在单核细胞中干扰SHP-2后对宫颈癌细胞的影响  

Effects of interference of SHP-2 in monocytes on cervical cancer cells

作　　者：穆庆[1] 魏恒[1] 张瑶[1] 罗清双[1] 张淑兰[1] 

机构地区：[1]中国医科大学附属盛京医院妇产科,辽宁沈阳110011

出　　处：《中国妇幼保健》2013年第14期2300-2303,共4页Maternal and Child Health Care of China

摘　　要：目的:研究宫颈感染HPV后是否能通过干扰SHP-2从而负调控IFN-β信号通路,最终导致宫颈癌的发生。方法:通过SiHa细胞与THP-1细胞共培养,利用SHP-2 siRNA干扰片段瞬时转染THP-1细胞,干扰SHP-2基因的表达。将SHP-2封闭肽以0、0.5、2.5μg/ml浓度和2、24 h培养THP-1细胞,收集培养后的细胞进行Western Blotting实验,进而检测SHP-2的表达,确定封闭的最佳时间和浓度。收集正常和不同处理因素的THP-1细胞,Western Blotting法检测THP-1细胞中SHP-2蛋白表达。将THP-1细胞及不同处理因素处理好的THP-1细胞分别与SiHa细胞共培养后,在选定的时间和浓度点测其OD值。收集正常THP-1细胞的培养液上清、THP-1细胞给予SHP-2封闭肽处理的培养液上清和THP-1细胞给予SHP-2 siRNA干扰的培养液上清进行ELISA实验。结果:给予SHP-2的SiRNA后SHP-2蛋白表达明显降低,在干扰后48 h、200 pmol时干扰效果最好。给予SHP-2的封闭抗体后SHP-2蛋白表达明显降低,在封闭后24 h、2.5μg/ml时抑制效果最好。THP-1细胞与SiHa细胞相互作用,从两个细胞作用时间来看,THP-1细胞对SiHa细胞杀伤作用随着时间的增长而增强,从24 h始就起明显作用。给予siRNA干扰的THP-1细胞对SiHa细胞的杀伤作用最强(OD值最低),给予封闭肽的次之,无处理因素的最弱(OD值最高)。SiRNA干扰组的浓度值最高,其次是封闭肽组的浓度值,无处理因素组(仅培养THP-1的细胞组)的浓度值最低,SiRNA干扰组与无处理因素组之间差异有统计学意义(P<0.05),封闭肽组与无处理因素组之间差异有统计学意义(P<0.05),SiRNA干扰组与封闭肽组之间差异无统计学意义(P>0.05)。结论:抑制THP-1细胞中SHP-2的表达后,THP-1细胞分泌的IFN-β明显增多,导致它对SiHa细胞的杀伤作用明显增强。SHP-2在宫颈癌的发生中起着重要作用,它发挥作用的机制是通过抑制IFN-β的信号转导,减少I型干扰素IFN-β的生成,使宫颈癌发生成为�Objective： To investigate whether SHP - 2 negatively regulated IFN - β signaling pathway after HPV infection, eventu- ally leading to cervical cancer. Methods： THP - 1 cells was co - cultured with SiHa cells. After THP - 1 ceils were transiently transfected with small interfering RNAs （siRNA） against human SHP- 2, gene silencing was monitored at the protein levels by immunoblotting using anti - SHP -2 antibodies. The SHP -2 blocking peptide at different concentrations （0, 0. 5, 2. 5 μg / ml） and time （2 h and 24 h）, ac- ted on the THP - 1 ceils. Western blotting was carried out to detect the expression of SHP - 2 to determine the closed best time and concentra- tion. THP - 1 ceils with normal and different treatment factors were collected, SHP - 2 protein expression in THP - 1 cells was detected by Western blotting. THP - 1 cells and the THP - 1 cells treated by different factors were cultured with SiHa cells individually, the results were measured in the selected points of time and the concentration. Cell culture supernatants were collected from normal, SHP - 2 blocking pep- tide treated and SHP - 2 siRNA treated THP - 1 cells. Cell culture supernatants were examined. Results： SHP - 2 protein expressions were decreased after SHP - 2 siRNA, the effect was best when the interference time was 48 h and concentration was 200 pmol. SHP - 2 protein ex- pressions were decreased after blocking peptide, the effect was best when the blocking time was 24 h and concentration was 2. 5 txg/ml. OD result in replacement of the THP - 1 cell culture medium was higher than that of original culture medium, the difference was statistically sig- nificant. From time to look at the two cells, THP - 1 cells killing effect on SiHa cells were more and more enhanced from the 24 hours. The interaction for THP - 1 cells of siRNA interfering on SiHa cells was strongest, THP - 1 cells of blocking peptide was stronger, and nothing done was weakest. There was statistically significant difference in the expression of IFN - β b

关 键 词：SHP-2 IFN—β宫颈癌THP-1细胞 

分 类 号：R737.33[医药卫生—肿瘤]