CAJ | 학술논문

目的探讨p53基因在电离辐射(IR)诱导的MCF-7细胞周期解耦联中的作用.方法构建RNAi表达载体,经磷酸钙共沉淀法转染293T细胞形成病毒包装颗粒,感染MCF-7后采用Western blot检测P53蛋白的表达,建立p53基因沉默模型.将p53野生型(+/+)和沉默模型(-/-)经电离辐射处理后,采用流式细胞术分别测定细胞周期并分析细胞多倍体的变化.结果与p53+/+组比较,p53-/-模型组G0/G1期细胞百分数减少,S期、G2期增加(P<0.01),倍体分析表明二倍体数减少,四倍体、八倍体均增加(P<0.01).在p53+/+和p53-/-细胞中,与假照射组比较,4 Gy照射后G0/G1期、S期细胞百分数减少,而G2期增多(P<0.01);倍体分析表明,照射后二倍体数减少,四倍体、八倍体均增加(P<0.01).与p53+/++IR组比较,p53-/-+IR组发生G0/G1期、S期细胞百分数减少,G2+M期增多(P<0.01),二倍体数减少,四倍体增多(P<0.01),八倍体无明显差别.结论电离辐射可以诱导细胞发生G2期阻滞和细胞周期解耦联;P53在电离辐射诱导的MCF-7细胞G2期阻滞中发挥作用,而在细胞周期解耦联中可能不发挥作用.
목적 탐토p53기인재전리복사(IR)유도적MCF-7세포주기해우련중적작용.방법 구건RNAi표체재체,경린산개공침정법전염293T세포형성병독포장과립,감염MCF-7후채용Western blot검측P53단백적표체,건립p53기인침묵모형.장p53야생형(+/+)화침묵모형(-/-)경전리복사처리후,채용류식세포술분별측정세포주기병분석세포다배체적변화.결과 여p53+/+조비교,p53-/-모형조G0/G1기세포백분수감소,S기、G2기증가(P<0.01),배체분석표명이배체수감소,사배체、팔배체균증가(P<0.01).재p53+/+화p53-/-세포중,여가조사조비교,4 Gy조사후G0/G1기、S기세포백분수감소,이G2기증다(P<0.01);배체분석표명,조사후이배체수감소,사배체、팔배체균증가(P<0.01).여p53+/++IR조비교,p53-/-+IR조발생G0/G1기、S기세포백분수감소,G2+M기증다(P<0.01),이배체수감소,사배체증다(P<0.01),팔배체무명현차별.결론 전리복사가이유도세포발생G2기조체화세포주기해우련;P53재전리복사유도적MCF-7세포G2기조체중발휘작용,이재세포주기해우련중가능불발휘작용.
Objective To explore the roles of p53 in ionizing radiation induced MCF-7 cell cycle uncoupling. Methods The p53 knock-down models was established in MCF-7 with retrovirus packaged particles from 293T cells through calcium acid phosphate co-precipitation, then Western blot was used to detect the protein expression. Flow cytometry(FCM) was used to analyze the cell cycle uncoupling and polyploid after irradiation. Results Compared with p53+/+ group, the percentages of G0/G1 cells in p53 -/- group decreased, while those of S and G2+M increased (P < 0.01). In polyploidy analysis 2N cells decreased, whereas both 4N and 8N cells =increased (P<0.01). Compared with sham-irradiation, 4 Gy X-ray led to the decrease of G0/G1, S cells, and the increase of G2+M cells. The increase of 2N cells and decrease of 4N and 8N cells were observed in both p53+/+ and p53-/- cells. Compared with p53+/+ +IR group, the decrease of G0/G1 and S cells and the increase of C2 + M cells were significant (P < 0.01) in p53-/-+ IR groups. 2N cells decreased, 4N cells increased, but no changes in 8 N cells occurred. Conclusion Radiation might induce G2 arrest and cycle uncoupling, p53 plays a role in the regulation of G2 arrest, but no role in cycle uncoupling.