Objectives: To explore the enhanced sensitization effect of peritumoral electroacupuncture (PEA) on doxorubicin (DOX) chemotherapy in mice with triple-negative breast cancer (TNBC).
Methods: Eighteen female Balb/c mice were randomly divided into the model, DOX, and EA+DOX groups, with 6 mice in each group. TNBC cells were injected into the mammary fat pad of mice to establish the breast cancer-bearing mice model. Mice of the DOX and EA+DOX groups were injected intraperitoneally with DOX solution at 4 mg/kg once a week for 4 weeks. For mice of the EA+DOX group, 4 filiform needles were inserted surrounding tumor tissues, followed by giving the mice with EA (1 mA, 2 Hz/100 Hz) stimulation for 3 min, once a week for 4 weeks. During the intervention, the tumor volume was measured every two days, and at the end of intervention, the weight of the tumor tissue was measured. The expression of Ki67 and proliferating cell nuclear antigen (PCNA) in tumor tissues was detected by immunohistochemistry. The protein expression levels of iron redox protein 1 (FDX1), lipoic acid synthase (LIAS), aconitase 2 (ACO2) and NADH：ubiquinone oxidoreductase core subunit V1 (NDUFV1), dihydrolipoamide S-acetyltransferase(DLAT), dihydrolipoyl succinyltransferase(DLST) in tumor tissues were detected by Western blot. The content of copper ions, pyruvic acid and α-ketoglutaric acid, succinic acid in tumor tissues was detected by copper ion kit, and the content of reactive oxygen species (ROS) in tumor tissues of mice was detected by fluorescence method.
Results: Compared with the model group, the tumor volume and weight, the expression levels of Ki67 and PCNA in tumor tissues were decreased (P<0.05, P<0.01) in the DOX and EA+DOX groups. The improvement of the above indicators was more significant in the EA+DOX group than that in the DOX group (P<0.05, P<0.01). Compared with the model and DOX groups, the contents of ROS, copper ions, pyruvic acid and α-ketoglutaric acid were increased (P<0.05, P<0.01), while the contents of succinic acid, and the expressions of FDX1, LIAS, ACO2 and NDUFV1, DLAT, DLST proteins were decreased (P<0.01) in the EA+DOX group.
Conclusions: The combination of PEA and DOX can effectively control the growth rate of tumors in mice with TNBC, which may contribute to its function in enhancing the chemotherapy efficacy of DOX on TNBC by promoting cuproptosis.
目的: 观察瘤周围刺联合阿霉素对三阴性乳腺癌（TNBC）荷瘤小鼠铜死亡的影响，探讨瘤周围刺增强阿霉素治疗TNBC敏感性的机制。方法: 雌性Balb/c小鼠随机分为模型组、阿霉素组和电针-阿霉素组，每组6只。于小鼠乳腺脂肪垫注射4T1细胞，构建移植瘤模型。阿霉素组和电针-阿霉素组均按照4 mg/kg的剂量腹腔注射阿霉素，每周1次，治疗4周；电针-阿霉素组在每次注射阿霉素后进行瘤周围刺电针治疗，每次3 min，每周1次，治疗4周。治疗期间，每隔2 d测量肿瘤体积，并在治疗结束后分离肿瘤测定质量。免疫组织化学法检测小鼠瘤体组织中Ki67、增殖细胞核抗原（PCNA）的表达，Western blot法检测小鼠瘤体组织中铁氧化还原蛋白1（FDX1）、硫辛酰合酶（LIAS）、乌头酸酶2（ACO2）、NADH：泛醌氧化还原酶核心亚基V1（NDUFV1）、二氢硫辛酰胺S-乙酰转移酶（DLAT）、二氢硫辛酰琥珀酰转移酶（DLST）共同的蛋白表达水平，比色法检测肿瘤组织中的Cu2+、丙酮酸、α-酮戊二酸和琥珀酸含量，荧光法检测小鼠瘤体组织中活性氧（ROS）相对荧光强度。结果: 与同时点模型组相比，阿霉素组和电针-阿霉素组小鼠的肿瘤体积和质量均减小（P<0.01，P<0.05），瘤体中Ki67和PCNA的表达水平降低（P<0.01），而电针-阿霉素组较阿霉素组变化更显著（P<0.01，P<0.05）。与模型组和阿霉素组比较，电针-阿霉素组瘤体中ROS和Cu2+含量升高（P<0.01），丙酮酸和α-酮戊二酸的含量升高（P<0.01），琥珀酸含量降低（P<0.01），FDX1、LIAS、ACO2、NDUFV1、DLAT和DLST蛋白表达降低（P<0.01）。结论: 瘤周围刺联合阿霉素可有效控制肿瘤生长速度，可能是通过影响铜死亡的方式增强了阿霉素对TNBC的化疗疗效。.
Keywords: Cuproptosis; Doxorubin; Electroacupuncture; Triple-negative breast cancer.