| Breast cancer is a common threat to women’s health and life, as well as the wildly existence of depression and anxiety disorders among breast cancer patients. Studies have shown that long-term depression and anxiety in patients with breast cancer will have a huge negative regulatory role in immune function, promoting the diseases. Myeloid-derived suppressor cells (MDSCs) is a specific group of immune suppressor cells, had become the hot spot in immune and tumor research in recent years. Natural killer T cells (NKT) is a group of immune cells characterized by both NK and T cells, as an " double-edged sword " in immunomodulatory. Breast cancer patients under long-term depression and anxiety and the tumor environment, which can produce large amounts of inflammatory cytokines, raising, amplification, and active of MDSC, starting immunosuppressive effect, promoting tumor progression and escape. The function of NKT cells can be remodeling by drug, thereby antagonizing the immunosuppressive effects of MDSC, improving immune function, and ultimately play the anti-tumor effect.Objective53breast cancer patients were treated by treatment by Liver-soothing and Spleen-fortifying Formula(LSSFF) for two weeks, to observe the changes of depressive and clinical symptoms by using Self-Rating Depression Scale (SDS) and TCM symptoms of depression assessment scale.Through basic research to reveal all changes leaded by Liver-soothing and Spleen-fortifying Formula(LSSFF) in the immune inhibitory mechanisms, and immune function of NKT cells remodeling mechanisms. To reveal the role of NKT cells and MDSC in the immune editing process of breast cancer. Provide new scientific basis for prevention and treatment of breast cancer. Add new theoretical foundation of" emotion theory" to TCM.MethodsClinical Study:Using Self-Rating Depression Scale (SDS) and TCM symptoms of depression assessment scale as assessment Methods, to observe the changes of depressive and clinical symptoms after the treatment of LSSFF for two weeks in53breast cancer patients treated.Basic research:built and evaluate depression of tumor-bearing model; take chemotherapy drug gemcitabine and LSSFF as the interventions. Collected samples of hippocampus, tumors, spleen and peripheral blood to detect MDSC immunosuppressive effects mediated by various aspects of indicator. Detect5-HT in hippocampus by HPLE-ECD; detect MDSC and its subtypes, NKT cells, T cell subsets in spleen by flow cytometry; detect the apoptosis of CD4+T and CD8+T cell; detect IL-6, IL-4, IL-13, IL-12, IFN-y, TGF-β and VEGF in peripheral blood by ELISA; assay NO and iNOS in spleen; and detect of the relevant nuclear transcription factors of MDSC in spleen by RT-PCR.ResultsResults of clinical studyStatistics showed that patients with breast cancer accompanied by depressive disorders accounted for67.92%, mild depression accounted for52.83%, moderate depression accounted for11.32%, accounting for3.77%of severe depression. After treatment of LSSFF for two weeks, patients accompanied by depressive disorders accounted for56.6%, mild depression accounted for43.4%, moderate depression accounted for9.43%, approximately3.77%of severe depression. Before treatment there were17cases less than53points of SDS standard score,9of them decreased after two weeks treatment of LSSFF. Among the patients more than53points before treatment, there were17cases decreased1-5points,5cases decreased6-10points; but there was no case decrease more than10points. After the treatment of LSSFF, clinical symptoms generally eased, scores dropped also obviously, mainly in segments of1-3and4-6.Detect neurotransmitters of5-HT in hippocampus by HPLE-ECDExpression of5-HT,5-HIAA and DA were significantly higher in TCM group, Chemotherapy group, Integrative Medicine group than the expression of the model control group(p<0.05). The expression of5-HT in TCM group was higher than chemotherapy group and the Integrative Medicine group (p<0.05); TCM group is higher in the expression of5-HIAA than Integrative Medicine group (p<0.05); The expression of DA in TCM group was significantly higher than in the chemotherapy group, Integrative Medicine group (p<0.01). The results showed that depression and tumor can reduce the expression of5-HT and other neurotransmitters, after drug intervention, the expression could improve, even higher than the normal group. Detect the expression of MDSC and its subtypes, NKT cells, T cell subsets in spleen by flow cytometryMDSC expressing in tumor groups were higher than the non-tumor groups (control and pure depression group), the model control group were higher as1times than the normal group; MDSC in chemotherapy group and Integrative Medicine group were less than the model control group(p <0.05).NKT cells expressing in model control group were lower than Integrative Medicine group, pure depression group and the normal group, with statistical significance (p <0.05). NKT cells express elevated after the intervention of Chinese and Western medicines.The results of CD4+/CD8+in each group:the ratio of the normal group was higher than other groups; model control group without the intervention had the lowest ratios; the ratio of the Integrative Medicine group was greater than the normal group (p 〈0.05).The results showed that depression and tumor can increase the expression of MDSC, improve immune inhibition function. After intervention of drug can up-regulate NKT and CD4+/CD8+, improve the immune function.Detect the apoptosis of CD4+T and CD8+T cell in spleenThe early apoptosis of CD8+T cells in the normal group had the lowest rate of1.28%, the model control group had the highest rate of8.78%. Among three treatment groups, result is Integrative Medicine Group<chemotherapy group<TCM group.The early apoptosis of CD4+T cells in the normal group had the lowest rate of2.91%, the model control group had the highest rate of7.33%. In the three treatment groups, the Integrative Medicine Group had the lowest early apoptosis rate.Detect cytokines associated MDSC in peripheral blood by ELISAIL-6, IL-4, IL-13, IFN-y, TGF-β and VEGF play negative role in immunosuppressive function caused by MDSC. Model control group had the highest expression of them; the expression of three treatment groups were lower than model control group.IL-12play a positive role in the regulation of immune function mediated by MDSC. The expression of IL-12in the model control group was lower than TCM group and Integrative Medicine Group(p<0.05). Indicating that after drug intervention, IL-12expression improved and effectively antagonize the expression of IL-13, limited the ctivation and amplification of MDSC, improving immune function. Assay the expression of NO and iNOS in spleenNO expression in Chemotherapy group, TCM group and model control group were significantly lower than the normal group (p <0.05), depression and tumor-bearing groups had the higher level of MDSC. After administration of the intervention, Chemotherapy group, TCM group and Integrative Medicine Group had lower NO expression than model control group, Integrative Medicine Group lower than model control group had Statistically significant difference(p <0.05). Indicated after LSSFF and chemotherapy, down-regulate the expression of MDSC, decreased its immunosuppression.iNOS expression in Chemotherapy group, TCM group and model control group were significantly higher than the normal group (p <0.05). In this study the expression of iNOS and MDSC expressed in the spleen had the same trend, drug intervention could decrease the immune suppression function, but no statistically significant difference (p>0.05) among the intervention groups.Detect of the relevant nuclear transcription factors of MDSC in spleen by RT-PCRThe expression of JAK1, JAK2, STAT1, STAT3, STAT6and Argl, iNOS in model control group were highest. This trend was amplified with the inflammatory cytokines started MDSC. After drug intervention, especially after the combined intervention can significantly decrease the JAK-STAT pathway gene expression. But pure herbs on each set of genes down-regulated inferior degree was lower than the Integrative Medicine Group and pure chemotherapy group.ConclusionClinical studies have shown that traditional Chinese medicine(LSSFF) can relieve depression in breast cancer patients, but the improvement was mainly reflected in mild depression and somatic symptoms; relieve clinic symptoms have a more significant effect.Basic research shows that depression and tumor-bearing state can decreasethe expression of5-HT and other neurotransmitters. And can secrete huge amounts of inflammatory cytokines, highly up-regulate of the genes in JAK-STAT signaling pathway, thereby activating and amplificating MDSC; upregulation the genes of Argl, iNOS, and its product NO significant increased, eventually forming a large number of ONOO-, destroy the body normal immune function, down-regulate the expression of NKT and T cell subsets, increased the early apoptosis of CD4+T and CD8+T cells, ultimately promote tumor growth and metastasis. TCM combination with chemotherapy improve immune function by remodeling function of NKT cells, through multiple targets and pathways. |
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