不同浓度曲古抑菌素A对鸡胚翅芽发育的影响

EFFECT OF DIFFERENT CONCENTRATION OF TRICHOSTATIN A ON CHICKEN WING BUD DEVELOPMENT

目的观察组蛋白去乙酰化酶(HDAC)抑制剂曲古抑菌素A(TSA)对鸡胚翅芽发育中某些基因表达水平的作用,探讨不同浓度TSA(75μmol/L、750μmol/L、1.5mmol/L)对鸡胚翅芽发育的影响,提高我们对染色质结构重组和表观遗传对基因表达的调控作用,以及对正在开发的抗癌药物的认识。方法以鸡胚翅芽作为实验模型,用TSA处理翅芽,对鸡胚胎实施原位杂交,检测与肌肉发育相关基因的表达,并采用硫酸耐尔蓝(Nile blue sulfate)法检测细胞凋亡状况。结果75μmol/L TSA能够抑制与肌肉发生相关的基因MyoD和Myf5在鸡胚翅芽的表达,经TSA(75μmol/L)处理的胚胎没有发生明显的细胞凋亡。但是随着TSA药物浓度的提高,TSA(≥750μmol/L)不仅强烈抑制相关基因的表达,而且出现翅芽外部畸形(外形改变、体积缩小)以及细胞凋亡。结论75μmol/LTSA调控某些重要的转录因子及有力地控制肌肉细胞的分化;高浓度TSA(≥750μmol/L)导致细胞凋亡和胚胎翅芽畸形;发育的鸡胚能够作为一个便利的、供体内研究药物(如HDAC抑制剂类等)的模型。

Objective To investigate the reaction of the limbs treated with different concentration of Trichostatin A （75umol/L,750 umol/L, 1 .Smmol/L）,a histone deacetylase （HDAC）-inhibitor. This may be useful to improve our understanding of the role of chromatin remodelling and epigenetic control of gene expression patterns and ultimately the development of drugs against cancer. Methods Using the chicken embryonic limb as an experimental model. The embryos received grafts of TSA soaked beads or PBS control beads into the right forelimb buds. Then they were submitted to in situ hybridization with probes and apoptosis test. Results TSA could modulate the expression of some myogenesis related genes, MyoD and Myf5 during chicken myogenesis. Using apoptosis staining methods, there was no significant apoptosis in the TSA （75umol/L） treated embryos. However the induction of morphological changes and apoptosis at specific stage was possible with high concentration of TSA. Conclusion TSA （75umol/L） regulates certain important transcriptional targets and strongly control muscle differentiation. Increasing the concentration of TSA （≥ 750umol/L） can induce apoptosis and embryonic limb malformations. Chicken limb development can serve as a convenient in vivo model for studying the effect of HDAC inhibitors.