CTCF通过SALL3/DNMT3A轴对肝癌干细胞自我更新和免疫逃逸的影响*

doi:10.3870/j.issn.1672-0741.25.11.015

摘要/Abstract

摘要： 目的探讨CCCTC结合因子(CTCF)通过SALL3/DNMT3A轴对肝癌干细胞自我更新和免疫逃逸的影响。方法将tLyP-1-EV-sh-CTCF转染至肝癌干细胞LCSCs中,为tLyP-1-EV-sh-CTCF-LCSCs组;将WZB117抑制剂与LCSCs共孵育,为WZB117抑制剂-LCSCs组;使用qRT-PCR检测肝癌细胞系HCCLM3细胞与LCSCs细胞中CTCF的mRNA表达水平,以及LCSCs组、tLyP-1-EV-sh-CTCF-LCSCs组和WZB117抑制剂-LCSCs组糖酵解标志物(HK2与PFKM)及细胞干性标志物(FBP1与PCK1)的mRNA的表达水平;使用细胞迁移与细胞克隆实验评价sh-CTCF与糖酵解抑制对LCSCs组、tLyP-1-EV-sh-CTCF-LCSCs组和WZB117抑制剂-LCSCs组细胞干性的影响;使用qRT-PCR检测sh-CTCF敲减对SALL3、DNMT3A表达的影响,以及过表达SALL3对DNMT3A的影响。结果与对照组(未转染)相比,tLyP-1-EV-sh-CTCF组细胞中CTCF mRNA与蛋白的表达量均较低(均P<0.05),PFKM与PCK1的mRNA表达降低(均P＜0.05),HK2与FBP1呈下降趋势(均P<0.05);WZB117抑制剂处理后,与转染sh-CTCF组比较,PFKM与PCK1的mRNA表达降低(均P＜0.05),HK2与FBP1呈下降趋势(均P＞0.05);过表达SALL3可抑制PD-L1表达并促进MHC-Ⅰ表达,抑制LCSCs自我更新并下调免疫逃逸相关分子表型,同时伴随糖酵解相关分子表达下调(均P＜0.05);tLyP-1-EV-sh-CTCF转染可上调SALL3表达、下调DNMT3A表达,并伴随糖酵解相关分子表达下调(均P＜0.05)。结论 sh-CTCF调控SALL3/DNMT3A轴与糖酵解相关标记物下调相一致,并伴随肝癌干细胞自我更新及免疫逃逸相关分子表型的减弱。

关键词: CTCF, SALL3/DNMT3A轴, 肝癌干细胞, 自我更新, 免疫逃逸

Abstract: Objective To investigate the effects of CTCF on self-renewal and immune escape of liver cancer stem cells through the SALL3/DNMT3A axis.Methods HCCLM3 cells and liver cancer stem cells(LCSCs)were assigned to the HCCLM3 group and LCSC group,respectively.LCSCs transfected with sh-CTCF delivered by tLyP-1-modified extracellular vesicles were defined as the tLyP-1-EV-sh-CTCF-LCSC group.LCSCs treated with the WZB117 inhibitor were defined as the WZB117 inhibitor-LCSC group.Quantitative reverse transcription PCR(qRT-PCR)was used to detect CTCF mRNA expression in the HCCLM3 and LCSC groups,as well as the mRNA expression levels of glycolysis-related markers(HK2 and PFKM)and stemness-related markers(FBP1 and PCK1)in the LCSC,tLyP-1-EV-sh-CTCF-LCSC,and WZB117 inhibitor-LCSC groups.Cell migration and colony formation assays were performed to evaluate the effects of CTCF knockdown and glycolysis inhibition on LCSC stemness.qRT-PCR was further used to assess the effects of CTCF knockdown on SALL3 and DNMT3A expression and the effect of SALL3 overexpression on DNMT3A expression.Results Compared with the untreated control group,cells transfected with tLyP-1-EV-sh-CTCF showed significantly reduced CTCF mRNA and protein expression levels(both P<0.05).Compared with the LCSC group,the tLyP-1-EV-sh-CTCF-LCSC group showed decreased mRNA expression of PFKM and PCK1(both P<0.05),while HK2 and FBP1 showed downward trends without statistical significance(both P>0.05).After WZB117 treatment,the mRNA expression levels of PFKM and PCK1 were also reduced(both P<0.05),whereas HK2 and FBP1 showed decreasing trends without statistical significance(both P>0.05).SALL3 overexpression inhibited PD-L1 expression,promoted MHC-I expression,suppressed LCSC self-renewal,and reduced immune escape-related molecular phenotypes,accompanied by downregulation of glycolysis-related molecules(all P<0.05).Transfection with tLyP-1-EV-sh-CTCF upregulated SALL3 expression and downregulated DNMT3A expression,accompanied by decreased expression of glycolysis-related molecules(all P<0.05).Conclusion CTCF knockdown is associated with downregulation of glycolysis-related markers through modulation of the SALL3/DNMT3A axis,accompanied by attenuated self-renewal and immune escape-related molecular phenotypes in liver cancer stem cells.

Key words: CTCF, SALL3/DNMT3A axis, hepatocellular carcinoma stem cell, self-renewal, immune evasion

中图分类号:

R735.7

朱恒, 吕慧娟, 王永瑞, 范宝霞, 韩莉, 张运贵. CTCF通过SALL3/DNMT3A轴对肝癌干细胞自我更新和免疫逃逸的影响^*[J]. 华中科技大学学报（医学版）, 2026, 55(4): 496-503.

Zhu Heng, Lyu Huijuan, Wang Yongrui, et al. CTCF Affects Self-renewal and Immune Escape of Hepatocellular Carcinoma Stem Cells Through SALL3/DNMT3A Axis[J]. Acta Medicinae Universitatis Scientiae et Technologiae Huazhong, 2026, 55(4): 496-503.

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CTCF通过SALL3/DNMT3A轴对肝癌干细胞自我更新和免疫逃逸的影响^*

CTCF Affects Self-renewal and Immune Escape of Hepatocellular Carcinoma Stem Cells Through SALL3/DNMT3A Axis

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