Yangyin Fuzheng Jiedu Prescription exerts anti-tumor immunity in hepatocellular carcinoma by alleviating exhausted T cells,Phytomedicine

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Yangyin Fuzheng Jiedu Prescription exerts anti-tumor immunity in hepatocellular carcinoma by alleviating exhausted T cells
Phytomedicine ( IF 6.7 ) Pub Date : 2021-08-19 , DOI: 10.1016/j.phymed.2021.153722
Fengna Yan ₁ , Xinhui Wang ₁ , Yuqing Xie ₁ , Xiaoli Liu ₁ , Lihua Yu ₁ , Peng Wang ₁ , Tenghui Li ₂ , Shanshan Wang ₂ , Weihong Li ₃ , Zhiyun Yang ₁

Affiliation

Background

: Yangyin Fuzheng Jiedu Prescription (YFJP), a formulated Chinese herbal medicine, has been used for several decades in the treatment of hepatocellular carcinoma (HCC). Previous studies have demonstrated its anti-tumor efficacy, but the mechanism of action remains uncharacterized. This study aims to evaluate the therapeutic effect of YFJP on H22 tumor-bearing mice.

Purpose

: This study aimed to evaluate the therapeutic effect of YFJP on H22 tumor-bearing mice.

Methods

: A total of 50 male H22 tumor-bearing mice were randomly divided into 6 groups and continuous administered either different doses of YFJP or cyclophosphamide (CTX) or normal saline. for 2 weeks. The tumor appearance was observed by taking photos, and the tumor volume, weight, spleen and thymus index were calculated. Morphology of tumor infiltrating lymphocytes and the CD8⁺ T lymphocytes were detected through HE staining immunohistochemistry respectively. The frequency of CD3⁺, CD8⁺ T cell subsets and co-inhibitory receptors PD-1, TIGIT, Tim-3 on CD8⁺ T cell in spleen, peripheral blood and tumor tissue was performed by flow cytometry. Meanwhile, the killing and apoptotic functions of CD8⁺ T cells in tumor tissues were also detected by the same method. The levels of cytokines in peripheral blood were detected by Milliplex map mouse highs sensitivity T Cell kit. The expression of T cell transcription factor T-bet and Eomes in tumor tissues were observed by Western blot.

Results

: We found that YFJP could effectively inhibit the solid tumor growth and spleen indexes, but showed little effect on the body weight in the established mouse model of HCC. Furthermore, we investigated the effect of YFJP on the phenotypic and functional changes of T cells. The results showed that YFJP could maintain the high ratio of CD3⁺ and CD8⁺ T cells in the peripheral blood, spleen, and tumor tissues while decreasing the expression of programmed cell death-1 (PD-1), T cell immunoglobulin and ITIM domain (TIGIT), T cell immunoglobulin domain and mucin domain-3 (Tim-3) in CD8⁺ T cells, respectively. Surprisingly, PD-1/Tim-3 double-positive T cells in the peripheral blood and tumor tissues were significantly decreased. Additionally, YFJP restored the cytotoxicity of tumor-infiltrating T cells and delayed their apoptosis in H22 tumor-bearing mice. In addition, treatment with YFJP significantly decreased the expression of inflammatory and immunosuppressive cytokines (including IL-1β, IL-6, and IL-10) in the serum and tumor tissues whereas enhancing that of effector cytokines TNF-α, and IFN-γ. Moreover, T cell transcription factors T-bet increased and Eomes degraded in the tumor tissues upon YFJP treatment.

Conclusion

: In conclusion, these results demonstrated that YFJP could simultaneously exert anti-tumor immune response in H22 tumor-bearing mice by alleviating T cell exhaustion and immunosuppression.

中文翻译：

养阴扶正解毒方通过缓解衰竭T细胞在肝癌中发挥抗肿瘤免疫作用

背景

：养阴扶正解毒方（YFJP）是一种中草药配方，在肝细胞癌（HCC）的治疗中已经使用了几十年。先前的研究已经证明了其抗肿瘤功效，但作用机制仍未明确。本研究旨在评估YFJP对H22荷瘤小鼠的治疗效果。

目的

：本研究旨在评估 YFJP 对 H22 荷瘤小鼠的治疗效果。

方法

：总共50只雄性H22荷瘤小鼠随机分为6组，连续给药不同剂量的YFJP或环磷酰胺（CTX）或生理盐水。2周。拍照观察肿瘤外观，计算肿瘤体积、重量、脾脏和胸腺指数。HE染色免疫组化分别检测肿瘤浸润淋巴细胞和CD8⁺ T淋巴细胞的形态学。CD3⁺、CD8⁺ T 细胞亚群和共抑制受体 PD-1、TIGIT、Tim-3在脾脏、外周血和肿瘤组织中的CD8⁺ T 细胞上的频率通过流式细胞术进行。同时，CD8⁺的杀伤和凋亡功能肿瘤组织中的T细胞也用同样的方法检测。Milliplex图谱小鼠高敏T细胞试剂盒检测外周血细胞因子水平。Western blot观察肿瘤组织中T细胞转录因子T-bet和Eomes的表达。

结果

：我们发现YFJP可以有效抑制实体瘤生长和脾脏指标，但对建立的HCC小鼠模型的体重影响不大。此外，我们研究了 YFJP 对 T 细胞表型和功能变化的影响。结果表明，YFJP可以维持外周血、脾脏和肿瘤组织中CD3⁺和CD8⁺ T细胞的高比例，同时降低程序性细胞死亡-1（PD-1）、T细胞免疫球蛋白和ITIM结构域的表达。 (TIGIT)、CD8^{+ 中的}T 细胞免疫球蛋白结构域和粘蛋白结构域-3 (Tim-3)T 细胞，分别。令人惊讶的是，外周血和肿瘤组织中的PD-1/Tim-3双阳性T细胞明显减少。此外，YFJP 恢复了肿瘤浸润性 T 细胞的细胞毒性，并延迟了 H22 荷瘤小鼠的细胞凋亡。此外，YFJP 治疗显着降低血清和肿瘤组织中炎症和免疫抑制细胞因子（包括 IL-1β、IL-6 和 IL-10）的表达，同时增强效应细胞因子 TNF-α 和 IFN-γ 的表达. 此外，在 YFJP 治疗后，肿瘤组织中的 T 细胞转录因子 T-bet 增加和 Eomes 降解。