 cycles). The maturity of oocytes, fertilization rate, embryo quality, and implantation and clinical pregnancy rates were evaluated. Results: The duration of ovarian stimulation, total dose of gonadotropins, serum estradiol levels on hCG administration day, and number of oocytes retrieved were not significantly different between the two groups. The total dose of GnRH antagonist was significantly lower in group B than group A ($2.5{\pm}0.9$ vs. $3.2{\pm}0.8$ ampoules, p<0.05). There was no premature luteinization in any of the subjects. The proportion of mature oocytes and fertilization rate were not significantly different in group B than group A (70.7% vs. 66.7%; 71.1% vs. 66.4%, respectively). There were no significant differences in the implantation or clinical pregnancy rates. Conclusion: Our prospective randomized study suggested that cessation of GnRH antagonist on the hCG administration day during a flexible multiple-dose protocol could reduce the total dose of GnRH antagonist without compromising its effects on pregnancy rates.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.4
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• pp.225-234
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• 2004

Objectives: To evaluate the efficacy of GnRH antagonist cetrorelix in women undergoing controlled ovarian hyperstimulation (COH) for in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) and to determine changes in serum hormone concentrations during cetrorelix administration. Methods: We performed a clinical trial on 30 patients undergoing COH with highly purified follicular stimulating hormone (HP-FSH) and gonadotropin releasing hormone antagonist (GnRHant), cetrorelix. FSH was administrated from day 2 or 3 of cycle with fixed dose and adjusted according to individual response. 0.25 mg of cetrorelix was injected daily subcutaneously from stimulation day 5 until the day of hCG administration. Daily ultrasound monitoring was performed for growing follicles and serum levels of luteinizing hormone (LH), estradiol ($E_2$) and progesterone were measured daily during cetrorelix administration. Up to 4 embryos were transferred. Results: Mean age of enrolled patients was $32.0{\pm}3.4$ years (mean $\pm$ S.D.). All of 30 patients underwent oocyte pick-up, and embryo transfer was done in 28 patients. The total and mean numbers of received oocytes were 196 and $6.5{\pm}4.7$, the number of fertilized eggs was 111, and the fertilization rate was 56.6%. Total duration of FSH administration was $9.2{\pm}2.2$ days and mean of $24.3{\pm}7.7$ ampules of HP-FSH was administered. Total duration of cetrorelix administration was $5.7{\pm}1.9$ days. Serum LH and progesterone levels were maintained in the range of $1.4{\sim}2.9\;mIU/mL$ and $0.3{\sim}0.6\;ng/mL$, which respectively reflected effective prevention of premature LH surge. Clinical pregnancies were achieved in 9 patients, and overall clinical pregnancy rate was 30.0% per oocyte retrieval, and 32.1% per embryo transfer. Conclusion: GnRH antagonist is safe and convenient for COH for IVF-ET and effective with optimal pregnancy rate.

• Journal of Embryo Transfer
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• v.20 no.3
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• pp.311-315
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• 2005

This study was conducted to investigate the effects of disregarding of heat detection on embryo production in superovulation of Hanwoo cows. Donors which showed 1 or 2 times of normal heat and had no abnormality of reproductive tract were selected The superovulation was performed injection of 2.5 mL FSH (Antorin R-10, Japan) 2 times on 5 days before next heat and continuously with the reduction of dose to 0.5 mL of first injection 2 times in a day for next 3 days. Otherwise, the donors of CIDR group were inserted CIDR plus (with the capsule of estradiol benzoate 10 mg) on Day 10 from standing heat for 9 days. On 6 days from insertion of CIDR, FSH was injected above same manners. The response according to the natural heat and CIDR were $82.2\%,\;89.7\%$, respectively. There were no difference between both treatments. The recovery rates of embryos were 7.7, 10.5, respectively and transferable embryos were 3.4, 3.8. There showed significant difference between both treatments (p<0.05). These results suggested that disregarding of heat detection in superovulation could be produced transferable embryos for embryo transfer and preserve the donors from the excess hormonal administration and maintain the economical lift span of genetically available Hanwoo donors.

• Development and Reproduction
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• v.13 no.4
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• pp.329-339
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• 2009

Cortisol is present in high concentration in the ovary and its receptor is expressed in the ovarian cells. Moreover, cortisol is known to have a role in steroid synthesis and cell metabolism in human granulosa and lutein cells. However, little is known of the role of cortisol presenting in high concentration in the follicles after LH surge on the granulosa-lutein cells. Therefore, the this study we evaluated the apoptosis and the production of progesterone $(P_4)$ and estradiol $(E_2)$ in the granulosa-lutein cells that are obtained during oocyte-retrieval after treatment with 5, 50, and $500{\mu}g/m\ell$ cortisol and 1 IU/$m\ell$ FSH. Results of DNA fragment analysis and TUNEL assay demonstrated that DNA fragmentation and the rate of apoptotic cells were increased in a dose-dependent manner showing a significant increase in 50 and $500{\mu}g/m\ell$ cortisol treated cells. We found, however, that FSH did not suppress the apoptosis of the cells induced by cortisol. In the results of chemiluminescence assay for $P_4$ and $E_2$, $P_4$ production was decreased by cortisol treatment, whereas $E_2$ was not changed. We also demonstrated that FSH did not inhibit the suppressive effect of GnRH on $P_4$ production as the result of apoptosis. The present study suggests that cortisol of high concentration could cause the apoptosis of human granulosa-lutein cells by suppressing the production of $P_4$. However, we need more studies to elucidate the mechanism by which cortisol induces apoptosis in human granulosa-lutein cells in view of the fact that our results are inconsistent with previous reported data.

• Journal of Embryo Transfer
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• v.27 no.3
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• pp.127-131
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• 2012

This study was performed to investigate the effects of parity and season on the embryo production in superovulated Hanwoo cows. Superovulation was performed from 1 to 8 times by repeated superovulation treatment of Hanwoo cows (n = 22). Irrespective of estrous cycle, donor cows were received a CIDR, progesterone (50 mg) and estradiol benzoate (2.5 mg). After 4.5 days, the donor cows were superovulated with total 28AU FSH (Antorin R-10) administrated twice daily in a decreasing dose for 4 days. On $6^{th}$ and $7^{th}$ of FSH injection, 2.5 mg and 15 mg $PGF_2{\alpha}$ were injected i.m, respectively. CIDR was removed at the $7^{th}$ FSH injection. The donor cows received $200{\mu}g$GnRH at 48 hrs after $1^{st}$ $PGF_2{\alpha}$ injection. The donor cows were artificially inseminated three times after estrous detection at 12 hr intervals and embryos were recovered 7 days after estrous detection. The mean number of total ova, transferrable embryos, degenerated embryos and unfertilized oocytes were $11.6{\pm}7.9$, $5.5{\pm}4.4$, $3.0{\pm}3.3$ and $2.6{\pm}4.1$ per donor cows, respectively. A higher number of total ova were recovered in parity 3~5 ($14.3{\pm}1.3$) than 1~2 ($8.9{\pm}1.9$, P<0.05). The number of recovered normal embryos is significantly higher in parity 3~5 ($7.3{\pm}0.8$) than that of over 6 ($3.7{\pm}1.5$). Significantly higher number of total ova and normal embryos were recovered in summer ($16.4{\pm}2.3$, $8.1{\pm}1.4$) than in autumn ($10.1{\pm}1.8$, $4.5{\pm}1.1$) and winter ($6.3{\pm}1.8$, $3.3{\pm}1.1$), respectively (P<0.05). Transferable embryos were significantly higher in summer ($7.6{\pm}1.3$) than in winter ($3.0{\pm}1.0$, P< 0.05). The results were showed that parity and season affecting on the production of embryos in superovulated Hanwoo.

• The Korean Journal of Physiology and Pharmacology
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• v.27 no.2
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• pp.167-176
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• 2023

This study aims to explore the impact of Rehmannioside D (RD) on ovarian functions of rats with diminished ovarian reserve (DOR) and its underlying mechanisms of action. A single injection of cyclophosphamide was performed to establish a DOR rat model, and fourteen days after the injection, the rats were intragastrically administrated with RD for two weeks. Rat estrus cycles were tested using vaginal smears. Ovarian tissues were histologically evaluated, the number of primordial, mature, and atretic follicles was calculated, and the apoptotic rate of granulosa cells. Follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E₂) levels were determined by ELISA assays. Protein levels of Forkhead Box O1 (FOXO1), KLOTHO, Bcl-2, and Bax were investigated in ovarian tissues of DOR rats. The binding between FOXO1 and KLOTHO was verified by ChIP assay. High-dose administration of RD into DOR rats improved their estrus cycles, increased ovarian index, enhanced the number of primordial and mature follicles, reduced the number of atretic follicle number, and ovarian granulosa cell apoptosis in addition to inhibiting FSH and LH levels and upregulating E₂ expression. FOXO1 and KLOTHO were significantly suppressed in DOR rats. FOXO1 knockdown partially suppressed the protective effects of RD on DOR rats, and KLOTHO overexpression could restore RD-induced blockade of DOR development despite knocking down FOXO1. FOXO1 antibody enriched KLOTHO promoter, and the binding between them was reduced in DOR group compared to that in sham group. RD improved ovarian functions in DOR rats and diminished granulosa cell apoptosis via the FOXO1/KLOTHO axis.

• Fisheries and Aquatic Sciences
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• v.14 no.4
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• pp.379-388
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• 2011

The mechanism by which gonadotropin-releasing hormone (GnRH) and dopamine (DA) control gonadotropin (GTH) release was studied in male and female rainbow trout using cultured pituitary cells obtained at different reproductive stages. The mechanisms of follicle-stimulating hormone (FSH) release by GnRH and DA could not be determined yet. However, basal and salmon-type GnRH (sGnRH)- or chicken-II-type GnRH (cGnRH-II)- induced luteinizing hormone (LH) release increased with gonadal maturation in both sexes. LH release activity was higher after sGnRH stimulation than cGnRH-II stimulation at maturing stages in both sexes. The GnRH antagonist ([Ac-3, 4-dehydro-$Pro^1$, D-p-F-$Phe^2$, D-$Trp^{3,6}$] GnRH) suppressed LH release by sGnRH stimulation in a dose-dependent manner, although the effect was weak in maturing fish. The role of DA as a GTH-release inhibitory factor differs during the reproductive cycle: the inhibition of sGnRH-stimulated LH release by DA was stronger in immature fish than in maturing, ovulating, or spermiated fish. DA did not completely inhibit sGnRH-stimulated LH release, and DA alone did not alter basal LH release. Relatively high doses ($10^{-6}$ or $10^{-5}M$) of domperidone (DOM, a DA D2 antagonist) increased LH release, which did not change with reproductive stage in either sex. The potency of DOM to enhance sGnRH-stimulated LH release was higher in maturing and ovulated fish than in immature fish. These data suggest that LH release from the pituitary gland is controlled by dual neuroendocrine mechanisms by GnRH and DA in rainbow trout, as has been reported in other teleosts. The mechanism of control of FSH release, however, remains unknown.