Effect of Breast Cancer Associated Fibroblast on Breast Caner Cells
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摘要: 目的 研究乳腺癌相关成纤维细胞（CAFs）对乳腺癌细胞株增殖和乳腺癌干细胞比例的影响。方法分离和鉴定人乳腺癌相关成纤维细胞和非肿瘤成纤维细胞（NAFs），用无血清培养液培养CAFs和NAFs，收集上清液，分别与乳腺癌细胞株MCF-7和MDA-MB-468共培养48h，通过细胞计数以及流式细胞术，检测其对乳腺癌细胞增殖和细胞周期的影响；并利用流式细胞术，检测乳腺癌干细胞（CD44+CD24-）的细胞比例。结果NAFs和CAFs中表达α-平滑肌肌动蛋白（α-SMA）的细胞比例分别为（6.00±0.57）% 、（53.33±2.33）%(P<0.001)；培养NAFs和CAFs的上清液对乳腺癌细胞株的增殖和细胞周期没有明显的影响，上清液处理48h后，MCF-7细胞对照组、NAFs组和CAFs组的G2/M期细胞比例分别为（5.32±0.02）%、（5.63±0.03）%、（6.03±0.03）%(P>0.05)；CAFs和NAFs均能诱导乳腺癌干细胞的产生，CAFs组和NAFs组的MCF-7干细胞比例分别为对照组的3.51和1.41倍(P<0.05)；MDA-MB-468干细胞比例分别为对照组的4.76和1.35倍(P<0.05)。结论乳腺CAFs高表达α-SMA，培养CAFs的上清液在体外对乳腺癌细胞的增殖没有明显影响，但能诱导乳腺癌干细胞的产生，提示其可能在肿瘤复发和转移中起重要作用。Abstract: Objective To study the effects of breast cancer associated fibroblasts(CAFs)on breast cancer sells in vitro. Methods The CAFs and non-cancer associated fibrolasts(NAFs) were isolated from primary breast tumors and their paired normal breast tissue.The CAFs and NAFs were cultured in 1640 medium without the supplement of serum，and the supernatant were collected.Breast cancer cell lines，MCF-7 and MDA -MB-468 were cultured in the collected supernatant.After 48h of culture，the cells were collected and calculated.Then the cell cycles were tested and the breast cancer stem cell markers CD44 and CD24 were tested by Flow Cytometry. Results We successfully isolated and cultured the NAFs and CAFs.The CAFs increased the expression of α-smooth muscle actin(α-SMA),a defining characteristic of myofibroblasts.The proportion of α-SMA positive cells in NAFs and CAFs were（6.00±0.57）%, （53.33±2.33）%(P<0.001),respectively.The CAFs and NAFs had no effects on breast cancer proliferation in vitro.After 48h of culture，the proportion of G2/M phase of MCF-7 were（5.32±0.02 ）%,（5.63±0.03）%,（6.03±0.03）%(P>0.05)for the control group,NAFs group and CAFs group respectively.Both of the NAFs and CAFs increased the proportion of CD44+CD24-cells in MCF-7 and MDA -MB-468.The proportions of CD44+CD24-cells in MCF-7 were 0.41 and 2.51 folds(P<0.05) higher in the NAFs group and CAFs group respectively than those in the control group.And there were 0.35 and 3.76 folds(P<0.05) higher in the MDA-MB-468 respectively. Conclusion The CAFs induces α-SMA expression，and has no effects on breast cancer proliferation in vitro，although significantly increases the rates of cancer stem cells.The CAFs might play an important role in the progresses of cancer recurrence and metastasis.
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Key words:
- Breast cancer /
- Cancer associated fibroblasts /
- Cancer stem cell
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