Effects of confluency, roscovitine and serum starvation on the cell-cycle synchronization and viability of sheep and goat adult fibroblasts

Eren, Aysel; Arat, Sezen; Tuna, Metin; Bircan, Rıfat

doi:10.1016/j.jbiotec.2014.07.156

Cited by 2 publications

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“…Compared with this study, it was seen that although the roscovitine inhibitor gave higher viable cell ratios compared to other groups, it remained low in G0/G1 ratios. Eren et al (2014) revealed that with roscovitine 66.64% of G0/G1 ratios were obtained in sheep adult fibroblast cells. These results were similar to this study and supported the data they obtained.…”

Section: Discussionmentioning

confidence: 99%

Can roscovitine and trichostatin A be alternatives to standard protocols for cell cycle synchronization of ovine adult and foetal fibroblast cells?

Yagcioglu

Ersoy

Demır

et al. 2023

Reprod Domestic Animals

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Synchronization of donor cells is an important step for the success of somatic cell nuclear transfer application and facilitates the development of embryos. Contact inhibition, serum starvation and different chemical agents are used in synchronizing different types of somatic cells. In this study, to synchronize the primary ovine adult (POF) and foetal (POFF) fibroblast cells to G0/G1 phases, the contact inhibition, the serum starvation, roscovitine and trichostatin A (TSA) methods were used. In the first part of the study, roscovitine (10, 15, 20 and 30 μM) and TSA (25, 50, 75 and 100 nM) were applied for 24 h to determine the optimal concentration for POF and POFF cells. In the second part, optimal concentrations of roscovitine and TSA for these cells were compared with contact inhibition and serum starvation methods. Cell cycle distribution and apoptotic activity analysis were performed by flow cytometry to compare this synchronization methods. Serum starvation method resulted in higher cell synchronization rate in both cells compared to other groups. Although contact inhibition and TSA also achieved high success rates of synchronized cell value, it was observed that the difference between serum starvation and these groups was significant (p < .05). When the apoptosis rates of the two cell types were examined, it was observed that the early apoptotic cells in contact inhibition and late apoptotic cells in the serum starvation were higher than the other groups (p < .05). Although the 10 and 15 μM concentrations of roscovitine gave the lowest apoptosis rates, it was observed that it failed to synchronize both the ovine fibroblast cells to G0/G1 phase. As a result, it was concluded that while roscovitine was not successful to synchronize both the POFF and POF cell lines, TSA (50 nM for POF cells and 100 nM for POFF cells) can be used efficiently as an alternative to the contact inhibition and the serum starvation methods.

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Section: Discussionmentioning

confidence: 99%

Can roscovitine and trichostatin A be alternatives to standard protocols for cell cycle synchronization of ovine adult and foetal fibroblast cells?

Yagcioglu

Ersoy

Demır

et al. 2023

Reprod Domestic Animals

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Methods of karyoplast cell cycle synchronization for increasing the efficiency of somatic cloning of farm animals

Zhukova

2024

Genes & Cells

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This article is aimed to the analysis of methods for synchronizing the cell cycle of the somatic cells (karyoplasts) upon somatic cloning of farm animals. To obtain cloned animals, the oocytes at the metaphase of the second meiotic division without prior activation are applied as recipient cells in most experiments. In most experiments to obtain cloned offspring, oocytes are used as recipient cells at metaphase of the second meiotic division without prior activation, which determines the choice of somatic cells as karyoplasts at the G0/G1 stage, the most optimal for subsequent nuclear reprogramming by oocyte cytoplasmic factors. The most commonly used methods for arresting cells in this phase are serum starvation and/or contact inhibition, which allows up to 90% of cells to be stopped at the G0/G1 stage. However, despite the effectiveness of these methods, they have a number of significant limitations, and therefore the addition of components to the culture medium of somatic cells that prevent the progression of cells through the cell cycle stages is becoming widespread. The advantage of using chemical inhibitors is the ability of a number of them to have a protective effect on somatic cells, preventing the induction of apoptotic changes. The efficacy of butyrolactone-I, mimosine and aphidicolin application is controversial but a number of studies attests the possible using of these drugs to synchronize the karyoplasts. Thus, there is a wide range of methods for effective synchronization of the cell cycle of karyoplasts, and when choosing the optimal method, one should pay attention to the type of cells, the species of animals from which they were obtained, the permissible duration of cultivation under given conditions in order to minimize the negative impact of conditions on karyoplast viability.

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Effects of confluency, roscovitine and serum starvation on the cell-cycle synchronization and viability of sheep and goat adult fibroblasts

Cited by 2 publications

References 0 publications

Can roscovitine and trichostatin A be alternatives to standard protocols for cell cycle synchronization of ovine adult and foetal fibroblast cells?

Can roscovitine and trichostatin A be alternatives to standard protocols for cell cycle synchronization of ovine adult and foetal fibroblast cells?

Methods of karyoplast cell cycle synchronization for increasing the efficiency of somatic cloning of farm animals

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