目的 探究芪珍胶囊对人结肠癌细胞HCT116及裸鼠移植瘤模型的作用及机制。方法 HCT116细胞给予芪珍胶囊进行干预，采用MTT法和流式细胞术检测细胞存活率和凋亡情况；采用Western blotting法考察芪珍胶囊对聚腺苷二磷酸核糖聚合酶（poly ADP-ribose polymerase，PARP）、B淋巴细胞瘤-2（B-cell lymphoma-2，Bcl-2）、半胱氨酸蛋白酶-3（Caspase-3）、Caspase-9、剪切型Caspase-9（cleaved Caspase-9）、哺乳动物雷帕霉素靶蛋白（mammalian target of rapamycin，mTOR）、磷酸化mTOR（p-mTOR）、腺苷酸活化蛋白激酶（adenosine monophosphate-activated protein kinase，AMPK）、p-AMPK、细胞外信号调节蛋白激酶（extraeellular signal regulated kinase，ERK）、p-ERK、p38和p-p38蛋白表达的影响；采用qRT-PCR法检测芪珍胶囊对程序性死亡分子配体-1（programmed death ligand 1，PD-L1）mRNA表达的影响。建立结肠癌裸鼠移植瘤模型，考察芪珍胶囊对裸鼠体质量、瘤体积和瘤质量的影响；采用ELISA法检测各组裸鼠血清中肿瘤坏死因子-α（tumor necrosis factor alpha，TNF-α）、白细胞介素-6（interleukin-6，IL-6）和IL-1β水平；采用qRT-PCR法检测各组裸鼠肿瘤组织核因子-κB（nuclear factor-κB，NF-κB）、TNF-α、IL-6、CD68、单核细胞趋化蛋白-1（monocyte chemoattractant protein-1，MCP-1）、诱导型一氧化氮合酶（inducible nitric oxide synthase，iNOS）和PD-L1 mRNA表达；采用Western blotting法考察各组裸鼠肿瘤组织PD-L1蛋白表达。以γ-干扰素（interferon-γ，IFN-γ）诱导HCT116细胞，考察芪珍胶囊对细胞存活率及PD-L1蛋白表达的影响。结果 芪珍胶囊显著抑制HCT116细胞存活率（P<0.01、0.001），显著促进细胞凋亡（P<0.01、0.001），显著下调细胞中Caspase-9、Caspase-3和Bcl-2蛋白表达水平（P<0.05、0.01），显著上调细胞中cleaved Caspase-9、cleaved PARP、p-mTOR/mTOR、p-AMPK/AMPK、p-ERK/ERK和p-p38/p38蛋白表达水平（P<0.05、0.01、0.001），显著下调细胞中PD-L1 mRNA表达水平（P<0.05）。结肠癌裸鼠移植瘤模型中，芪珍胶囊显著抑制移植瘤生长（P<0.05、0.01），降低氟尿嘧啶导致的裸鼠体质量减轻，显著降低血清中TNF-α、IL-6和IL-1β水平（P<0.05、0.01、0.001），显著降低肿瘤组织中NF-κB、TNF-α、IL-6、CD68、MCP-1、iNOS和PD-L1 mRNA表达水平（P<0.05），显著降低肿瘤组织中PD-L1蛋白表达水平（P<0.01）。芪珍胶囊显著抑制IFN-γ诱导的HCT116细胞中PD-L1蛋白表达水平（P<0.001）。结论 芪珍胶囊能够通过诱导细胞凋亡、抑制炎性反应及PD-L1表达等途径抑制结肠癌的发生发展，可能是一种潜在的免疫检查点抑制剂。

Objective To explore effect and mechanism of Qizhen Capsule (芪珍胶囊) on human colon cancer cell HCT116 and transplanted tumor models in nude mice. Methods HCT116 cells were treated with Qizhen Capsules for intervention. MTT and flow cytometry were used to detect cell survival and apoptosis; Western blotting was used to investigate the effects of Qizhen Capsules on poly ADP-ribose polymerase (PARP), B-cell lymphoma-2 (Bcl-2), Caspase-3, Caspase-9, cleaved Caspase-9, mTOR, phosphorylated mTOR (p-mTOR), adenosine monophosphate-activated protein kinase (AMPK), p-AMPK, extraeellular signal-regulated protein kinase (ERK), p-ERK, p38 and p-p38 expressions; qRT-PCR was used to detect the effect of Qizhen Capsule on programmed death ligand 1 (PD-L1) mRNA level. Transplanted tumor model of colon cancer in nude mice was established, effects of Qizhen Capsule on body weight, tumor volume and tumor weight of nude mice were investigated; ELISA was used to detect tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6) and IL-1β levels in serum of nude mice; qRT-PCR was used to detect nuclear factor-κB (NF-κB), TNF-α, IL-6, CD68, monocyte chemoattractant protein-1 (MCP-1), inducible nitric oxide synthase (iNOS) and PD-L1 mRNA levels in tumor; Western blotting was used to investigate PD-L1 expression in tumor tissues of nude mice in each group. IFN-γ was used to induce HCT116 cells to investigate the effect of Qizhen Capsule on cell survival rate and PD-L1 expression. Results Qizhen Capsule significantly inhibited the survival rate of HCT116 cells (P<0.01, 0.001), significantly promoted cell apoptosis (P<0.01, 0.001), significantly down-regulated expressions of Caspase-9, Caspase-3 and Bcl-2 in cells (P<0.05, 0.01), significantly up-regulated expressions of cleaved Caspase-9, cleaved PARP, p-mTOR/mTOR, p-AMPK/AMPK, p-ERK/ERK and p-p38/p38 in cells (P<0.05, 0.01, 0.001), significantly down-regulated PD-L1 mRNA level in cells (P<0.05). In transplanted tumor model of colon cancer in nude mice, Qizhen Capsules significantly inhibited the growth of transplanted tumors (P<0.05, 0.01), reduced the body weight of nude mice caused by fluorouracil, significantly reduced TNF-α, IL-6 and IL-1β levels in serum (P<0.05, 0.01, 0.001), significantly reduced mRNA levels of NF-κB, TNF-α, IL-6, CD68, MCP-1, iNOS and PD-L1 in tumor tissues (P<0.05), significantly reduced PD-L1 expression in tumor tissues (P<0.01). Qizhen Capsules significantly inhibited PD-L1 expression in HCT116 cells induced by IFN-γ (P<0.001). Conclusion Qizhen Capsules can inhibit the occurrence and development of colon cancer by inducing cell apoptosis, inhibiting inflammation and PD-L1 expression, which may be a potential immune checkpoint inhibitor.

R285.5

国家自然科学基金面上项目（81973521）；浙江省科技厅一流学科药学学科基金资助项目（2018-2020）