目的 探讨柴胡皂苷D（SSD）对食管鳞状细胞癌（ESCC）Eca109细胞生长和缺氧诱导因子2α（HIF-2α）/核转录因子-κB p65（NF-κB p65）通路的影响。方法 用不同浓度SSD（0、3、6、9、12、15、18、21、24 μmol/L）处理人正常食管鳞状上皮细胞Het-1a和ESCC细胞Eca109筛选SSD的实验浓度。将Eca109细胞分为对照（NC）组、HIF-2α抑制剂（PT2385）组、SSD低、高浓度（SSD-L、SSD-H）组、SSD-H+空载慢病毒（LV-NC）组、SSD-H+HIF-2α重组慢病毒（LV-HIF-2α）组。MTT法检测细胞活力；流式细胞术检测细胞凋亡；ELISA检测细胞培养液中TNF-α、IL-1β、IL-8水平；Western blot检测细胞增殖、凋亡和HIF-2α/NF-κB p65通路相关蛋白表达；建立Eca109细胞荷瘤裸鼠模型验证SSD对Eca109细胞体内生长的影响。结果 选择6、12 μmol/L SSD为实验浓度。与NC组比较，PT2385组、SSD-L、SSD-H组细胞活力、细胞培养液中TNF-α、IL-1β、IL-8水平、PCNA、Bcl-2、HIF-2α、p-NF-κB p65/NF-κB p65蛋白水平、裸鼠皮下移植瘤体积和肿瘤重量、血清TNF-α、IL-1β、IL-8水平以及HIF-2α、NF-κB p65蛋白阳性表达降低，细胞凋亡率、Bax、Cleaved-caspase3蛋白水平升高（P＜0.05或P＜0.01）；过表达HIF-2α可显著削弱SSD对Eca109细胞增殖和裸鼠皮下移植瘤生长的抑制作用（P＜0.05或P＜0.01）。结论 SSD可抑制Eca109细胞增殖，诱导细胞凋亡，并抑制Eca109细胞荷瘤裸鼠皮下移植瘤生长，其作用可能与抑制HIF-2α/NF-κB p65通路的活化有关。

Objective To investigate the effects of saikosaponin D (SSD) on the growth of esophageal squamous cell carcinoma (ESCC) Eca109 cells and the hypoxia-inducible transcription factor-2α (HIF-2α)/nuclear factor-κB p65 (NF-κB p65) pathway. Methods Different concentrations of SSD (0, 3, 6, 9, 12, 15, 18, 21, 24 μmol/L) were used to treat Het-1a normal esophageal squamous epithelial cells and Eca109 ESCC cells to screen the experimental concentration of SSD. Eca109 cells were separated into control (NC) group, HIF-2α inhibitor (PT2385) group, SSD low and high concentration (SSD-L, SSD-H) groups, SSD-H+empty lentivirus (LV-NC) group, and SSD-H+HIF-2α recombinant lentivirus (LV-HIF-2α) group. MTT assay was applied to detect cell viability. Flow cytometry was applied to detect cell apoptosis. ELISA was applied to detect the levels of TNF-α, IL-1β, and IL-8 in cell culture media. Western blot was applied to detect cell proliferation, apoptosis, and protein expression related to the HIF-2α/NF-κB p65 pathway. The Eca109 cell tumor bearing nude mouse model was established to verify the effect of SSD on the in vivo growth of Eca109 cells. Results 6 and 12 μmol/L SSD were selected as experimental concentrations. Compared with the NC group, the cell viability, TNF-α, IL-1β, IL-8 levels in cell culture medium, PCNA, Bcl-2, HIF-2α, p-NF-κB p65/NF-κB p65 protein levels, subcutaneous transplant tumor volume and weight in nude mice, serum TNF-α, IL-1β, IL-8 levels, and positive expression of HIF-2α and NF-κB p65 proteins in the PT2385 group, SSD-L group, and SSD-H group were lower, the apoptosis rate, Bax, and Cleaved-caspase3 protein levels were higher (P<0.05 or P<0.01). Overexpression of HIF-2α prominently weakened the inhibitory effects of SSD on the proliferation of Eca109 cells and the growth of subcutaneous transplanted tumors in nude mice (P<0.05 or P<0.01). Conclusion SSD can inhibit the proliferation of Eca109 cells, induce cell apoptosis, and suppress the growth of subcutaneous transplanted tumors of Eca109 cells in nude mice. Its effect may be related to the inhibition of the activation of the HIF-2α/NF-κB p65 pathway.

河南省中医药科学研究专项课题（2024ZYZD03）