title: Human menopausal and pregnant mare serum gonadotrophins in murine superovulation regimens for transgenic applications.
abstract: Superovulation is a fundamental procedure for generating transgenic rodents. While various methods exist, zygote yield/quality remain suboptimal, making these techniques open to refinement. All require a follicle stimulating and a luteinising effect. The former can be induced by pregnant mare serum gonadotrophin (PMSG) or other compounds like human menopausal gonadotrophin (HMG). While HMG can double zygote yield compared to PMSG, no study has compared their effects on embryo quality. Embryo yield could also be increased with PMSG: timing administration at estrus may further improve follicular recruitment. This study compared: (i) the efficacy of HMG/PMSG for producing viable embryos for microinjection; and (ii) the effect of HMG/PMSG administration at estrus on embryo yield. Whitten effect-induced estrous C57/Bl6xCBA F(1) hybrid mice were superovulated as follows: PMSG (day 1; 5 IU intraperitoneally) or HMG (days 1 and 2; 1 IU intramuscularly); all received human chorionic gonadotrophin (hCG) on day 3 (5 IU, intraperitoneally). Zygotes were retrieved following mating, morphologically assessed and microinjected with innocuous ZhAT1R construct (expressing LacZ reporter and human angiotensin II type 1 receptor) before transfer to pseudopregnant recipients. Pups were tested for the transgene by Southern blot. Neither HMG nor PMSG proved superior in improving embryo yield, morphology and short-term post-microinjection survival. However, HMG group micromanipulated embryos all failed to establish a pregnancy/generate transgenic pups, unlike their PMSG counterparts. While HMG can be used for superovulation, it appears to increase embryo vulnerability to the long-term effects of microinjection. Furthermore, the embryo yields associated with HMG can be replicated by timing PMSG injection to coincide with Whitten effect-induced estrus.
authors.affiliation: Molecular Medicine--Level 9, Wellcome Trust Brenner Building, St. James's University Hospital, Beckett Street, Leeds LS9 7TF, UK. d.a.brooke@leeds.ac.uk
authors.forename: D A
authors.initials: DA
authors.lastname: Brooke
authors.forename: N M
authors.initials: NM
authors.lastname: Orsi
authors.forename: J F X
authors.initials: JF
authors.lastname: Ainscough
authors.forename: S E
authors.initials: SE
authors.lastname: Holwell
authors.forename: A F
authors.initials: AF
authors.lastname: Markham
authors.forename: P L
authors.initials: PL
authors.lastname: Coletta
mesh_terms: D000818:Animals; D015139:Blotting, Southern; D004624:Embryo Transfer / Q000662:veterinary; D005260:Female; D018014:Gene Transfer Techniques / Q000662:veterinary*; D006064:Gonadotropins, Equine / Q000008:administration & dosage*; D006801:Humans; D008596:Menotropins / Q000008:administration & dosage*; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008822:Mice, Transgenic; D008845:Microinjections / Q000662:veterinary; D011247:Pregnancy; D011555:Pseudopregnancy / Q000662:veterinary; D044140:Receptor, Angiotensin, Type 1 / Q000235:genetics; D013480:Superovulation; D020858:Tissue and Organ Harvesting / Q000662:veterinary; D019076:Transgenes; D015053:Zygote
chemical_list: D006064:Gonadotropins, Equine; D044140:Receptor, Angiotensin, Type 1; D008596:Menotropins