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Regular Article |
a Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, Massachusetts 01003
b Hematech LLC, Manhattan, Kansas 66502
c Hematech LLC, Worcester, Massachusetts 01605
A study was undertaken to determine whether injection of porcine sperm factors (pSF), which trigger oscillations in intracellular calcium concentration ([Ca2+]i) in mammalian oocytes, could be used to activate bovine oocytes during nuclear transfer. To date, only combined treatments that induce a monotonic rise in [Ca2+]i and inhibit either phosphorylation or protein synthesis have been utilized in nuclear transfer. Several doses of pSF were tested. Injection of 5 mg/ml pSF triggered [Ca2+]i oscillations that resembled those associated with fertilization with respect to amplitude and periodicity, and as a result, a high percentage of oocytes underwent activation. Furthermore, this concentration of pSF supported in vitro and in vivo development up to 60–90 days of gestation, comparable to development in control nuclear transfer embryos. Nevertheless, neither activation procedure supported development as well as did fertilization. The effectiveness of pSF as an activating agent in bovine oocytes may have been compromised because pSF was unable to support oscillations past 3–5 h postinjection and a second injection was necessary to extend the [Ca2+]i oscillations. Likewise, a single injection of pSF failed to trigger downregulation of the inositol 1,4,5-trisphosphate receptor 1 subtype, whereas a second injection downregulated the receptor in a manner similar to that seen in fertilized oocytes. These results demonstrate that soluble factor(s) from porcine sperm can support early development in bovine nuclear transfer embryos; however, the efficacy may be limited because of the premature cessation of the induced oscillations.
1 This work was supported by USDA grants (97-2919, 99-2371) to R.A.F.
2 Correspondence: James M. Robl, Hematech LLC, 4 Biotech Park, 377 Plantation Dr., Worcester, MA 01605. FAX: 508 792 0782; jrobl{at}hematech.com
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