ABSTRACT
ObjectiveTo explore the effects of modified Danggui Beimu Kushen pills on tumor growth and T-cell subsets in H22 hepatocellular carcinoma-bearing mice and to provide an experimental basis for the treatment of hepatocellular carcinoma with modified Danggui Beimu Kushen pills combined with immune checkpoint antibodies. MethodA H22 hepatocellular carcinoma-bearing mouse model was established. The modeled mice were randomized into model， cisplatin， low- （4 g·kg-1·d-1）， medium- （8 g·kg-1·d-1）， and high-dose （16 g·kg-1·d-1） modified Danggui Beimu Kushen pills groups. After continuous administration for 14 days， the mice were sacrificed on day 15. The tumor volume was measured on days 0， 4， 8， 12， 15 of drug administration. Tumors were weighed and thymus index and spleen index were calculated. Spleen lymphocytes were co-cultured with H22 hepatoma cells， and the tumor cell-killing rate was detected by the cell counting kit-8 （CCK-8）. Real-time polymerase chain reaction was carried to determine the mRNA levels of programmed cell death protein-1 （PD-1） and lymphocyte activation gene-3 （LAG-3） in spleen and tumor tissues. The number of CD4+ and CD8+ T cells and the expression of PD-1 and LAG-3 were detected by immunohistochemistry （IHC）. ResultOn day 8 of drug administration， tumor volumes in all treatment groups decreased compared with that in the model group. On day 15， both tumor volume and tumor weight were significantly lower in the treatment groups than in the model group （P<0.01）， with the cisplatin group showing the most pronounced reduction. Compared with the model and cisplatin groups， medium- and high-dose modified Danggui Beimu Kushen pills increased the thymus index （P<0.01）. Compared with the model group， all treatment groups showed increased spleen index （P<0.05， P<0.01）， with the cisplatin group showing the most significant increase. Compared with the model and cisplatin groups， all the groups of modified Danggui Beimu Kushen pills demonstrated increased number of CD4+ and CD8+ T cells and tumor cell-killing rate in the spleen and tumor tissues （P<0.01） and down-regulated mRNA and protein levels of LAG-3 （P<0.05， P<0.01）. The high-dose group of modified Danggui Beimu Kushen pills had lower mRNA level of PD-1 in the tumor tissue than the model and cisplatin groups （P<0.01）. ConclusionModified Danggui Beimu Kushen pills may promote the proliferation and tumor microenvironment infiltration of CD4+ and CD8+ T cells in H22 tumor-bearing mice by down-regulating LAG-3 expression， thereby improving T-cell immune activity and inhibiting tumor growth. This study provides an experimental basis for the combination of modified Danggui Beimu Kushen pills and immune checkpoint antibodies in the treatment of hepatocellular carcinoma.
ABSTRACT
Objective To analyze the material basis and molecular mechanisms of Danggui Beimu Kushen (DBK) Pills in treating prostatic diseases based on the method of integrated pharmacology. Method The platform of Integrative Pharmacology of Traditional Chinese Medicine (TCM-IP, www.tcmip.cn) was utilized to predict the main active ingredients and functional targets of DBK Pills in treating prostatic disease, key targets were screened for enrichment analysis of pathways, and the network of “herb-core component-key target-main pathway” was constructed, and the possible mechanisms of DBK Pills in treating prostatic diseases were explored. Results A total of 532 candidate key targets for the treatment of prostatic diseases by DBK Pills were predicted, and 1 840 terms of gene function and 194 signal pathways were analyzed by gene ontology (GO) and KEGG, respectively. The network analysis of “herb-core component-key target-main pathway” showed that 65 core components were predicted, including 29 ingredients from Angelica sinensis, 11 from Fritillaria thunbergii and 26 from Sophora flavescens. Those predicted components acted on the key targets of prostatic diseases, such as transcription factor binding, negative regulation of apoptosis, et al, through the estrogen, apoptosis, chemokines and other signal pathways, and thus played a role in the regulation of cell cycle, apoptosis and proliferation imbalance, which might be the molecular mechanisms of DBK Pills for the treatment of prostatic disease. Conclusion DBK Pills regulate the development of BPH, prostate cancer and other diseases through multiple pathways with multi-component interacting with multiple targets.
ABSTRACT
Objective To study the effects of modified Danggui Beimu Kushen Pills on the expressions of TLR2, TLR4, TLR6, TRAF6 and MyD88 in tumor tissues on MFC gastric cancer bearing mice; To discuss relevant mechanism of action. Methods MFC gastric cancer bearing mice were employed to perform anti-tumor experiment in vivo in this study. A total of eligible 48 mice were randomly divided into model group, DDP positive control group, modified Danggui Beimu Kushen Pills high-dose and low-dose groups, modified Danggui Beimu Kushen Pills high-dose and low-dose combined with DDP groups. The treatment was conducted once a day, and lasted for 14 continuous days. After the last administration of gavage orally treatment, all mice were anaesthetized and killed by cervical dislocation method to obtain tumor tissue completely for further HE staining measure and detection of TLR2, TLR4, TLR6, TRAF6 and MyD88 in tumor tissue with the method of RT-qPCR and immunohistochemistry. Meanwhile, the tumor growth was observed and the general conditions of mice were recorded. Results The model group was rich in tumor cells; the sizes of cells were different; the volume was large; the nucleus was deeply stained and the heterotypic shape was obvious, and the small focal necrosis was seen. The number of tumor cells in each administration group was reduced; the arrangement was loose; the cell volume was significantly reduced, and the nuclear pyknosis was reduced. Cell necrosis significantly increased; the number of interstitial blood vessels decreased; collagen fibers increased, especially in modified Danggui Beimu Kushen Pills high-dose and low-dose combined with DDP groups. Compared with the model group, the expressions of TLR2, TLR4, TLR6, TRAF6 and MyD88 mRNA and protein decreased in each administration group. TLR2, TLR4, TLR6, TRAF6 and MyD88 were lighter and weakly positive expressed in modified Danggui Beimu Kushen Pillshigh-dose and low-dose combined with DDP groups, the protein changes were more obvious Compared with DDP positive control group, modified Danggui Beimu Kushen Pills high-dose and low-dose groups. Conclusion Modified Danggui Beimu Kushen Pills can down-regulate TLR2, TLR4, TLR6, TRAF6 and MyD88 expression of tumor tissue in MFC gastric cancer bearing mice at both mRNA and protein levels to play anti-tumor pharmacology action.