Chen Yan, Gao Bo, Pan Yun, Wang Qingqing, Zhang Qiurong
College of Clinical Medicine, Dali University, Dali, Yunnan, 671000, P.R. China.
Department of Pathology, The First Affiliated Hospital of Dali University, Jiashibo Road 32, Dali, Yunnan, 671000, P.R. China.
Ann Hematol. 2024 Dec;103(12):5817-5833. doi: 10.1007/s00277-024-06062-7. Epub 2024 Nov 4.
MiR-525-5p functions as an oncomiRNA or tumor suppressor, and has been reported in various cancer types, including laryngeal squamous cell carcinoma, glioma, breast cancer, and cervical cancer. However, the biological functions and precise mechanisms of miR-525-5p remain unclarified in Burkitt's lymphoma (BL). This study aimed to explore the roles of miR-525-5p in BL, with the goal of ascertaining its regulatory effects on the nuclear factor-kappaB (NF-κB) signaling pathway by targeting Myeloid differentiation factor 88 (MyD88). The expression levels of miR-525-5p and MyD88 were measured by quantitative real-time PCR and immunohistochemical staining, respectively. The effects of miR-525-5p overexpression on BL cell proliferation, colony-forming, and migration were evaluated by cell counting kit-8, soft agar colony-forming, and transwell assays, while cell cycle and cell apoptosis were analyzed by flow cytometry. Possible interactions between miR-525-5p and MyD88 was examined via luciferase reporter assay. The expression of MyD88 and NF-κB signaling pathway-related proteins, including p65, p-p65, IκBa, and p-ΙκBa was determined by western blotting. BL cells overexpressing miR-525-5p were treated with phorbol 12-myristate 13-acetate (PMA), and Hoechst 33258 staining and Calcein AM/EthD-I staining were used to analyze the changes in chemotherapy sensitivity of BL cells to doxorubicin (DOX). Compared with reactive lymphoid hyperplasia, miR-525-5p was dramatically downregulated in BL tissues, while the rate of MyD88 protein positivity was significantly increased. Upregulation of miR-525-5p suppressed cell proliferation, colony-forming, and migration, induced cell cycle arrest and apoptosis, and enhanced the chemosensitivity to DOX in BL cells. MiR-525-5p targeted MyD88 to inhibit the activation of NF-κB signaling pathway. PMA treatment reactivated the NF-κB pathway and reversed apoptosis mediated by miR-525-5p overexpression. These findings revealed that miR-525-5p acts as a tumor suppressor, targeting MyD88 to modulate proliferation, cell cycle progression, and apoptosis in BL cells by regulation of NF-κB signaling pathway.
微小RNA-525-5p(miR-525-5p)可作为癌基因或肿瘤抑制因子发挥作用,并且已在多种癌症类型中被报道,包括喉鳞状细胞癌、神经胶质瘤、乳腺癌和宫颈癌。然而,miR-525-5p在伯基特淋巴瘤(BL)中的生物学功能及确切机制仍不清楚。本研究旨在探讨miR-525-5p在BL中的作用,目标是通过靶向髓样分化因子88(MyD88)来确定其对核因子-κB(NF-κB)信号通路的调控作用。分别采用定量实时聚合酶链反应(qRT-PCR)和免疫组织化学染色检测miR-525-5p和MyD88的表达水平。通过细胞计数试剂盒-8(CCK-8)、软琼脂集落形成和Transwell实验评估miR-525-5p过表达对BL细胞增殖、集落形成和迁移的影响,同时通过流式细胞术分析细胞周期和细胞凋亡情况。通过荧光素酶报告基因实验检测miR-525-5p与MyD88之间可能的相互作用。采用蛋白质免疫印迹法检测MyD88及NF-κB信号通路相关蛋白(包括p65、磷酸化p65、IκBa和磷酸化IκBa)的表达。用佛波酯12-肉豆蔻酸酯13-乙酸酯(PMA)处理过表达miR-525-5p的BL细胞,并采用Hoechst 33258染色和钙黄绿素AM/碘化丙啶(EthD-I)染色分析BL细胞对多柔比星(DOX)化疗敏感性的变化。与反应性淋巴组织增生相比,miR-525-5p在BL组织中显著下调,而MyD88蛋白阳性率显著升高。miR-525-5p上调可抑制BL细胞的增殖、集落形成和迁移,诱导细胞周期阻滞和凋亡,并增强对DOX的化疗敏感性。miR-525-5p靶向MyD88以抑制NF-κB信号通路的激活。PMA处理可重新激活NF-κB通路,并逆转miR-525-5p过表达介导的细胞凋亡。这些研究结果表明,miR-525-5p作为一种肿瘤抑制因子,通过靶向MyD88调控NF-κB信号通路,从而调节BL细胞的增殖、细胞周期进程和凋亡。