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Title: ULTRASTRUCTURE AND KARYOTYPE EXAMINATION OF PREIMPLANATION ELONGATION STAGE NUCLEAR TRANSFER BOVINE BLASTOCYSTS DEVELOPED IN THE SHEEP UTERUS.

Author
item Talbot, Neil
item Powell, Anne
item GARRETT, WESLEY - 1265-45-00
item EDWARDS, JANICE - 1265-45-00
item REXROAD, JR., CAIRD - 1265-45-00

Submitted to: Tissue and Cell
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/7/1999
Publication Date: N/A
Citation: N/A

Interpretive Summary: The study examined whether development of bovine nuclear cloned embryos in the sheep uterus resulted in normal morphology and chromosome content. Noteworthy ultrastructural features held in common to those embryos grown in the sheep uterus were the presence of retroviral-like particles elalorated from the embryonic disc cells, the presence of monocilia on embryonic disc cells, distinct alternative mitochondrial morphology betwee epiblast cells and trophectoderm cells, and the presence of crystalloid bodies in the trophectoderm cells. In comparison to development in the cow uterus, sheep uterine incubation caused increased cell death of the epiblast cells. Chromosomal analysis of nine nuclear cloned bovine embryos developed in the sheep uterus showed no abnormal chromosome complements. These results indicated that using the sheep uterus for the growth of nuclear cloned embryos could be useful for the assessment of their early development and that our nuclear cloning protocol resulted in embryos with normal chromosome complements.

Technical Abstract: The study examined whether development of bovine nuclear transfer (NT) blastocysts in the sheep uterus resulted in morphologically and karyotypically normal elongation stage bovine blastocysts. Seven-day in vitro-produced (IVP) NT, parthenogenic, of fertilized bovine blastocysts were surgically transferred into sheep uteri. Sheep were sacrificed after 7-8 d, and blastocysts were flushed from their uteri. One of each kind of IVP bovine blastocyst was recovered from sheep uteri for analysis by transmission electron microscopy (TEM), and nine NT blastocysts were used to establish cell cultures that were analyzed for chromosome complement. TEM analysis of in vivo-derived elongation stage bovine and ovine blastocysts were done for comparative purposes. Most ultrastructural features of the 13 d - 19 d blastocysts were similar to earlier stage blastocysts except that distinct alternative mitochondrial morphologies were found between epiblast and trophectoderms cells. Also, singly occurring cilium were observed in bovine epiblast and endoderm cells, and retrovirus-like particles were elaborated by these same cell types. Development in the sheep uterus of IVP bovine blastocysts resulted in the presence of crystalloid bodies in the trophectoderm cells, and apoptotic and necrotic cells were observed in the epiblast tissue. Thus, in vivo incubation in the sheep uterus allowed nearly normal development to the elongated blastocyst stage and could be useful for assessment of NT bovine blastocyst developmental competence. Cell cultures derived from from the NT blastocysts had normal chromosome complements suggesting that activation by ionomycin and 6-dimethyl-aminopurine (DMAP) did not cause detrimental changes in ploidy in those blastocysts that developed.