中华预防医学杂志
中華預防醫學雜誌
중화예방의학잡지
CHINESE JOURNAL OF
2014年
9期
809-814
,共6页
董超%张凤梅%赵锡鹏%郭公社%罗月%凤志慧
董超%張鳳梅%趙錫鵬%郭公社%囉月%鳳誌慧
동초%장봉매%조석붕%곽공사%라월%봉지혜
乳腺肿瘤%BRCA1蛋白质%DNA损伤%HCC1937%MCF7
乳腺腫瘤%BRCA1蛋白質%DNA損傷%HCC1937%MCF7
유선종류%BRCA1단백질%DNA손상%HCC1937%MCF7
Breast neoplasms%BRCA1 protein%DNA damage%HCC1937%MCF7
目的：通过比较DNA损伤修复基因乳腺癌易感基因1（ breast cancer susceptibility gene 1，BRCA1）和抑癌基因p53在HCC1937和MCF7两种乳腺癌细胞系中的功能状态及对DNA损伤修复反应的应答特性，研究两种细胞系在功能上的差异。方法应用western-blotting法检测两种乳腺癌细胞系中BRCA1和p53蛋白的表达，并检测了经过10 Gy电离辐射下1、4、8 h后BRCA1蛋白在MCF7、HCC1937和HCC1937野生型BRCA1（HCC1937 wtBRCA1）细胞系中的表达水平。同时用免疫荧光法观察BRCA1蛋白在MCF7和HCC1937细胞系细胞内的分布和焦点形成情况，并对电离辐射下的BRCA1、Rad51蛋白的焦点形成百分比进行计算。进一步用流式细胞仪检测两种细胞系的细胞周期变化。结果 MCF7细胞中野生型的BRCA1和p53主要分布于细胞核内，这两种蛋白对DNA损伤反应有应答。10 Gy 8 h照射条件下，MCF7细胞系中BRCA1蛋白焦点形成百分比较无电离辐射高[（59.40±3.66）％比（11.80±3.51）％，t＝16.26，P＜0.05]；MCF7细胞系照射组中Rad51蛋白焦点形成百分比较无电离辐射高[（73.90±8.66）％比（16.70±3.76）％，t＝10.49，P＜0.05]，照射组p53蛋白[（82.54±1.04）比（23.75±0.51），t＝87.90，P＜0.05]和p21蛋白[（90.95±1.13）比（50.19±0.89），t＝49.11，P＜0.05]表达水平均较无电离辐射高，细胞在G1期蓄积。与MCF7细胞相比，在HCC1937细胞中的BRCA1和p53都产生了变异，在细胞核内两种蛋白较少。10 Gy 8 h照射条件下HCC1937细胞中无BRCA1蛋白焦点形成、p53和p21几乎无诱导表达以及细胞在G1和G2-M期无明显的蓄积。当恢复野生型BRCA1在HCC1937细胞内表达后，10 Gy 8 h照射条件下Rad51蛋白焦点形成百分比较无电离辐射高[（61.70±4.03）％比（6.22±2.27）％，t＝20.78，P＜0.05]，细胞在G2-M期的蓄积增多。结论乳腺癌细胞系HCC1937和MCF7在应答DNA损伤修复反应时具有不同的功能特性。
目的：通過比較DNA損傷脩複基因乳腺癌易感基因1（ breast cancer susceptibility gene 1，BRCA1）和抑癌基因p53在HCC1937和MCF7兩種乳腺癌細胞繫中的功能狀態及對DNA損傷脩複反應的應答特性，研究兩種細胞繫在功能上的差異。方法應用western-blotting法檢測兩種乳腺癌細胞繫中BRCA1和p53蛋白的錶達，併檢測瞭經過10 Gy電離輻射下1、4、8 h後BRCA1蛋白在MCF7、HCC1937和HCC1937野生型BRCA1（HCC1937 wtBRCA1）細胞繫中的錶達水平。同時用免疫熒光法觀察BRCA1蛋白在MCF7和HCC1937細胞繫細胞內的分佈和焦點形成情況，併對電離輻射下的BRCA1、Rad51蛋白的焦點形成百分比進行計算。進一步用流式細胞儀檢測兩種細胞繫的細胞週期變化。結果 MCF7細胞中野生型的BRCA1和p53主要分佈于細胞覈內，這兩種蛋白對DNA損傷反應有應答。10 Gy 8 h照射條件下，MCF7細胞繫中BRCA1蛋白焦點形成百分比較無電離輻射高[（59.40±3.66）％比（11.80±3.51）％，t＝16.26，P＜0.05]；MCF7細胞繫照射組中Rad51蛋白焦點形成百分比較無電離輻射高[（73.90±8.66）％比（16.70±3.76）％，t＝10.49，P＜0.05]，照射組p53蛋白[（82.54±1.04）比（23.75±0.51），t＝87.90，P＜0.05]和p21蛋白[（90.95±1.13）比（50.19±0.89），t＝49.11，P＜0.05]錶達水平均較無電離輻射高，細胞在G1期蓄積。與MCF7細胞相比，在HCC1937細胞中的BRCA1和p53都產生瞭變異，在細胞覈內兩種蛋白較少。10 Gy 8 h照射條件下HCC1937細胞中無BRCA1蛋白焦點形成、p53和p21幾乎無誘導錶達以及細胞在G1和G2-M期無明顯的蓄積。噹恢複野生型BRCA1在HCC1937細胞內錶達後，10 Gy 8 h照射條件下Rad51蛋白焦點形成百分比較無電離輻射高[（61.70±4.03）％比（6.22±2.27）％，t＝20.78，P＜0.05]，細胞在G2-M期的蓄積增多。結論乳腺癌細胞繫HCC1937和MCF7在應答DNA損傷脩複反應時具有不同的功能特性。
목적：통과비교DNA손상수복기인유선암역감기인1（ breast cancer susceptibility gene 1，BRCA1）화억암기인p53재HCC1937화MCF7량충유선암세포계중적공능상태급대DNA손상수복반응적응답특성，연구량충세포계재공능상적차이。방법응용western-blotting법검측량충유선암세포계중BRCA1화p53단백적표체，병검측료경과10 Gy전리복사하1、4、8 h후BRCA1단백재MCF7、HCC1937화HCC1937야생형BRCA1（HCC1937 wtBRCA1）세포계중적표체수평。동시용면역형광법관찰BRCA1단백재MCF7화HCC1937세포계세포내적분포화초점형성정황，병대전리복사하적BRCA1、Rad51단백적초점형성백분비진행계산。진일보용류식세포의검측량충세포계적세포주기변화。결과 MCF7세포중야생형적BRCA1화p53주요분포우세포핵내，저량충단백대DNA손상반응유응답。10 Gy 8 h조사조건하，MCF7세포계중BRCA1단백초점형성백분비교무전리복사고[（59.40±3.66）％비（11.80±3.51）％，t＝16.26，P＜0.05]；MCF7세포계조사조중Rad51단백초점형성백분비교무전리복사고[（73.90±8.66）％비（16.70±3.76）％，t＝10.49，P＜0.05]，조사조p53단백[（82.54±1.04）비（23.75±0.51），t＝87.90，P＜0.05]화p21단백[（90.95±1.13）비（50.19±0.89），t＝49.11，P＜0.05]표체수평균교무전리복사고，세포재G1기축적。여MCF7세포상비，재HCC1937세포중적BRCA1화p53도산생료변이，재세포핵내량충단백교소。10 Gy 8 h조사조건하HCC1937세포중무BRCA1단백초점형성、p53화p21궤호무유도표체이급세포재G1화G2-M기무명현적축적。당회복야생형BRCA1재HCC1937세포내표체후，10 Gy 8 h조사조건하Rad51단백초점형성백분비교무전리복사고[（61.70±4.03）％비（6.22±2.27）％，t＝20.78，P＜0.05]，세포재G2-M기적축적증다。결론유선암세포계HCC1937화MCF7재응답DNA손상수복반응시구유불동적공능특성。
Objective The functional characters of MCF 7 and HCC1937 cell lines were compared through the activity of BRCA 1 and p53 following DNA damage in order to provide more research evidence for the related studies in both breast cancer cell lines.Methods The protein level of BRCA1 and p53 in two breast cancer cell lines and the protein level of BRCA 1 in MCF7,HCC1937 and HCC1937 wtBRCA1 breast cancer cell lines treated with 10Gy after 1 h,4 h or 8 h were detected by western blotting analysis.The distribution and foci formation of BRCA 1 in the cells were observed through immunostaining assay and the percentage of BRCA1 or Rad51 foci formation after ionizing radiation was calculated.Cell cycle profiling was analyzed using flow cytometry.Results Most of BRCA1 and p53 localized in nucleus , and both proteins responded to DNA damage in MCF 7 cells.In MCF7 cells, BRCA1 and Rad51 foci formation respectively increased to (59.40 ±3.66)%from(11.80 ±3.51)%(t=16.26,P<0.05)and (73.90 ±8.66)% from (16.70 ±3.76)%(t=10.49,P<0.05) after 10 Gy 8 h;p53 and p21 protein level was further separately induced and enhanced to (82.54 ±1.04) from (23.75 ±0.51) (t =87.90,P<0.05) and (90.95 ± 1.13) from (50.19 ±0.89)( t=49.11,P<0.05) after 10 Gy 8 h;and the cells were accumulated in G1 phase.In contrast to MCF7, in HCC1937 cell line, both of BRCA1 and p53 were defective in nucleus since both proteins were mutated;in response to DNA damage , BRCA1 foci formation was not found , p53 and p21 was not induced; there was no cell accumulation in both of G 1-S and G2-M phases.However , after complementation of wild-type BRCA1 in HCC1937 cells, DNA damage-induced Rad51 foci formation increased to (61.70 ±4.03)%from (6.22 ±2.27)%( t=20.78,P<0.05) and accumulation of cells in G2-M phase was also restored after 10 Gy 8h , which was similar to that of in MCF7 cells.Conclusions We have identified that BRCA1 and p53 have dramatically different functions in MCF 7 and HCC1937 cell lines in response to DNA damage.
