肿瘤相关巨噬细胞通过FOXM1/Wnt/β-catenin通路影响乳腺癌内分泌抵抗

Tumor-associated macrophages affect breast cancer endocrine resistance through FOXM1/Wnt/β-catenin pathway

目的探讨肿瘤相关巨噬细胞(tumor associatedmacrophages,TAMs)通过叉头框M1(forkhead boxM1,FOXM1)/Wnt/β-catenin通路影响乳腺癌内分泌抵抗。方法培养他莫昔芬耐药的乳腺癌细胞,将THP-1细胞诱导成巨噬细胞(M),并进一步诱导成为肿瘤相关巨噬细胞(tumor-associated macrophages,TAMs)。MΦ在MCF-7细胞的条件培养基(conditionedmedium,CM)中培养24h后被定义为MS细胞,MΦ在MCF-7R细胞的CM中培养24h后被定义为MR细胞,MCF-7细胞在巨噬细胞的CM中培养24h后被定义为MCF-7(MΦ)细胞,MCF-7细胞在MS细胞的CM中培养24h后被定义为MCF-7(MS)细胞,MCF-7细胞在MR细胞的CM中培养24h后被定义为MCF-7(MR)细胞。借助CCK8与Transwell实验检测细胞活性与侵袭能力。qRT-PCR与Westernblot检测各组细胞中CD163、Wnt1、β-catenin、FOXM1蛋白水平。结果与MS(mRNA:1.49±0.12)(蛋白:1.15±0.12)相比,MR(mRNA:2.33±0.16)(蛋白:1.52±0.11)中CD163表达更高(t=7.28,P=0.002)(t=3.94,P=0.017),表明他莫昔芬耐药的乳腺癌细胞能诱导更多MΦ极化成TAMs。TAMs增加乳腺癌细胞中FOXM1的表达,FOXM1进一步促进Wnt/β-catenin通路激活。与MCF-7(MΦ)组细胞相比,MCF-7(MS)和MCF-7(MR)组细胞的活性与侵袭增强,MCF-7(MR)组细胞增幅最显著。较MCF-7(MΦ)组细胞,MCF-7(MS)与MCF-7(MR)组细胞中Wnt1、β-catenin、FOXM1水平显著升高,极化成最多TAMs的MCF-7(MR)组细胞中Wnt1、β-catenin、FOXM1水平最高。较MCF-7组,MCF-7(MR)组与MCF-7+pcDNA-FOXM1组中Wnt1、β-catenin水平均升高,细胞活性与侵袭能力增强,较MCF-7(MR)组,MCF-7(MR)+si-FOXM1组中Wntl、β-catenin水平降低,细胞活性与侵袭能力减弱。结论TAMs通过上调FOXM1并激活Wnt/β-catenin促进乳腺癌细胞内分泌抵抗.

Objective To explore the influence of tumor-associated macrophages(TAMs)on endocrine resistance in breast cancer through the forkhead box M1(FOXM1)/Wnt/β-catenin pathway.Methods Tamoxifen-resistant breast cancer cells were cultured,THP-1 cells were induced into macrophages(MQ),and further induced into TAMs.After being cultured in the conditioned medium(CM)of MCF-7 cells for 24 hours,MQ were defined as MS cells.After being cultured in the CM of MCF-7R cells for 24 hours,MQ were defined as MR cells.MCF-7 cells,after being cultured in the CM of macrophages for 24 hours,were defined as MCF-7(MO)cells.MCF-7 cells,after being cultured in the CM of MS cells for 24 hours,were defined as MCF-7(MS)cells.MCF-7 cells,af-ter being cultured in the CM of MR cells for 24 hours,were defined as MCF-7(MR)cells.Cell viability and invasion ability were evaluated using CCK-8 and Transwell assays.The protein levels of CD163,Wntl,β-catenin,and FOXM1 in different groups were examined by qRT-PCR and Western blot.Results Compared to the MS group(mRNA:1.49±0.12,protein:1.15±0.12),CD163 expression was higher in the MR group(mRNA:2.33±0.16,protein 1.52±0.11)(t=7.28,P=0.002)(t=3.94,P=0.017),indicating that tamoxifen-resistant breast cancer cells can induce polarization of more MQ into TAMs.TAMs increased the expression of FOXM1 in breast cancer cells,which further activated the Wnt/β-catenin pathway.Compared to the MCF-7(MQ)group,the MCF-7(MS)and MCF-7(MR)groups showed enhanced cell viability and invasion,with the most significant increase observed in the MCF-7(MR)group.Compared with MCF-7(MO)cells,the levels of Wntl,β-catenin,and FOXM1 in MCF-7(MS)and MCF-7(MR)cells were significantly increased,with the highest levels observed in the MCF-7(MR)group with the most TAM polarization.Compared to the MCF-7 group,both the MCF-7(MR)and MCF-7+pcDNAFOXM1 groups showed increased levels of Wntl andβ-catenin,enhanced cell viability and invasion.Compared to the MCF-7(MR)group,the MCF-7(MR)+si-FOXM1 group showed reduced levels of Wntl andβ-catenin,weakened cell viability and invasion.Conclusion TAMs promote endocrine resistance in breast cancer by upregulating FOXM1 and activating the Wnt/β-catenin pathway.