| 刘红光 文波 康颖 祖旭宇 赵茂勋 王雄宇.三阴性乳腺癌干细胞的富集及CD44+CD24-/low、CXCR4 表达测定[J].,2015,15(4):640-644 |
| 三阴性乳腺癌干细胞的富集及CD44+CD24-/low、CXCR4 表达测定 |
| Enrichment of Triple Negative Breast Cancer StemCells and Detecting thePropotion of CD44+CD24-/low and CXCR4 Protein Expression |
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| DOI： |
| 中文关键词: 三阴性乳腺癌干细胞 癌球 CD44+CD24-/low CXCR4 |
| 英文关键词: Triple negative Breast cancer stemcells Mammosheres CD44+CD24-/low CXCR4 |
| 基金项目:湖南省卫生厅一般项目（B2010-039）；南华大学研究生科研创新项目(2013XCX28) |
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| 中文摘要: |
| 目的：探讨MDA-MB-231 细胞经无血清培养富集三阴性乳腺癌干细胞，观察再成球、集落形成及CD44+CD24-/low、CXCR4
表达。方法：将MDA-MB-231 乳腺癌细胞进行微球体培养，取培养第7-9 天的微球体,判断干细胞富集的程度；比较不同细胞浓度
对癌球细胞成球率影响；流式细胞仪测定CD44+CD24-/low含量；Western blot 法分析CXCR4 蛋白表达；单个癌球细胞再成球能
力；观察癌球与贴壁细胞集落形成。结果：1).在含20 ng/mL EGF，10 ng/mL bFGF，2%b27 无血清培养基中可培养三阴性乳腺癌癌
球，1× 104/mL、2× 104/mL、3× 104/mL、4× 104/mL、5× 104/mL细胞浓度癌球细胞成球率分别为（5.61± 0.02）%、（3.23± 0.54）%、
（2.28± 0.48）%、（1.05± 0.13）%、（0.91± 0.01）%，组间比较差异有统计学意义P 值均＜0.05。2).贴壁MDA-MB-231细胞与癌球细
胞CD44+CD24-/low 含量分别为（38.54 ± 2.00）%VS (66.35 ± 2.06）%，差异有统计学意义P=0.003。3).癌球细胞CXCR4蛋白
表达高于贴壁MDA-MB-231 细胞，灰度扫描分析差异有统计学意义，P=0.03。4).单个癌球细胞具有再成球能力。5).软琼脂糖集落
形成能力癌球需200 个细胞即可见集落形成，而贴壁细胞需1 000 个MDA-MB-231 细胞。结论：1.通过无血清培养可以富集三阴
性乳腺癌干细胞，低细胞密度有利于癌球形成。2.癌球中CD44+CD24-/low含量高于贴壁MDA-MB-231 细胞。3.CXCR4 在癌球中
表达高于贴壁MDA-MB-231 细胞。 |
| 英文摘要: |
| Objective:In order to acquire triple-negative breast cancer stem cells by adherent MDA-MB-231 cells were cultured in
serum-free medium. In the experiment, the morphological observation of the sphere re-formation and colony formation was carried out, in
addition, the protein expression of CD44+CD24-/low and CXCR4 was detected.Methods:Human MDA-MB-231 breast cancer cell lines
were plated in stem cell conditioned culture system to generate mammospheres. Samples of mammospheres in 7-9 days were taken to
evaluate the effects of different MDA-MB-231cell density on the mammosphere-formation efficiency; the proportion of CD44+CD24-/low
was analyzed by flow cytometry, CXCR4 protein expression was tested by western blot, sphere re-formation and colony formation were
assessed by morphological observation.Results:Triple negative breast cancer mammospheres can be enriched in serum-free medium
con- taining 20 ng/mL EGF, 10 ng/mL bFGF and 2%b27. The mammosphere formation efficiency was (5.61± 0.02) %, (3.23± 0.54) %,
(2.28± 0.48) %, (1.05± 0.13) %and (0.91± 0.01)% when the MDA-MB-231 cell culturing density was 1× 104/mL, 2× 104/mL, 3×
104/mL, 4× 104/mL and 5× 104/mL, respectivel. There was a significantly difference among these groups (P<0.05). The proportion of
CD44+ CD24-/low in MDA-MB-231 adherent cells and mammosheres were (38.54 ± 2.00) % VS (66.35 ± 2.06) %, P=0.003. CXCR4
protein expression in mammosheres cells was higher than MDA-MB-231 adherent cells, P=0.03. Single mammosphere cell has the ability
to generate sphere again. Colony formations of mammosheres need 200 cells and adherent MDA-MB-231 need 1000 cells in soft agarmedia.Conclusion:1.Triple negative breast cancer stem cells can be enriched in serum-free medium, low cell density was beneficial to the
formation of mammosheres. 2. Mammosheres had a higher CD44+CD24-/low proportion than MDA-MB-231 adherent cells. 3.CXCR4
protein expression in mammosheres was higher than in MDA-MB-231 adherent cells. |
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