Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, 210023, China; Jiangsu Joint International Research Laboratory of Chinese Medicine and Regenerative Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China.
Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, 210023, China; Department of Biochemistry and Molecular Biology, School of Medicine & Holistic Integrative Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China.
J Ethnopharmacol. 2023 Nov 15;316:116735. doi: 10.1016/j.jep.2023.116735. Epub 2023 Jun 5.
Based on the notion of traditional Chinese medicine, the theory of invigorating the circulation of blood is a prominent treatment for cancer in clinic. Therefore, Salvia miltiorrhiza Bunge, as a representative of Chinese medicine of invigorating the circulation of blood, has been proved to be an effective medicinal herb for treating cancer.
To clarify the anti-cancer effect of Salvia miltiorrhiza Bunge aqueous extract (SMAE) on colorectal cancer (CRC) and investigate whether the therapeutic effect of SMAE was mediated by attenuating the infiltration of tumor-associated macrophages (TAMs) into the tumor microenvironment (TME).
High-performance liquid chromatography (HPLC) was used for determined the main compounds of SMAE. MC38 cells were subcutaneously injected into the mice to establish the mouse model of CRC. Tumor growth curve was detected by tumor volume measurement. The model group received distilled water irrigation once a day. SMAE-treated group received 5 g/kg or 10 g/kg SMAE once a day. Anti-PD-L1 treated group received 5 mg/kg anti-PD-L1 once every three days. Protein expression of Cox2 and PD-L1 was determined by Western blot assay. The secretion levels of PGE2, IL-1β, IL-6, MCP-1, and GM-CSF were evaluated through ELISA. The mRNA expression of CSF1, CCL2, CXCL1, CXCL2, and CXCL3 was measured by using RT-qPCR. Staining of Ki67, TUNEL and Caspase3 was used to investigate cell proliferation and apoptosis. Immunohistochemical staining was used to determine CD8 T cell distribution. H&E staining was used to confirm histopathological changes. The expressions of F4/80 and CD68 were measured by flow cytometry to identify macrophages in tumors and lymph nodes. The number of CD8 T cells and the expression of PD-1, IFN-γ, and Granzyme B (GZMB) were determined by flow cytometry.
SMAE significantly retarded the growth of MC38 mouse colorectal cancer. SMAE strikingly inhibited the expression of Cox2 and impaired the secretion of PGE2 in tumors, contributing to the attenuated intra-tumoral infiltration of TAMs via Cox2/PGE2 cascade. Meanwhile, SMAE augmented anti-tumor immunity by the elevated proportion of IFN-γ CD8 T cells and GZMB CD8 T cells, which decreased the tumor load. Furthermore, the combination of SMAE and anti-PD-L1 showed a higher therapeutic efficacy than either monotherapy in controlling tumor growth in MC38 xenograft model.
SMAE attenuated the infiltration of TAMs into tumors and synergized with anti-PD-L1 to treat CRC via modulating Cox2/PGE2 cascade.
基于中药的理念，活血化瘀理论是临床上治疗癌症的主要方法。因此，丹参作为活血化瘀中药的代表，已被证明是治疗癌症的有效药物。
阐明丹参水提物（SMAE）对结直肠癌（CRC）的抗癌作用，并探讨 SMAE 的治疗效果是否通过减轻肿瘤相关巨噬细胞（TAMs）浸润肿瘤微环境（TME）来介导。
采用高效液相色谱法（HPLC）测定 SMAE 的主要成分。将 MC38 细胞皮下注射到小鼠中建立 CRC 小鼠模型。通过肿瘤体积测量检测肿瘤生长曲线。模型组每天接受蒸馏水灌胃一次。SMAE 处理组每天接受 5g/kg 或 10g/kg SMAE 一次。抗 PD-L1 处理组每三天接受 5mg/kg 抗 PD-L1 一次。Western blot 检测 Cox2 和 PD-L1 蛋白表达。通过 ELISA 评估 PGE2、IL-1β、IL-6、MCP-1 和 GM-CSF 的分泌水平。采用 RT-qPCR 测定 CSF1、CCL2、CXCL1、CXCL2 和 CXCL3 的 mRNA 表达。用 Ki67、TUNEL 和 Caspase3 染色检测细胞增殖和凋亡。免疫组织化学染色检测 CD8 T 细胞分布。H&E 染色确认组织病理学变化。用流式细胞术测量 F4/80 和 CD68 的表达以鉴定肿瘤和淋巴结中的巨噬细胞。用流式细胞术测定 CD8 T 细胞的数量以及 PD-1、IFN-γ 和 Granzyme B（GZMB）的表达。
SMAE 显著延缓 MC38 小鼠结直肠癌的生长。SMAE 显著抑制 Cox2 的表达，并通过 Cox2/PGE2 级联破坏肿瘤内 TAMs 的浸润。同时，SMAE 通过增加 IFN-γ CD8 T 细胞和 GZMB CD8 T 细胞的比例增强抗肿瘤免疫，从而降低肿瘤负荷。此外，与单药治疗相比，SMAE 与抗 PD-L1 联合治疗在控制 MC38 异种移植模型中的肿瘤生长方面具有更高的疗效。
SMAE 通过调节 Cox2/PGE2 级联减轻 TAMs 浸润肿瘤，并与抗 PD-L1 协同治疗 CRC。
