The transcription factor nuclear factor kB (NF-kB) plays a key role in the delayed xenograft rejection (DXR). One of the important objects in the field is how to inhibit the NF-kB activity at optimal level. Thus, a mo...The transcription factor nuclear factor kB (NF-kB) plays a key role in the delayed xenograft rejection (DXR). One of the important objects in the field is how to inhibit the NF-kB activity at optimal level. Thus, a modified E1A gene (E1AD) containing function domain (1—80 aa) and nuclear localization domain (139—243 aa) was used and cloned into an eucaryotic expression vector pcDNA3 to transfect the porcine aortic endothelial cells (PAEC). The stable transfectants were screened with G418. E1AD gene was able to be stably expressed in the PAEC and could not affect the growth of PAEC as analyzed by RT-PCR and cell growth rate. Reporter gene assay demonstrated that E1AD was capable of inhibiting NF-kB activity in the PAEC in-duced by TNF-a without sensitizing to apoptosis, and the rate of inhibition was 53%. Furthermore, E1AD inhibited the expression of a NF-kBdependent inflammatory gene E-selectin in the cells, and the rate of inhibition was 63%. In summary, the usage of E1AD gene may be a new strategy to overcome DXR in the xenotransplantation.展开更多
背景与目的:腺病毒5型早期区1A基因(early region 1A,E1A)是新近发现的一个具有抑癌作用的基因,E1A蛋白能够从正、负两种途径调控多种基因的表 达,具有诱导肿瘤细胞分化、逆转其恶性表型、降低其体内致瘤性和抗转移等活性,但E1A基因在...背景与目的:腺病毒5型早期区1A基因(early region 1A,E1A)是新近发现的一个具有抑癌作用的基因,E1A蛋白能够从正、负两种途径调控多种基因的表 达,具有诱导肿瘤细胞分化、逆转其恶性表型、降低其体内致瘤性和抗转移等活性,但E1A基因在头颈淋巴结转移鳞癌治疗中的作用尚少见报道。本研究的目的是探讨E1A基因对头颈淋巴结转移鳞癌细胞体外生长的抑制作用和化、放疗增敏作用及其初步的作用机理。方法:经脂质体介导,将真核细胞高效表达E1A基因的重组质粒peDNA3-E1A导入人舌鳞癌的淋巴结转移癌细胞系686LN-1。通过测绘生长曲线和计算倍增时间,观察E1A基因对该细胞系生长的抑制作用。用MTT法测定转染E1A基因前后,癌细胞对顺铂、泰素、5-氟尿嘧啶和博来霉素等化疗药物以及放疗敏感性的改变。流式细胞术和免疫细胞化学染色检测转染前后的细胞周期分布以及p53和HER-2/neu表达情况。结果:转染E1A基因细胞群体生长缓慢,倍增时间延长(是转染空载体细胞的1.48倍)。与686LN-1-vect比较,686LN-1-E1A细胞对顺铂、博来霉素、泰素和射线的敏感性(IC_(50))分别提高了8倍、20倍、10倍和1倍。细胞周期出现G_2/M期阻滞;免疫细胞化学染色显示E1A基因能抑制HER-2/neu的表达。展开更多
基金This work was supported by Beijing "248"High-Tech Program China.
文摘The transcription factor nuclear factor kB (NF-kB) plays a key role in the delayed xenograft rejection (DXR). One of the important objects in the field is how to inhibit the NF-kB activity at optimal level. Thus, a modified E1A gene (E1AD) containing function domain (1—80 aa) and nuclear localization domain (139—243 aa) was used and cloned into an eucaryotic expression vector pcDNA3 to transfect the porcine aortic endothelial cells (PAEC). The stable transfectants were screened with G418. E1AD gene was able to be stably expressed in the PAEC and could not affect the growth of PAEC as analyzed by RT-PCR and cell growth rate. Reporter gene assay demonstrated that E1AD was capable of inhibiting NF-kB activity in the PAEC in-duced by TNF-a without sensitizing to apoptosis, and the rate of inhibition was 53%. Furthermore, E1AD inhibited the expression of a NF-kBdependent inflammatory gene E-selectin in the cells, and the rate of inhibition was 63%. In summary, the usage of E1AD gene may be a new strategy to overcome DXR in the xenotransplantation.
文摘背景与目的:腺病毒5型早期区1A基因(early region 1A,E1A)是新近发现的一个具有抑癌作用的基因,E1A蛋白能够从正、负两种途径调控多种基因的表 达,具有诱导肿瘤细胞分化、逆转其恶性表型、降低其体内致瘤性和抗转移等活性,但E1A基因在头颈淋巴结转移鳞癌治疗中的作用尚少见报道。本研究的目的是探讨E1A基因对头颈淋巴结转移鳞癌细胞体外生长的抑制作用和化、放疗增敏作用及其初步的作用机理。方法:经脂质体介导,将真核细胞高效表达E1A基因的重组质粒peDNA3-E1A导入人舌鳞癌的淋巴结转移癌细胞系686LN-1。通过测绘生长曲线和计算倍增时间,观察E1A基因对该细胞系生长的抑制作用。用MTT法测定转染E1A基因前后,癌细胞对顺铂、泰素、5-氟尿嘧啶和博来霉素等化疗药物以及放疗敏感性的改变。流式细胞术和免疫细胞化学染色检测转染前后的细胞周期分布以及p53和HER-2/neu表达情况。结果:转染E1A基因细胞群体生长缓慢,倍增时间延长(是转染空载体细胞的1.48倍)。与686LN-1-vect比较,686LN-1-E1A细胞对顺铂、博来霉素、泰素和射线的敏感性(IC_(50))分别提高了8倍、20倍、10倍和1倍。细胞周期出现G_2/M期阻滞;免疫细胞化学染色显示E1A基因能抑制HER-2/neu的表达。
