靶向垂体特异转录因子-1的小干扰RNA对大鼠生长激素垂体腺瘤GH3细胞生物学行为的影响

Effect of pituitary-specific transcription factory small interfering RNA on biological behaviors of growth hormone pituitary adenoma GH3 cells in rats

目的观察小干扰RNA（siRNA）抑制大鼠垂体生长激素腺瘤细胞（GH3）中垂体特异转录因子．1（Pit-1）的基因表达对其生物学行为的影响。方法设计合成针对Pit-1基因的siRNA，经脂质体包裹转染GH3细胞。采用Westernblot、逆转录-聚合酶链反应（RT—PCR）法测定经siRNA转染24h后；Pit．1蛋白和Pit-1mRNA表达；用细胞计数试剂盒（CCK-8）细胞增殖-毒性检测法检测细胞生长增殖的变化；用流式细胞仪检测细胞凋亡的改变；制备Transwell小室测定GH3细胞体外侵袭能力的变化。结果siRNA显著地抑制Pit-1蛋白和Pit-1mRNA的表达水平，分别降低58．18％和75．82％；CCK．8法结果显示siRNA组细胞增长率明显低于与阴性对照组；流式细胞仪分析显示，siRNA组细胞凋亡率[（9．25000±0．99695）％]较阴性对照组[（2．89330±0．66516）％]增加；，siRNA组穿过Matrigel胶及滤膜的细胞数[（92．91±53．25）个]，与阴性对照组[（7．50±7．95）个]比较侵袭力明显减弱，差异有统计学意义（P〈0．05）。结论Pit-1siRNA能够有效地抑制GH3细胞的生长、促进其凋亡，降低侵袭力。

Objective To observe the effects of pituitary-specific transcription factory （ Pit-1 ） small interfering RNA （siRNA） on biological behaviors of growth hormone pituitary adenoma GH3 cells in rats. Methods Design and synthesis of the Pit-1 gene of siRNA, mixed with liposome transfection of GH3 cells. Using protein immunoblot assay （ Western blotting）, reverse transcription polymerase chain reaction （RT-PCR） assay for the determination, after siRNA translated 24 h, of Pit-1 protein and Pit-1 mRNA ex- pression ;with cell counting kit-8 （CCK-8） cell proliferation-toxicity test in the detection of cell growth in- crement change;by flow cytometry for detection of apoptosis changes ; system preparation of Transwell cell determination of GH3 cells in vitro invasion ability changes. Results SiRNA significantly inhibited Pit-1 protein and the Pit-1 mRNA the expression level decreased by 58. 18% and 75.82%, respectively; CCK-8 method results showed that the siRNA group cell growth significantly lower than the negative control group; siRNA group increased apoptosis rate （9. 250 00 ± 0. 996 95 ） % compared with the negative control group （2. 893 30 ±0. 665 16） % ;siRNA group through the Matrigel gum and cell membrane （92, 91± 53.25 ） months ,with the negative control group （7. 50 ± 7.95 ） comparison invasion force significantly weakened, and the difference was statistically significant （ P 〈 0. 05 ）. Conclusion Pit-1 siRNA can effectively inhibit GH3 cell growth, promote their apoptosis, reduced invasiveness.