Experimental Study On Liver Metastases Of Colorectal Carcinoma Treated By Blood-activating And Stasis-eliminating Herbs And Its Interaction With Erythrocyte Immunity

Object:Supported by the National natural science foundation, we choose classical traditional Chinese medicine (TCM) prescriptions of promoting blood circulation and removing blood stasis, namely Guizhifuling Pills and Danshen Tang, observe their effects on the metastatic behavior of malignant tumor, explore the possible mechanisms of such effects and their interaction with erythrocyte immunity.Method:1. Preparation of TCM prescription Guizhifuling Pills (Guizhi 9g, Chishao 9g, Danpi 9g, Shaoyao 9g, Fuling 9g) and Danshen Tang (Danshen 30g) were chosen. It will switched into the equivalent dosage while feeding the mice.2. Replication of the mouse model and grouping of the miceLiver metastases were replicated in BALB/c mice by intrasplentic injection ofcolonic adenocarcinoma cells(CT-26) with 1 X 106 cells in 0.1 ml solution. Erythrocyte immunodeficiency mice model were replicated by feeding the low-selenium forage for 10 weeks.96 BALB/c mice were randomly divided into 2 groups, namely group I and II which were divided into 4 small groups respectively, each of the small group contain 12 mice. Group I were the mice with normal erythrocyte immunity: (D Normal control group, (2) Pathological control A group (receive intrasplenic injection of CT-26 cells), (3) Guizhifuling Pill A group and ? Danshen Tang A group (receive both of the intrasplenic injection of CT-26 cells and TCM prescription). Group II were mice with erythrocyte immunodeficiency: ? RBC-hypoimmunity group, (6) Pathological control B group( erythrocyte immunodeficiency + intrasplenic injection of CT-26 cells), ? Guizhifuling pill B group and ? Danshen Tang B (erythrocyte immunodeficiency + intrasplenic injection of CT-26 cells + treatment of TCM prescription).3. The indicators of the observation3.1 The liver weight and number of the metastic tumor were surveyed.3.2 The detection of RBC immune adherence function (RIAF)The indicators of RIAF included RBC C3b receptor rosette(R-C3bR), RBC immune complex receptor rosette(R-ICR) and RBC immune adherence tumor cell receptor rosette(RITCR). A RBC adhered to more than 2 microzyme were defined as positive and the RBC rosettes were counted.3.3 The expression of human telomerase reverse transcriptase(hTERT), P53, C-erbB-2 and Bcl-2 were detected by SABC immunohistochemical(IH) method. Brown was defined as positive expression in the IH method.Result:1. The hepatic metastasis models of colonic adenocarcinoma with different RIF was successfully replicated. We used CT-26 murine colon adenocarcinoma cells syngeneic to BALB/c mice, and it is characterized by high and stable repetition and metastasis incidence, and similar metastasis pathway to patients. The method for mice model replication is feasible with stability, reliability and repeatability and can meet the demand of related researches.2. Tumor metastasis in the group mice with erythrocyte immunodeficiency were severer with significant difference (P<0.05) compared with the normal group mice inoculated with tumor cells. After all group animals were administrated with TCM prescriptions, the findings indicate that Guizhifuling Pills and Danshen tang can inhibit tumor metastasis in normal group animals inoculated with tumor cells. However, effects of the two prescriptions don't display remarkable difference compare with the pathological control B group (P >0.05), and there was remarkable difference between erythrocyte immunodeficiency mice and the normal mice both inoculated with tumor cells and treated by the two prescriptions (P <0.05).3. The expression of hTERT, P53, C-erbB-2 and Bcl-2 in the models receiving treatment with Guizhifuling Pills differed significantly from those of pathological control A group (P <0.05), whereas between Danshen tang and the model group, only the expression of hTERT showed significant difference.Conclusion:1. Being erythrocyte immunodeficiency, the liver metastasis of colorectal carcinoma was observed obviously. The blood-activating and stasis-eliminating herbs could inhibit carcinoma metastasis exactly. In group I , the tumor inhibitive effect ofGuizhifuling Pills is better than Dansken tang(P <0.05). However, in group II, there is no significant difference for the tumor inhibitive effect between Guizhifuling Pills and Danshen tang.2. Guizhifuling Pills can reduce the expressions of hTERT, p53, c-erbB-2 and Bcl-2 in the mice models, while Danshen tang can only reduce the expression of hTERT without similar effects on the other 3 oncogenes. Such inhibitory effects of the prescriptions on hTERT and the oncogenes might explain their actions to inhibit malignant tumor metastasis.3. Mechanism of inhibiting tumor metastasis: ? related to the function of erythrocyte immunity, (2) controlling the expression of telomerase, (3) controlling the expression of P53,C-erbB-2,Bcl-2.4. The inhibiting effects of the TCM of blood circulation promotion by removal of blood stasis on tumor metastasis depend on erythrocyte immune function in great extent.