猪成纤维细胞细胞周期同期化对核移植胚胎发育的影响

Effects of Cell Cycle Synchronization of Fibroblasts on the Development of Porcine Nuclear Transfer Embryos

供体细胞所处的细胞周期及细胞周期同期化的方法对于体细胞核移植（somatic cell nuclear transfer，SCNT）的成功非常重要，本研究对血清饥饿培养处理与培养至完全汇合后的猪成纤维细胞周期同期化水平进行了检测。利用不同方法对猪成纤维细胞同期化处理后，通过流式细胞仪对细胞的细胞周期分布比率进行了检测。将细胞进行血清饥饿24～72h，显著地增加了Go／G。期的细胞百分率（92．2％-93．7％VS．77．8％，P〈0．05）。将细胞培养至完全汇合后再培养24-48h，GgG。期的细胞比例类似于血清饥饿法（94．4%，89．6%）。血清饥饿24h后，置换为10％FBS能逆转至生长期。用这两种不同方法处理后的体细胞作为核移植的供体构建重构胚，分裂率与囊胚率差异不显著（P〉0．05）。结果表明，猪成纤维细胞通过血清饥饿法或者培养至汇合完全均能有效地将细胞周期同期化至G0/G1期，且均可作为体细胞核移植的供体细胞。

Donor cell in cell cycle and cell cycle synchronization are very important for the success of somatic cell nuclear transfer （SCNT）. In this study, we examined the effects of serum starvation, culture to confluence on cell cycle characteristics of porcine fibroblast cells. The effect of the various methods of cell cycle synchronization was determined by flow cytomertry. 24-72 h serum starvation increased （P〈0.05） the proportion of cells at the Go/G1 phase （92.2%-93.7%） as compared to the control group （77.8%）. A similar increase in the percentage of G0/G1 was ob- tained in culturing to confluence group （94.4% and 89.6%）. The serum starvation cells were fully reversible with 10% FBS （fetal bovine serum） after serum deprivation 24 h. While used these fibroblasts synchronized by serum deprivation or culturing to confluence as SCNT donors, there was no significant difference in the cleavage and blastocyst development rate of reconstructed oocytes of two groups （P〉0.05）. In conclusion, porcine fibroblasts were effectively synchronized at Go/G1 stages of the cell cycle by serum starvation or culturing to confluence, and both can be porcine SCNT donors.