WAS-like 蛋白在肠癌组织中低表达并抑制人结肠癌细胞的干性

李宏丹; 杨成

doi:10.16098/j.issn.0529-1356.2022.01.007

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解剖学报 ›› 2022, Vol. 53 ›› Issue (1) : 50-59. DOI: 10.16098/j.issn.0529-1356.2022.01.007

WAS-like 蛋白在肠癌组织中低表达并抑制人结肠癌细胞的干性

李宏丹^1* 杨成²

作者信息 +

Wiskot-Aldrich syndrome-like lower expressed in colon cancer tissues and inhibited the stemness of human colon cancer

LI Hong-dan^1* YANG Cheng²#br#

Author information +

文章历史 +

摘要

目的探讨WAS-like 蛋白(WASL)在肠癌的表达情况，以及对肠癌细胞干性的影响。方法运用UALCAN数据库预测WASL蛋白的表达量与肠癌组织中的表达及其与肠癌患者的性别、临床分期、淋巴结转移的关系；组织免疫荧光和细胞免疫荧光染色验证WASL在肠癌组织和细胞中的表达。Western blotting检测人正常结直肠黏膜细胞（FHC细胞）与肠癌细胞HCT-116和SW-480中WASL蛋白的表达情况及WASL的转染效率；应用干细胞球（sphere）形成实验和集落形成实验检测转染后，WASL对肠癌细胞中癌干细胞（CSCs）的影响；Real-time PCR检测CSCs相关基因八聚体结合转录因子4（OCT4），性别决定区Y框蛋白2（SOX2），Lin28，Krüppel样因子（KLF）的表达；采用缺氧，血清剥夺和对5-氟尿嘧啶（5-FU）耐药实验检测WASL对肠癌细胞干性的影响。结果 WASL在肠癌组织中mRNA和蛋白均低表达（P<0.05）；WASL在肠癌组织中低表达，在HCT-116和SW-480中，WASL的表达明显弱于FHC细胞；成功构建稳定WASL表达的肠癌细胞系；Real-time PCR显示，WASL抑制了CDCs相关基因OCT4，SOX2，Lin28，Krüppel样因子（KLF）的表达；干细胞球形成和集落形成实验发现，WASL抑制肠癌细胞形成干细胞球形成和集落形成；缺氧实验和血清剥夺实验发现WASL上调后，能够抑制肠癌对缺氧和缺血清耐受，5-FU耐药实验显示，WASL也能抑制肠癌细胞对5-FU的耐受。结论 WASL蛋白在肠癌组织和肠癌细胞中低表达，并与肠癌的不良预后有关，WASL蛋白的过表达能够抑制肠癌细胞的干性。

Abstract

Objective To explore the expression of Wiskot-Aldrich syndrome-like（WASL）in colon cancer tissues and its function on colon cancer stemness. Methods The UALCAN database was used to analyze the expression of WASL in colon cancer tissues and its correlation among the gender, lymphnode stage and metastasis in colon cancer patients. Immunohistofluorescence and immunocyte fluorescence were used to verify the date from UALCAN. Western blotting was used to detect the expression and transfection efficiency of WASL in human normal colorectal mucosal cells (FHC cells) and colon cancer cells HCT-116, SW-480. Sphere formation assay and colony formation were used to detect the effect of WASL on the stemness of colon cancer. Hypoxia assay, serum deprivation, drug resistance assay were used to detect the stemness of colon cancer cells. Results WASL was lower expressed in colon cancer tissues (P<0.05). WASL was lower expressed in colon cancer tissues than peri-cancer tissues. WASL was obviously lower expressed in HCT-116 and SW-480 than FHC cells. WASL overexpression stable cells were constructed suscessfully in HCT-116 and SW-480.Real-time PCR showed WASL obviously down-regulated the expression of cancer stemcells(CSCs) related markers ［octamer binding transcription factor 4(OCT4), sex determining region Y-box 2(SOX2), Lin28, krüppel-like factor(KLF)］. Sphere formation assay and colony formation assay showed WASL could inhibited the sphere formation and colony formation. Hypoxia assay and serum deprivation showed WASL inhibited the resistance of cancer cells to hypoxia and serum deprivation. Drug resistance assay showed WASL inhibit the resistance of cancer cells to 5-fluorouracil(5-FU). Conclusion WASL is lower expressed in colon cancer tissues and cells, and is related with poor prognosis of colon cancer. Overexpression of WASL can inhibit the stemness of colon cancer cells.

导出引用

李宏丹杨成. WAS-like 蛋白在肠癌组织中低表达并抑制人结肠癌细胞的干性[J]. 解剖学报. 2022, 53(1): 50-59 https://doi.org/10.16098/j.issn.0529-1356.2022.01.007

LI Hong-dan YANG Cheng. Wiskot-Aldrich syndrome-like lower expressed in colon cancer tissues and inhibited the stemness of human colon cancer[J]. Acta Anatomica Sinica. 2022, 53(1): 50-59 https://doi.org/10.16098/j.issn.0529-1356.2022.01.007

中图分类号： R735.3

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