Treatment with Proteasome Inhibitor MG132 during Cloning Improves Survival and Pronuclear Number of Reconstructed Rat Embryos
Treatment with Proteasome Inhibitor MG132 during Cloning Improves Survival and Pronuclear Number of Reconstructed Rat Embryos
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DOI：
10.1089/clo.2008.0038
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发表时间：
2008-12-01
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影响因子：
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通讯作者：
Kashiwazaki, Naomi
中科院分区：
文献类型：
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作者：
Nakajima, Noriaki;Inomata, Tomo;Kashiwazaki, Naomi
在包括大鼠在内的几种哺乳动物物种中，已经使用体细胞核移植（SCNT）成功地克隆了动物。然而，在大鼠的情况下，额外的治疗与MG 132，蛋白酶体抑制剂，去核前的卵母细胞似乎需要成功克隆，因为排卵的大鼠卵母细胞自发激活，因此，他们的抑制是成功克隆的关键。先前对大鼠的研究表明，与小鼠卵母细胞相比，成熟卵母细胞可能具有较低的细胞生长抑制因子（CSF）活性，导致SCNT后重建胚胎中过早染色体凝聚的发生率较低。已知通常观察到具有两个以上原核的小鼠。在核转移卵母细胞中，诱导过早的染色体凝聚后，这意味着成功的重编程。这导致我们假设MG 132处理不仅影响自发激活的抑制，而且影响重构大鼠胚胎的重编程和发育能力。如果是这样，在SCNT期间和/或之后延长MG 132治疗可能进一步改善存活率。然而，在大鼠和其他物种中，MG 132处理对SCNT后重构胚胎的影响非常有限。我们在这里表明，在SCNT期间和之后延长MG 132治疗可提高活化后重建大鼠胚胎的存活率和原核数量。与对照胚胎相比，在SCNT之前、期间和之后处理的这些重建胚胎显示出显著更高的p34（cdc 2）激酶活性，涉及CSF活性。另一方面，p34（cdc 2）激酶活性没有恢复在核移植卵母细胞没有MG 132，这表明去核对重构卵母细胞的发育有不利影响。总之，在SCNT期间MG 132处理通过维持高CSF活性增加了重构大鼠胚胎的存活率和原核数。数据表明，MG 132治疗至少对大鼠SCNT是不可或缺的。
In several mammalian species including rats, successfully cloned animals have been generated using somatic cell nuclear transfer (SCNT). However, in the case of rats, additional treatment with MG132, a proteasome inhibitor, before enucleation of oocytes seems to be required for successful cloning because ovulated rat oocytes are spontaneously activated, and hence, their suppression is the key to successful cloning. A previous study on rats demonstrated that matured oocytes potentially possess lower cytostatic factor (CSF) activity compared to mouse oocytes, resulting in a low incidence of premature chromosome condensation in the reconstructed embryos after SCNT. It is known that mice having more than two pronuclei are generally observed. in nuclear-transferred oocytes after induction of premature chromosome condensation, which implies successful reprogramming. This leads us to the hypothesis that MG132 treatment affects not only the inhibition of spontaneous activation but also the reprogramming and developmental ability of reconstructed rat embryos. If so, prolonged MG132 treatment during and/or after SCNT may further improve the survivability. However, the effect of MG132 treatment on reconstructed embryos after SCNT has been very limited in rats and other species. We show here that prolonged MG132 treatment during and after SCNT improves survival and the number of pronuclei in reconstructed rat embryos after activation. These reconstructed embryos treated before, during, and after SCNT showed significantly higher p34(cdc2) kinase activity involving CSF activity compared to that of the control embryos. On the other hand, p34(cdc2) kinase activity was not recovered in nuclear-transferred oocytes without MG132, which suggested that the enucleation had detrimental effects on the development of reconstructed oocytes. Taken together, MG132 treatment during SCNT increases survival and pronuclear numbers in reconstructed rat embryos via maintenance of high CSF activity. The data suggest that MG132 treatment is indispensable for at least rat SCNT.