目的 鉴定三叶青醇提物（ethanol extract of Tetrastigma hemsleyanum，SYQ）入血成分并探讨其抗4T1乳腺癌的作用机制。方法 采用超高效液相色谱-四极杆-飞行时间串联质谱（ultra-high performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry，UHPLC-Q-TOF-MS/MS）分析SYQ的入血成分；通过MTT法、克隆形成与划痕实验检测SYQ含药血清对4T1乳腺癌细胞增殖和迁移的影响。构建4T1荷瘤小鼠模型，设置对照组、模型组、环磷酰胺（20 mg/kg）组和SYQ低、中、高剂量（5、10、20 g/kg）组，给药干预后，ELISA检测血清中白细胞介素-6（interleukin-6，IL-6）和肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）水平；苏木素-伊红（hematoxylin-eosin，HE）染色观察肿瘤组织病理变化；免疫组化检测肿瘤组织CD68和Ki67表达；Western blotting法检测肿瘤组织中TNF-α/蛋白激酶B（protein kinase B，Akt）/信号传导与转录激活因子3（signal transducer and activator of transcription 3，STAT3）信号轴及凋亡相关蛋白表达。结果 UHPLC-Q-TOF-MS/MS鉴定出19种入血成分，主要包括黄酮类（芦丁、橘皮素、水仙苷）以及苯丙素类和其他苷类化合物（连翘苷E、达伦苷B）。体外实验结果显示，SYQ含药血清显著抑制4T1细胞增殖、迁移及克隆形成（P＜0.01、0.001）。体内实验结果显示，SYQ显著抑制荷瘤小鼠肿瘤体积增长（P＜0.001），高剂量组抑瘤率达25.95%；降低血清中IL-6和TNF-α水平（P＜0.05、0.001），诱导肿瘤组织坏死并显著降低CD68和Ki67表达（P＜0.001），同时下调肿瘤组织中TNF-α、p-Akt/Akt、p-STAT3/STAT3及B细胞淋巴瘤-2（B-cell lymphoma-2，Bcl-2）表达水平（P＜0.05、0.01、0.001）。结论 SYQ可通过降低炎症因子水平、抑制肿瘤细胞增殖及诱导肿瘤组织坏死，从而发挥抗乳腺癌作用，其机制可能与抑制TNF-α/Akt/STAT3信号轴有关。

Objective To identify the blood-absorbed constituents of ethanol extract of Tetrastigma hemsleyanum (SYQ) and investigate its anti-breast cancer activity and underlying mechanisms. Methods Blood-absorbed components of SYQ were identified using ultra-high performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry. The effects of SYQ drug containing serum on proliferation and migration of 4T1 cells were evaluated by MTT, colony formation and wound-healing assays. A 4T1 tumor-bearing mouse model was established, control group, model group, cyclophosphamide (20 mg/kg) group, SYQ low-, medium-, and high-dose (5, 10, 20 g/kg) groups were set up. After drug intervention, ELISA was used to detect the levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in serum. Hematoxylin-eosin (HE) staining was used to observe pathological changes in tumor tissue. Immunohistochemistry was used to detect CD68 and Ki67 expressions in tumor tissue. Western blotting was used to detect the expressions of TNF-α/protein kinase B (Akt)/signal transducer and activator of transcription 3 (STAT3) signaling axis and apoptosis related proteins in tumor tissues. Results A total of 19 blood components were identified by UHPLC-Q-TOF-MS/MS, mainly including flavonoids (rutin, tangeretin, narcissoside), phenylpropanoids, and other glycoside compounds (forsythoside E, darendoside B). The in vitro experimental results showed that SYQ drug containing serum significantly inhibited the proliferation, migration and colony formation of 4T1 cells (P < 0.01, 0.001). The in vivo experimental results showed that SYQ significantly inhibited tumor volume growth in tumor bearing mice (P < 0.001), with a tumor inhibition rate of 25.95% in high-dose group. SYQ reduced the levels of IL-6 and TNF-α in serum (P < 0.05, 0.001), induced tumor tissue necrosis and significantly reduced the expressions of CD68 and Ki67 (P < 0.001), while downregulated the expression levels of TNF-α, p-Akt/Akt, p-STAT3/STAT3 and B-cell lymphoma-2 (Bcl-2) in tumor tissue (P < 0.05, 0.01, 0.001). Conclusion SYQ could exerts anti-breast cancer effects by reducing the levels of inflammatory factors, inhibiting tumor cell proliferation and inducing tumor tissue necrosis. The mechanism may be related to the inhibition of TNF-α/Akt/STAT3 signaling axis.

R285.5

江苏省人民医院专项发展基金（ZXFZ2026017）；南京中医药大学中药学一级重点学科开放项目（ZYXPY2024-008）