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目的 利用动物模型阐明CCL23对肝细胞癌恶性进展的影响及作用机制，验证靶向CCL23/CCR1/AKT信号轴的治疗潜力。方法 通过qRT-PCR筛选出本底低表达CCL23的肝细胞癌细胞系，构建慢病毒介导的CCL23过表达细胞株。采用CCK-8增殖检测、平板克隆实验、Transwell实验及EdU实验验证CCL23过表达对肝细胞癌细胞恶性表型的影响。通过BALB/c nu/nu小鼠皮下移植瘤模型(慢病毒/AAV双重验证)、尾静脉肺转移模型及免疫健全鼠模型，评估过表达CCL23对肿瘤生长与转移的影响。利用蛋白免疫印记技术解析AKT通路活化、转移相关蛋白及CD8蛋白的表达情况。使用CCR1的拮抗剂BX471进行阻断，确认CCL23/CCR1/AKT通路的生物学功能依赖性。结果 1)筛选出具有CCL23本底低表达的肝细胞癌细胞系MHCC97H,成功构建慢病毒介导的CCL23过表达细胞株。2)体外实验结果证明，与对照组相比，CCL23过表达组细胞的增殖、迁移及侵袭能力均降低。3)慢病毒及AAV介导的CCL23过表达均显著抑制BALB/c nu/nu小鼠皮下肿瘤生长，且肿瘤组织中Ki-67、p-Akt及PCNA蛋白表达水平均显著降低。4)尾静脉注射肺转移模型中，CCL23过表达组肺转移灶数量及面积显著减少，且E-cadherin表达上调、N-cadherin及p-Akt表达水平下调。5)在免疫健全小鼠模型中，CCL23过表达同样可抑制皮下瘤生长，且CD8蛋白表达增加。结论 CCL23通过CCR1/AKT信号轴抑制肝细胞癌生长和肺转移并增强CD8~+T细胞浸润。CCL23/CCR1/AKT信号轴是肝细胞癌的潜在治疗靶点。
Abstract：Objective To elucidate the effect and mechanism of CCL23 on the malignant progression of hepatocellular carcinoma using animal models, and to verify the therapeutic potential of targeting the CCL23/CCR1/AKT signaling axis. Methods Hepatocellular carcinoma cell lines with low basal expression of CCL23 were screened by qRT-PCR, and lentivirus-mediated cell lines with CCL23 overexpression were constructed. CCK-8 proliferation assay, colony formation assay, Transwell assay and EdU assay were used to verify the effect of CCL23 overexpression on the malignant phenotypes of hepatocellular carcinoma cells. The effects of CCL23 overexpression on tumor growth and metastasis were evaluated using a subcutaneous xenograft tumor model in BALB/c nu/nu mice(validated by both blot was used to analyze the activation of the AKT pathway, the expression of metastasis-related proteins and CD8 protein. The biological function dependence of the CCL23/CCR1/AKT pathway was confirmed by blocking with BX471, an antagonist of CCR1. Results 1) Hepatocellular carcinoma cell line MHCC97H with low endogenous CCL23 expression was selected, and a CCL23-overexpressing cell line was successfully established via lentiviral transduction. 2) In vitro experiments demonstrated that, compared to the control group, cells in the CCL23-overexpression group exhibited reduced proliferation, migration, and invasion capabilities. 3) Both lentivirus-mediated and AAV-mediated CCL23 overexpression significantly suppressed subcutaneous tumor growth in BALB/c nu/nu mice. Furthermore, protein expression levels of Ki-67, p-Akt, and PCNA were significantly decreased in the tumor tissues. 4) In the tail vein injection lung metastasis model, the CCL23-overexpression group showed a significant reduction in both the number and area of lung metastatic foci. Additionally, E-cadherin expression was upregulated, while N-cadherin and p-Akt expression levels were downregulated. 5) In immunocompetent mouse models, CCL23 overexpression also inhibited subcutaneous tumor growth and was associated with increased CD8 protein expression. Conclusion CCL23 inhibits the growth and lung metastasis of hepatocellular carcinoma and enhances the infiltration of CD8~+T cells through the CCR1/AKT signaling axis. The CCL23/CCR1/AKT signaling axis is a potential therapeutic target for hepatocellular carcinoma.
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基本信息:
DOI：10.13880/j.cnki.65-1174/n.2025.22.029
中图分类号:R735.7
引用信息:
[1]王凤桐,侯卜文,舒敏,等.CCL23通过CCR1/AKT信号轴抑制肝细胞癌生长及转移[J].石河子大学学报(自然科学版),2025,43(06):702-711.DOI:10.13880/j.cnki.65-1174/n.2025.22.029.
基金信息:
新疆维吾尔自治区天山英才科技创新团队项目(2023TSYCTD0020)
2025-12-03
2025-12-03
2025-12-03