Spermatogenic cells as an alternative source of DNA for the production of bovine embryos by microinjection /
Microinsemination techniques have the potential to be applied for the propagation of many species for the treatment of certain forms of infertility, or in cases where AI or IVF are not feasible. In exotic and endangered species, sperm as well as oocytes are rare commodities, and intracytoplasmic inj...
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| Format: | Thesis Book |
| Language: | English |
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[Place of publication not identified] :
[publisher not identified] ;
2002.
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| Online Access: | http://proxy.library.tamu.edu/login?url=http://proquest.umi.com/pqdweb?did=726460281&sid=1&Fmt=2&clientId=2945&RQT=309&VName=PQD |
| Summary: | Microinsemination techniques have the potential to be applied for the propagation of many species for the treatment of certain forms of infertility, or in cases where AI or IVF are not feasible. In exotic and endangered species, sperm as well as oocytes are rare commodities, and intracytoplasmic injection of either sperm, or spermatogenic cells offers the most economical use of the available resources. Male gametes, such as round spermatids, ejaculated, epididymal and testicular spermatozoa, have been injected into the bovine oocyte using different approaches (conventional or Piezo methods) resulting in viable embryos. However, these techniques are still inefficient. The outcomes of cytoplasmic injections of ejaculated spermatozoa (ICSI), intact round spermatids (ROSI), round spermatid nuclei (ROSNI), testicular sperm heads (TSHI), and subzonal injection-fusion of intact round spermatids (ROSZI-EP) were compared. Bovine spermatogenic cells were recovered from abattoir testes and injected either "fresh-chilled" or frozen-thawed into bovine oocytes. Fertilization, cleavage, embryonic development rates and cell number at the blastocyst stage were compared among treatments. Two additional studies were performed. Three protocols for bovine spermatogenic cell cryopreservation were compared and short-term storage of testicular tissue and/or spermatogenic cell suspensions were analyzed. Cell viability following the different treatments was assessed using the trypan blue exclusion test. In initial studies very few blastocysts were produced by conventional injections using mature sperm and round spermatids. However, ionomycin-CHX as an exogenous activator enhanced blastocyst rates and was chosen for use in combination with the Piezo-injections in the second part of this study. Isolation of spermatogenic cells yielded over 74% cell viability after testicle storage for 24 h at 4°C was obtained. Survival rates were low (<38%) following cryopreservation of mixed spermatogenic cells suspension. When ROSNI, TSHI, ICSI, ROSZI-EP were applied using the Piezo injector, fertilization rates of over 31% were observed, cleavage rates ranged from 39-65%, while blastocysts rates ranged from 0-20%. These results demonstrate that "fresh-chilled" or frozen-thawed spermatogenic cells can be injected into the cytoplasm or into the PVS of the bovine oocyte for the production of bovine embryos. |
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| Item Description: | Vita. "Major Subject: Physiology of Reproduction". |
| Physical Description: | xv, 156 leaves : illustrations ; 28 cm. Issued also on microfiche from University Microfilm Inc. |
| Bibliography: | Includes bibliographical references (leaves 143-152). |