[1]唐娜,周略,成志强,等.手术切除的结直肠癌组织中的肿瘤干细胞的培养及鉴定[J].南方医科大学学报,2019,(04):415.[doi:10.12122/j.issn.1673-4254.2019.04.06]
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手术切除的结直肠癌组织中的肿瘤干细胞的培养及鉴定()
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《南方医科大学学报》[ISSN:1673-4254/CN:44-1627/R]

卷:
期数:
2019年04期
页码:
415
栏目:
出版日期:
2019-04-15

文章信息/Info

Title:
Culture and identification of tumor stem cells from surgically resected colorectal cancer tissues
作者:
唐娜周略成志强邓永键丁彦青
关键词:
肿瘤干细胞结直肠癌原代培养
Keywords:
cancer stem cells colorectal cancer primary culture
DOI:
10.12122/j.issn.1673-4254.2019.04.06
摘要:
目的获得结直肠癌患者手术切除癌组织的肿瘤干细胞(CSC),培养并鉴定其干细胞特性。方法通过原代培养手术切除 结直肠癌组织获得结直肠癌细胞,再通过有限稀释法和无血清培养法筛选结直肠癌CSC,软琼脂集落实验检测CSC和对照组人 结直肠癌细胞株SW480 的增殖能力;用低数量级细胞裸鼠皮下成瘤实验,检测结直肠癌CSC和SW480 的成瘤能力;并运用 Western blot及免疫荧光方法,检测CSC和SW480的免疫表型。结果临床结直肠癌标本原代培养的CSC,加入血清培养可使 CSC分化。软琼脂集落形成实验结果显示,原代培养的CSC克隆形成率为56.64%和54.45%,对照组人结直肠癌细胞株SW480 的克隆形成率为4.41%,CSC与SW480集落形成率的差异具有统计学意义(P<0.01)。裸鼠皮下成瘤实验中,原代培养的CSC 皮下注射500 个细胞可形成皮下瘤,而SW480 皮下注射500 个细胞不形成皮下瘤。CSC 表达CD133、CD44,不表达CK7; SW480细胞不表达CD133、CD44,表达CK7。结论原代培养的手术切除结直肠癌组织标本获得的肿瘤细胞,再行无血清培养 可形成CSC,鉴定具有CSC的自我更新及分化能力。
Abstract:
Objective To obtain cancer stem cells (CSCs) from surgically resected colorectal cancer specimens and identify their stem cell characteristics. Methods Colorectal cancer tissue specimen obtained from a patient undergoing radical resection of colorectal cancer were implanted in nude mice, and the xenograft was harvested 1 month later to obtain purified tumor cells by enzyme digestion and adherent culture. The CSCs were screened by limiting dilution method and serum-free culture to identify their phenotypes. Soft agar colony assay was used to assess the proliferative ability of the CSCs and human colorectal cancer cell line SW480. The tumorigenic ability of the isolated CSCs and SW480 cells was evaluated by observing their subcutaneous tumor formation in nude mice. Western blotting and immunofluorescence assay were used to detect the immunophenotype of the CSCs and SW480 cells. Results The primary cultured CSCs from clinical specimens of colorectal cancer underwent differentiation in the presence of serum in the culture. Soft agar colony formation assay showed that the CSCs had a colony formation rate above 50%, significantly higher than the rate of colorectal cancer SW480 cells (4.41%; P<0.01). In nude mice, subcutaneous injection of 500 CSCs was sufficient to result in subcutaneous tumor formation, while the injection of 500 SW480 cells failed to form any subcutaneous tumors. The CSCs expressed CD133 and CD44 but not CK7, while SW480 cells expressed CK7 but not CD133 or CD44. Conclusion CSCs can be derived by primary culture of cancer cells obtained from surgically resected colorectal cancer tissue followed by serum-free culture, and the CSCs obtained have self-renewal and differentiation abilities.

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更新日期/Last Update: 1900-01-01