Synchronization of porcine fetal fibroblast cells with topoisomerase-inhibitor hoechst 33342

Animal Reproduction Science
B Kühholzer, R S Prather

Abstract

Proper synchronization of donor nuclei has been shown to have a major influence on the developmental potential of nuclear transfer embryos. In the present study, a protocol was established to synchronize porcine fetal fibroblasts in the G2 stage of the cell cycle. Cell cycle analyses were performed by flow cytometry. Cells were pre-synchronized by serum deprivation or aphidicoline-treatment; then incubated in medium containing 0.1 microg/ml Hoechst 33342 (H342). The fluorochrome H342 has been shown to be a topoisomerase-inhibitor that can inhibit progression through the cell cycle. There was no significant difference in the percentage of fibroblasts in G2/M whether cells were pre-synchronized in medium supplemented with 0.1% serum for 48h or 0.5% serum for 6 days. Compared with controls, pre-synchronization in early S-phase before incubation in H342 increased the proportion of G2/M fibroblasts; also an increase from 0 and 6 versus 12h culture in complete medium before incubation in H342 resulted in an increased percentage of cells in G2/M at the end of the synchronization period (9.3 and 13.1% versus 33.7%; P<0.001). Neither an increase in the concentration of H342 (0.1-1.0 microg/ml) nor a longer exposure time (12h versus 18h ...Continue Reading

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Citations

Jun 24, 2003·Il Farmaco·Sabiha AlperIsmail Yalçin
May 15, 2002·Cloning and Stem Cells·András DinnyésIan Wilmut
Sep 13, 2006·Environmental Health Perspectives·Samuel C McNeelyJ Christopher States
Sep 30, 2008·Journal of Dentistry·Ryo NishikioriMasayuki Okazaki
Nov 3, 2007·Reproduction in Domestic Animals = Zuchthygiene·M D GoissisJ A Visintin
Oct 1, 2009·Reproduction in Domestic Animals = Zuchthygiene·C H ChorescaS C Park
Jun 1, 2002·Molecular Endocrinology·G Alexander AbelTheo Rein

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