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  • Title: Neutralization of tumor necrosis factor alpha (TNF alpha) action on cell proliferation in rat blastocysts by antisense oligodeoxyribonucleotides directed against TNF alpha p60 receptor.
    Author: Pampfer S, Vanderheyden I, Vesela J, De Hertogh R.
    Journal: Biol Reprod; 1995 Jun; 52(6):1316-26. PubMed ID: 7543296.
    Abstract:
    Antisense oligodeoxyribonucleotide inhibition of gene expression was used to test whether the p60 form of the tumor necrosis factor alpha (TNF alpha) receptor (Rp60) is responsible for mediating the negative effect of TNF alpha on the development of rat blastocysts in vitro. The antisense oligonucleotide was designed to overlap the translation initiation codon of the TNF alpha Rp60 mRNA. Preliminary experiments showed that concentrations of oligonucleotides above 10 microM in the culture medium were embryotoxic over 24 h. When used at nontoxic concentrations (8 microM), antisense oligonucleotides specifically decreased the abundance of intact TNF alpha Rp60 transcripts by 80% within 3 h of exposure. In contrast to results with control embryos, mRNA for the second form of TNF alpha receptor, TNF alpha Rp80, was detected in blastocysts exposed to antisense oligonucleotides to TNF alpha Rp60. Antisense oligonucleotides to TNF alpha Rp60 blocked the 25-30% decrease in cell proliferation induced by 50 ng/ml TNF alpha added to a standard culture medium and by 5 ng/ml TNF alpha added to a medium that had been conditioned by rat uterine cells. Sense oligonucleotides had no such protective effect. Because uterine cells from diabetic rats secrete higher levels of TNF alpha than those from control rats, antisense oligonucleotides were also tested in a medium that had been conditioned by diabetic uterine cells (cell-secreted TNF alpha concentration was 50 pg/ml in this medium, and no exogenous TNF alpha was added). Addition of antisense oligonucleotides to TNF alpha Rp60 improved the quality of this medium with respect to cell proliferation but failed to correct the high frequency of dead cells observed in the blastocysts.
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