文章摘要
王荣国.高迁移率族蛋白B1调节巨噬细胞极化对喉鳞状细胞癌细胞增殖和Akt/Wnt/β-catenin信号的影响[J].安徽医药,待发表.
高迁移率族蛋白B1调节巨噬细胞极化对喉鳞状细胞癌细胞增殖和Akt/Wnt/β-catenin信号的影响
投稿时间:2024-05-09  录用日期:2024-05-31
DOI:
中文关键词: 高迁移率族蛋白B1  巨噬细胞极化  喉鳞状细胞癌  增殖  Akt/Wnt/β-catenin信号通路
英文关键词: 
基金项目:2021年河北医学科学研究课题计划:20210800
作者单位地址
王荣国* 河北省人民医院 河北省石家庄市和平西路348号
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中文摘要:
      目的 探究高迁移率族蛋白B1(high mobility group protein, HMGB1)对巨噬细胞极化及喉鳞状细胞癌(laryngeal squamous cell carcinoma, LSCC)细胞增殖的影响及可能的机制。方法 生物信息学和免疫组化染色分析HMGB1在LSCC中的表达及与巨噬细胞的相关性。佛波酯诱导THP-1细胞分化为M0巨噬细胞,IL-4和IL-13刺激M0巨噬细胞向M2型极化。细胞骨架染色、免疫荧光染色和流式细胞术分析巨噬细胞分化和极化。通过RNA干涉技术建立沉默HMGB1的LSCC和M2巨噬细胞。收集LSCC细胞条件培养基(conditioned medium, CM)培养M2巨噬细胞,收集巨噬细胞CM培养LSCC细胞。qRT-PCR和Western blot检测mRNA和蛋白的表达。CCK-8、EdU和克隆形成实验检测LSCC细胞活力和增殖能力。结果 HMGB1在LSCC中高表达且与M2巨噬细胞浸润正相关。敲低HMGB1抑制LSCC细胞活力和增殖能力。与对照组或CM-siNC组相比,CM-siHMGB1组巨噬细胞体积变小,呈圆形且光滑,与M0巨噬细胞相似。M0巨噬细胞经siHMGB1转染的LSCC细胞CM处理后,其M2型标志物CD163、CD206、IL-10、TGF-β1和Arg-1的表达显著减少。此外,与CM-THP-1和CM-M0组相比,CM-M2处理的LSCC细胞活力和增殖能力明显增加;而经CM-M2-siHMGB1处理后,LSCC细胞活力和增殖能力低于CM-M0和CM-M2-siNC培养的细胞。经CM-M2和CM-M2-siNC培养的LSCC细胞中Akt和GSK-3β的磷酸化水平和β-catenin蛋白表达及TCF/LEF转录活性较CM-M0培养组明显升高;而CM-M2-siHMGB1处理的LSCC细胞中p-Akt、p-GSK-3β和β-catenin的表达及TCF/LEF转录活性显著低于CM-M2-siNC处理组。结论 HMGB1在LSCC中高表达且通过诱导巨噬细胞M2型极化促进LSCC细胞增殖并激活Akt/Wnt/β-catenin信号通路。
英文摘要:
      Objective To explore the effects of high mobility group protein (HMGB1) on macrophage polarization and laryngeal squamous cell carcinoma (LSCC) cell proliferation and the possible mechanisms. Methods Bioinformatics and immunohistochemical staining were used to analyze HMGB1 expression in LSCC and its correlation with macrophages. Phorbol myristate acetate was used to induce THP-1 cells to differentiate into M0 macrophages, and IL-4 and IL-13 was used stimulate M0 macrophages to polarize M2-type. Macrophage differentiation and polarization were analyzed by cytoskeleton staining, immunofluorescence staining and flow cytometry. HMGB1-silenced LSCC and M2 macrophages were established by RNA interference technique. M2 macrophages were cultured in conditioned medium (CM) of LSCC cells, and macrophage CMs were collected for culture of LSCC cells. qRT-PCR and Western blot assays were performed to measure mRNA and protein expressions. CCK-8, EdU and colony formation assay were conducted to assess the viability and proliferation of LSCC cells. Result HMGB1 was highly expressed in LSCC and positively correlated with M2 macrophages. HMGB1 knockdown inhibited the viability and proliferation of LSCC cells. Compared with the control or CM-siNC group, the macrophages in CM-siHMGB1 group were smaller, round and smooth, similar to M0 macrophages. The expressions of M2 markers CD163, CD206, IL-10, TGF-β1 and Arg-1 were significantly reduced in M0 macrophages treatment with CM from siHMGB1-transfected LSCC cells. In addition, compared with CM-THP-1 and CM-M0 groups, the viability and proliferation of LSCC cells with CM-M2 treatment were considerably increased. However, after CM-M2-siHMGB1 culture, the viability and proliferation of LSCC cells were significantly lower than those with CM-M0 and CM-M2-siNC culture. The phosphorylation of Akt and GSK-3β and β-catenin protein expression as well as TCF/LEF transcriptional activity in LSCC cells cultured by CM-M2 and CM-M2-siNC were remarkably higher than those in CM-M0-cultured group, while the levels of p-Akt, p-GSK-3β and β-catenin and the transcriptional activity of TCF/LEF in LSCC cells treated with CM-M2-siHMGB1 were markedly lower than those with CM-M2-siNC treatment. Conclusion HMGB1 was highly expressed in LSCC and promoted LSCC cell proliferation and activated the Akt/Wnt/β-catenin signaling pathway by inducing M2-type polarization of macrophages.
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