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Ovarian stimulation with highly purified human FSH and ultrasound-guided oocyte retrieval in Papio cynocephalus anubis
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Theriogenology OVARIAN STIMULATION WITH HIGHLY PURIFIED HUMAN FSH AND ULTRASOUND-GUIDED OOCYTE RETRIEVAL IN Papio cynoeephalus anubis
S. Cseh, 1 J. Corselfi, ~ P. Chan, 2 S. Nehlsen-Carmarella, 3 W. Kelln, 3 O. Fagoaga,3 L. Bailey4, A. SzalayI 1Center for Molecular Biology and Gene Therapy, Department of Microbiology and Molecular Genetics, 2Department of Obstetrics and Gynecology, 3Immunology Center, Department of Pathology, and 4Department of Surgery, School of Medicine, Loma Linda University, Loma Linda, CA, USA, 92350 Baboon gonadotropins are not available; thus, human preparations of varying purity have been used to induce controlled ovarian stimulation (COS). However, the baboon eventually produces antibody to the human hormone and neutralizes subsequent treatments. The objective of this study was to investigate the repeated use of a highly purified FSH (hpFSH) for COS in baboons. The preparation is considered >95% pure and contains negligible LH activity (FertinexTM,Serono, 1996). A modified human ovulation induction protocol was applied 3 times in a 12-month period to an adult baboon (13 years old, 17.5 kg) with a history of regular menstrual cycles (33-34 days). A long-acting GnRHa-contalning goserelin acetate (3.6 rag, ZoladexR, ICI Pharma) was implanted s.c. on cycle day 23 (luteal phase, P4 13.8 ng/ml). Serum estradiol (E2) and progesterone (P4) levels were obtained .by radioimmunoassay. Baboon anti-human FSH antibody formation was determined by ELISA. Menses occurred M0 days after GnRHa implantation. Daily hpFSH injections (75 IU, s.c.) were started ~ 10 days following menses. When the majority of follicles were >5 mm in diameter and the E2 levels reached their maximum, 2000 IU hCG was admires"tered i.m. to induce ovulation and oocyte maturation. Thirty to 34 hr after hCG administration, transabdominal follicular aspiration was performed using a variable frequency transvaginal transducer with ultrasound. Oocytes were assessed for meiotic maturity 3 hr after retrieval. CmRHa pituitary suppression was evident by low E2 levels (<_20 pg/ml) and absence of perineal turgeses in the days just prior to hpFSH injection. E2 levels began to rise ~0 7 days after beginning hpFSH treatment. Sonographic evidence of the follicular development was not observed trans-vaginally or -abdominally until treatment day ~ 10 when several follicles 3 mm in diameter and multiple smaller follicles became visible. The serum E2 reached plateau levels of 207 and 306 pg/ml and the size of the follicles increased to >5 mm in diameter in the first two treatment cycles between treatment Day 10 and 13. In the third treatment cycle however, the maximum E2 level was measured on Day 9 (101 pg/ml) and on Day 11 the E2 level dropped to baseline (<20pg/ml). Sera obtained after the first treatment cycle revealed no anti-human FSH IgM or IgG. However, while no antibodies were detectable during the second cycle; IgG was detected 1 day (tiler <16) and 6 days (titer 512) post-treatment; this decreased by 28 days (titer 128). Weak antibody (<16 titer) was detectable up until the third treatment was started; this disappeared during treatment, and rose on the last day of treatment (titer 16), and one day (tiler 32) and 29 days (titer 256) post-treatment. Fourteen and 21 morphologically normal oocytes, all at metaphase II and appropriate for in vitro insemination, were collected in the first and second treatments. No eggs could be collected in the third cycle. In conclusion, highly purified human FSH is effective to induce COS and oocyte maturation in the baboon suppressed with GnRHa. However, repeated superovulation of the same animal is limited by the development of neutralizing anti-human FSH antibody (IgG) and failed induction of follicular development.
Theriogenology OVARIAN STIMULATION WITH HIGHLY PURIFIED HUMAN FSH AND ULTRASOUND-GUIDED OOCYTE RETRIEVAL IN Papio cynoeephalus anubis
S. Cseh, 1 J. Corselfi, ~ P. Chan, 2 S. Nehlsen-Carmarella, 3 W. Kelln, 3 O. Fagoaga,3 L. Bailey4, A. SzalayI 1Center for Molecular Biology and Gene Therapy, Department of Microbiology and Molecular Genetics, 2Department of Obstetrics and Gynecology, 3Immunology Center, Department of Pathology, and 4Department of Surgery, School of Medicine, Loma Linda University, Loma Linda, CA, USA, 92350 Baboon gonadotropins are not available; thus, human preparations of varying purity have been used to induce controlled ovarian stimulation (COS). However, the baboon eventually produces antibody to the human hormone and neutralizes subsequent treatments. The objective of this study was to investigate the repeated use of a highly purified FSH (hpFSH) for COS in baboons. The preparation is considered >95% pure and contains negligible LH activity (FertinexTM,Serono, 1996). A modified human ovulation induction protocol was applied 3 times in a 12-month period to an adult baboon (13 years old, 17.5 kg) with a history of regular menstrual cycles (33-34 days). A long-acting GnRHa-contalning goserelin acetate (3.6 rag, ZoladexR, ICI Pharma) was implanted s.c. on cycle day 23 (luteal phase, P4 13.8 ng/ml). Serum estradiol (E2) and progesterone (P4) levels were obtained .by radioimmunoassay. Baboon anti-human FSH antibody formation was determined by ELISA. Menses occurred M0 days after GnRHa implantation. Daily hpFSH injections (75 IU, s.c.) were started ~ 10 days following menses. When the majority of follicles were >5 mm in diameter and the E2 levels reached their maximum, 2000 IU hCG was admires"tered i.m. to induce ovulation and oocyte maturation. Thirty to 34 hr after hCG administration, transabdominal follicular aspiration was performed using a variable frequency transvaginal transducer with ultrasound. Oocytes were assessed for meiotic maturity 3 hr after retrieval. CmRHa pituitary suppression was evident by low E2 levels (<_20 pg/ml) and absence of perineal turgeses in the days just prior to hpFSH injection. E2 levels began to rise ~0 7 days after beginning hpFSH treatment. Sonographic evidence of the follicular development was not observed trans-vaginally or -abdominally until treatment day ~ 10 when several follicles 3 mm in diameter and multiple smaller follicles became visible. The serum E2 reached plateau levels of 207 and 306 pg/ml and the size of the follicles increased to >5 mm in diameter in the first two treatment cycles between treatment Day 10 and 13. In the third treatment cycle however, the maximum E2 level was measured on Day 9 (101 pg/ml) and on Day 11 the E2 level dropped to baseline (<20pg/ml). Sera obtained after the first treatment cycle revealed no anti-human FSH IgM or IgG. However, while no antibodies were detectable during the second cycle; IgG was detected 1 day (tiler <16) and 6 days (titer 512) post-treatment; this decreased by 28 days (titer 128). Weak antibody (<16 titer) was detectable up until the third treatment was started; this disappeared during treatment, and rose on the last day of treatment (titer 16), and one day (tiler 32) and 29 days (titer 256) post-treatment. Fourteen and 21 morphologically normal oocytes, all at metaphase II and appropriate for in vitro insemination, were collected in the first and second treatments. No eggs could be collected in the third cycle. In conclusion, highly purified human FSH is effective to induce COS and oocyte maturation in the baboon suppressed with GnRHa. However, repeated superovulation of the same animal is limited by the development of neutralizing anti-human FSH antibody (IgG) and failed induction of follicular development.