Teng-Long-Bu-Zhong Granules Inhibits Tumor Growth And Metastasis Of CT26 Colorectal Cancer And Its Mechanism

Objective: To study the therapeutic effect and mechanism of Teng-Long-Bu-Zhong Granules(TLBZG)on growth and metastasis of murine CT26 colorectal cancer.Methods: The experimental study was divided into two parts.The first part,BALB/c mice were subcutaneously transplanted with CT26 cells.When the tumor tissue grew to a measurable range,the mice were divided into the control group,5-fluorouracil(5-Fu)group,compound Banmao capsule group,TLBZG group,5-Fu + compound Banmao capsule,5-Fu + low dose of TLBZG group,5-Fu + middle dose of TLBZG group and 5-Fu + high dose of TLBZG group.Mice were intragastrically administered with sterile water,compound Banmao capsule,different doses of TLBZG or/and intraperitoneally injected with 5-Fu.The length and width of the tumor were measured every 3 days.After 2 weeks of treatment,blood samples were collected and subjected to CD3+,CD4+,CD8+ T cells and NK cells detection by flow cytometry.Mice were sacrificed after 3 weeks of treatment,the tumors were removed and weighed.Apoptosis,cell senescence,protein expression,and cytokines were detected by Terminal deoxynucleotidyl transferase d UTP nick end labeling(TUNEL)assay,?-galactosidase staining,Western blot/immunohistochemistry and ELISA,respectively.In second part,lung metastasis of CT26 colorectal cancer was established in BALB/c mice by tail vein injection.Mice were randomly divided into control group,5-Fu group,compound Banmao capsule group,TLBZG group 5-Fu + compound Banmao capsule,5-Fu + low dose TLBZG group,5-Fu + middle dose of TLBZG group and 5-Fu + high dose of TLBZG group.Mice were intragastrically administered with sterile water,compound Banmao capsule,different doses of TLBZG or/and intraperitoneally injected with 5-Fu.Mice were sacrificed after 3 weeks of treatment,lungs were weighed and metastatic nodules in lung surface was counted.Metastatic nodules in lung tissues were detected by hematoxylin and eosin(HE)staining.Protein expression and phosphorylation were detected by immunohistochemistry and Western blot.Results: TLBZG inhibited CT26 tumor growth.TLBZG down-regulated PI3 K and AKT phosphorylation,activated Caspase-3,8 and 9,promoted PARP cleavage and induced apoptosis in CT26 colorectal cancer.TLBZG also increased p16 and p21 expression,down-regulated RB phosphorylation,inhibited E2F1 target gene CDK2 and Cyclin-E1 expression and promoted cell senescence.In addition,TLBZG reduce HIF1? expression,inhibit VEGF expression and angiogenesis.Furthermore,TLBZG increased CD3+,CD4+,CD8+ T lymphocytes and NK cells,up-regulated IL-4,IL-12 and IFN-? in mice bearing CT26 colorectal cancer.TLBZG enhanced the therapeutic effect of 5-Fu.TLBZG reduced metastasis nodules in lung surface and lung weight.TLBZG also inhibitd the expression of Wnt/?-catenin and its downstream target genes MMP-2 and MMP-9.TLBZG enhanced the therapeutic effect of chemotherapy.Conclusions: 1.TLBZG can inhibit the growth and metastasis of CT26 colorectal cancer cells and improve the effect of chemotherapy.2.TLBZG induces apoptosis in CT26 colorectal cancer which is related to down-regulation of phosphorylation of PI3 K and AKT,activation of Caspase-3,8 and 9 and PARP cleavage.3.TLBZG induces cell senescence in CT26 colorectal cancer which is related to up-regulation of p16 and p21,down-regulation of RB phosphorylation and E2F1 target genes CDK2 and Cyclin-E1.4.TLBZG inhibits angiogenesis in CT26 colorectal cancer which is related to down-regulation of HIF1?-VEGF signaling.5.TLBZG enhances immune function in mice bearing CT26 colorectal cancer and are associated with up-regulation of CD3+,CD4+ and CD8+ T cells and NK cells and IL-4,IL-12 and IFN-?.6.TLBZG inhibits metastasis of CT26 colorectal cancer which is related down-regulation of Wnt/?-catenin signaling and its target genes MMP-2 and 9.