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目的 探讨牛黄?麝香联合使用对人肝癌细胞SMMC-7721及裸鼠肝癌皮下移植瘤的影响,探讨其抗肝癌机制.方法 将牛黄?麝香联合应用于体内、体外两种肝癌模型.体外培养人肝癌SMMC-7721细胞系,实验分为牛黄?麝香处理组和对照组,采用不同浓度(生药量0.625、1.25、2.5、5 mg/mL)牛黄?麝香萃取液进行干预,细胞增殖及毒性检测(CCK-8)实验检测肝癌细胞增殖能力,伤口愈合实验检测肝癌细胞迁移能力.制备人肝癌细胞裸鼠皮下移植瘤模型,选择18只5周龄SPF级雄性BALB/c裸鼠随机分为模型组(0.9％ 生理盐水0.2 mL/d),牛黄?麝香组[牛黄45.5 mg/(kg·d)+麝香13 mg/(kg·d)]与顺铂组(DDP,每周腹腔注射一次5 mg/kg),每组6只,各组裸鼠均给药14天.观察裸鼠皮下移植瘤瘤体体积及质量变化,采用实时荧光定量PCR(RT-PCR)和蛋白免疫印迹法(Western blot)检测肝癌裸鼠皮下移植瘤模型中磷脂酰肌醇-3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白(PI3K/AKT/mTOR)通路、凋亡相关因子p70 S6 Kinase(S6K)、Bax、Bcl-2、cas-pase-3和caspase-9的表达.原位末端转移酶标记技术(TUNEL)用于凋亡细胞的定量分析.结果 CCK-8实验表明,牛黄?麝香联合使用抑制肝癌细胞增殖能力优于使用单味药,且作用效果呈浓度?时间依赖性(P<?0.01).伤口愈合实验表明,牛黄?麝香联合使用对肝癌细胞迁移能力有明显的抑制作用(P<?0.01).肝癌裸鼠皮下移植瘤模型实验发现,牛黄?麝香组瘤体体积及质量均低于模型组(P<?0.01),牛黄?麝香组与DDP组PI3K/AKT/mTOR信号通路及S6K蛋白表达水平均明显下降(P<?0.01),牛黄?麝香组抑凋亡基因Bcl-2的表达下调(P<?0.05),促凋亡基因Bax及凋亡相关因子caspase-3和caspase-9的表达显著上调(P<?0.01).TUN-EL实验也进一步证实,二者联用能够促进肝癌细胞凋亡(P<?0.01).结论 牛黄?麝香联合使用能抑制人肝癌细胞SMMC-7721的增殖及迁移,并有效抑制肝癌裸鼠皮下移植瘤的生长,其机制可能与下调PI3K/AKT/mT-OR通路、调控凋亡相关蛋白caspase-3、caspase-9、Bcl-2和Bax的表达及促进细胞凋亡密切相关.“,”Objective To investigate the effects of Niuhuang (Bovis Calculus, BC) and Shexiang (Moschus) (BC-Moschus) on human hepatocellular carcinoma (HCC) cells SMMC-7721 and a nude mouse model of subcutaneous xenografts, and to explore its anti-HCC mechanism.Methods The BC-Moschus combination was applied to two liver cancer models in vivo and in vitro. SMMC-7721 was divided into the BC-Moschus group and the control group, and dif-ferent doses (rude drug dosage 0.625, 1.25, 2.5, and 5 mg/mL) of BC-Moschus extract were used for the intervention. The proliferation ability of HCC cells was detected using the Cell Counting Kit-8 (CCK-8) assay, and the migration ability was detected by a wound healing as-say. A subcutaneous xenograft model was prepared using nude mice with human HCC. Spe-cific pathogen-free-grade BALB/c nude mice (5-week-old) were randomly divided into the following groups (n = 6 per group): control (0.9％ physiological saline 0.2 mL/d), BC-Moschus [BC 45.5 mg/(kg·d) + Moschus 13 mg/(kg·d)], and cisplatin (DDP, intraperitoneal injection 5 mg/kg per week) groups. All groups were administered for 14 d. The volume and mass of the subcutaneous xenografts in nude mice were observed. The expression levels of phos-phatidylinositol-3 kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mT-OR) pathway, apoptosis-associated factor p70 S6 Kinase (S6K), Bax, Bcl-2, caspase-3, and cas-pase-9 in nude mice subcutaneous xenografts were measured by real-time quantitative PCR (RT-qPCR) and Western blot. Terminal Deoxynucleotidy Transferase-Mediated dUTP Nick-End Labeling (TUNEL) was used for quantitative analysis of apoptotic cells. Results The CCK-8 assay demonstrated that the BC-Moschus combination inhibited HCC cell proliferation in a superior manner to the use of BC and Moschus alone, and the inhibi-tion effect was dose- and time-dependent (P < 0.01). The wound healing assay showed that the BC-Moschus combination inhibited HCC cell migration (P < 0.01). In the subcutaneous xenograft model of nude mice with human HCC, we found that the tumor volume and weight of the BC-Moschus group were lower than those of the control group (P < 0.01). The levels of the PI3K/AKT/mTOR signaling pathway and S6K protein in the BC-Moschus and DDP groups were significantly decreased (P < 0.01). The expression level of the anti-apoptotic gene Bcl-2 was downregulated (P < 0.05), and the expression of the pro-apoptotic gene Bax and apoptosis-related factors caspase-3 and caspase-9 were significantly upregulated (P <0.01). The TUNEL assays further confirmed that the combination of the BC-Moschuas could promote HCC (P < 0.01).Conclusion The BC-Moschus combination inhibited the proliferation and migration ability of HCC cells SMMC-7721 and effectively inhibited the growth of subcutaneous xenografts in nude mice. The mechanism may be closely related to the downregulation of the PI3K/AKT/mTOR pathway, regulation of apoptosis-related protein caspase-3, caspase-9, Bcl-2, and Bax expression, and promotion of apoptosis.