Serum-Starved Apoptotic Fibroblasts Reduce Blastocyst Production but Enable Development to Term after SCNT in Cattle

Miranda, M. S.; Bressan, Fabiana Fernandes; Zecchin, Karina G.; Vercesi, Aníbal E.; Mesquita, Lígia Garcia; Merighe, Giovana Krempel Fonseca; King, W. A.; Ohashi, Otávio Mitio; Pimentel, José Rodrigo Valim; Perecin, Felipe; Meirelles, Flávio Vieira

doi:10.1089/clo.2009.0028

Cited by 30 publications

(15 citation statements)

References 48 publications

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“…This may be attributed to the induction of apoptosis, which affected the development of cloned embryos. Miranda et al (2009) showed that blastocyst production and quality after SCNT were reduced using apoptotic cells induced by serum starvation. Park et al (2004) found that treatment of donor cells with putative apoptosis inhibitors reduced apoptosis after SCNT and improved the development of cloned embryos.…”

Section: Discussionmentioning

confidence: 99%

Production of hGFAP-DsRed transgenic Guangxi Bama mini-pigs via somatic cell nuclear transfer

Lü

Yang

et al. 2015

Genet. Mol. Res.

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ABSTRACT. The mini-pig is a useful animal model for human biomedical research due to its physiological similarity to humans and the ease of handling. In order to optimize the efficiency of production of transgenic Bama mini-pigs through somatic cell nuclear transfer (SCNT), we examined the effects of contact inhibition, roscovitine treatment, and serum starvation on the cell cycle synchronization and transgenic cloned embryo development in vivo and in vitro after nuclear transfer. The analysis showed that the rates of G0/G1 stage cells in the contact inhibition (92.11%) and roscovitine treatment groups (89.59%) were significantly higher than in serum starvation group (80.82%). A higher rate of apoptosis was seen in the serum starvation group (14.13%) compared to the contact inhibition and roscovitine treatment groups (6.71 and 2.46% respectively, P < 0.05). There was a significant decrease in blastocyst yield in the serum starvation group (14.19%) compared to the roscovitine treatment and contact inhibition groups (21.31 and 20.32% respectively, P < 0.05). A total of 1070 transgenic cloned embryos derived from the three treatment groups were 16286 L.L. Hu et al.©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 14 (4): 16285-16296 (2015) transferred to surrogate sows; one pregnancy was established and three embryos from the roscovitine treatment group successfully completed gestation. These results indicate that the roscovitine treatment was more effective at synchronizing transgenic kidney cells in Bama mini-pigs and allowed reconstructed embryos to develop to full term.

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Section: Discussionmentioning

confidence: 99%

Production of hGFAP-DsRed transgenic Guangxi Bama mini-pigs via somatic cell nuclear transfer

Lü

Yang

et al. 2015

Genet. Mol. Res.

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“…Since all doses resulted in significantly higher proliferation/viability according to the MTT assay, 2 mM of VPA was used in the subsequent experiments as this concentration has been shown to increase the rates of induced pluripotent stem cells (iPSCs) [21] production and SCNT in mice [14]. It is noteworthy that based on the MTT assay, the treatment with VPA increased cell proliferation/viability while these cells were not expected to proliferate because of serum starvation to synchronize the cell cycle [46], [47]. However, this increase might be caused as a side effect of VPA on cell metabolism as the MTT assay is based on activity of metabolic enzymes that reduces tetrazolium salts [30].…”

Section: Discussionmentioning

confidence: 99%

Development to Term of Cloned Cattle Derived from Donor Cells Treated with Valproic Acid

Sangalli

Chiaratti

et al. 2014

PLoS ONE

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Cloning of mammals by somatic cell nuclear transfer (SCNT) is still plagued by low efficiency. The epigenetic modifications established during cellular differentiation are a major factor determining this low efficiency as they act as epigenetic barriers restricting reprogramming of somatic nuclei. In this regard, most factors that promote chromatin decondensation, including histone deacetylase inhibitors (HDACis), have been found to increase nuclear reprogramming efficiency, making their use common to improve SCNT rates. Herein we used valproic acid (VPA) in SCNT to test whether the treatment of nuclear donor cells with this HDACi improves pre- and post-implantation development of cloned cattle. We found that the treatment of fibroblasts with VPA increased histone acetylation without affecting DNA methylation. Moreover, the treatment with VPA resulted in increased expression of IGF2R and PPARGC1A, but not of POU5F1. However, when treated cells were used as nuclear donors no difference of histone acetylation was found after oocyte reconstruction compared to the use of untreated cells. Moreover, shortly after artificial activation the histone acetylation levels were decreased in the embryos produced with VPA-treated cells. With respect to developmental rates, the use of treated cells as donors resulted in no difference during pre- and post-implantation development. In total, five clones developed to term; three produced with untreated cells and two with VPA-treated cells. Among the calves from treated group, one stillborn calf was delivered at day 270 of gestation whereas the other one was delivered at term but died shortly after birth. Among the calves from the control group, one died seven days after birth whereas the other two are still alive and healthy. Altogether, these results show that in spite of the alterations in fibroblasts resulting from the treatment with VPA, their use as donor cells in SCNT did not improve pre- and post-implantation development of cloned cattle.

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“…The blastocysts produced from urine-and tail skin-derived cells were, however, of superior quality compared to those produced from ear skin cells, in terms of a lower AI. This could be due to a higher level of apoptosis in ear skin cells because the AI has been reported to be higher in bovine blastocysts derived from annexin-positive apoptotic cells than in those derived from annexin-negative nonapoptotic cells (Miranda Mdos et al, 2009). However, a lower level of apoptosis was not reflected in the relative transcript levels of proapoptotic genes CAS-PASE3 and CASPASE9 and in the expression level of P53, which were found to be similar among the blastocysts produced from the three types of cells.…”

Section: Urine-and Tail Skin-derived Cells For Scntmentioning

confidence: 99%

Production of a Cloned Buffalo (Bubalus bubalis) Calf from Somatic Cells Isolated from Urine

Madheshiya

Sahare

Jyotsana

et al. 2015

Cellular Reprogramming

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This study was aimed at isolation of cells from urine and skin on the ventral part of the tails of healthy adult female buffaloes (Bubalus bubalis), an area rarely exposed to solar radiation, establishment of the cells in culture, and their use as donor cells for production of buffalo embryos by handmade cloning (HMC). The blastocyst rate and total cell number of urine-and tail skin-derived embryos were similar to those of control embryos derived from ear skin cells; however, their apoptotic index was lower ( p < 0.05) than that of control blastocysts. The global level of histone H3 acetylated at lysine 9 (H3K9ac) was similar in the three types of donor cells and in urine-and tail skin-derived HMC blastocysts and in vitro-fertilized (IVF) blastocysts (controls). The global level of histone H3 trimethylated at lysine 27 (H3K27me3) in the cells was in the order ( p < 0.05) urine ‡ tail skin > ear skin-derived cells, whereas in blastocysts, it was higher ( p < 0.05) in urine-and tail skin-derived HMC blastocysts than that in IVF blastocysts. The expression level of CASPASE3, CAS-PASE9, P53, DNMT1, DNMT3a, OCT4, and NANOG, which was similar in HMC blastocysts of three the groups, was lower ( p < 0.05) than that in IVF blastocysts, whereas that of HDAC1 was similar among the four groups. Following transfer of urine-derived embryos (n = 10) to five recipients (two embryos/recipient), one of the recipients delivered a normal calf that is now 5 weeks old.

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Serum-Starved Apoptotic Fibroblasts Reduce Blastocyst Production but Enable Development to Term after SCNT in Cattle

Cited by 30 publications

References 48 publications

Production of hGFAP-DsRed transgenic Guangxi Bama mini-pigs via somatic cell nuclear transfer

Production of hGFAP-DsRed transgenic Guangxi Bama mini-pigs via somatic cell nuclear transfer

Development to Term of Cloned Cattle Derived from Donor Cells Treated with Valproic Acid

Production of a Cloned Buffalo (Bubalus bubalis) Calf from Somatic Cells Isolated from Urine

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