| Basis for selection: Breast cancer ranks first in the incidence and mortality of female cancer,which is a serious threat to women’s health.Chemotherapy is the main means of breast cancer treatment at all stages,but because of the rapid growth of tumor blood vessels and structural abnormalities,it is difficult to deliver chemotherapy drugs,which is an urgent problem to be solved.Vascular normalization is a newly proposed theory in recent years,aiming to improve the efficiency of chemotherapy drug delivery by promoting the normalization of tumor vascular structure and function.Modern pharmacological studies have shown that Chinese herbs for promoting blood circulation and removing blood stasis can improve the hypoxic microenvironment of tumor,the hypercoagulable state of blood and eliminate microcirculation disorders.Salvianolic acid A,as one of the main active monomer components of Salvia miltiorrhiza,is widely used in the treatment of cardiovascular and cerebrovascular diseases.However,A large number of research data prove that salvianolic acid A plays a significant role in anti-tumor and adjuvant tumor chemotherapy,but relevant studies only stay in the pharmacodynamic evaluation stage,and the pharmacological mechanism is still unclear.Based on the above background,this study explored the mechanism of salvianolic acid A in "activating blood circulation" assisted by tumor chemotherapy from the perspective of "tumor vascular normalization".Objective: In this study,the orthotopic transplantation model of mouse breast cancer was used as the in vivo pharmacological study model,the angiogenesis model of endothelial cell proliferation induced by lactic acid was used as the in vitro activity study model,and the role of salvianolic acid A in promoting tumor vascular normalization and improving the efficacy of paclitaxel chemotherapy was investigated by integrating cell metabolomics technology,aiming to clarify the possible pharmacological mechanism of salvianolic acid A in promoting vascular normalization.Research methods:(1)A mouse breast cancer orthotopic transplantation model was established by injecting mouse breast cancer cells 4T1 into the fourth pair of nipple fat pads of female Balb/c mice.After the model was established,the animals were divided into control group,paclitaxel monotherapy group,salvianolic acid A monotherapy group,and combined administration group.The effects of salvianolic acid A on improving paclitaxel chemotherapy and inhibiting tumor metastasis were evaluated based on tumor mass,tumor volume,mouse body weight,expression levels of TGF-β and CSF-1,which are related to tumor metastasis.The expression of endothelial cell marker CD31,pericellular marker NG2 and hypoxic microenvironment marker CA-IX were used to characterize the improvement of salvianolic acid A on tumor vascular structure and function.(2)Different concentrations of lactic acid were used to stimulate human umbilical vein endothelial cells HUVEC,and cell proliferation was used as the model construction index to determine the optimal lactate modeling dose.After the successful construction of the model,two concentrations of salvianolic acid A were given for intervention,and the proliferation of endothelial cells was analyzed by MTT colorimetry,Ki-67 immunofluorescence,Ed U staining,immunowestern blot and cell counting.(3)UHPLC-Q-TOF-MS method was used to analyze the cell samples treated with lactic acid and salvianolic acid A,and the differential metabolic components were identified based on multivariate statistical analysis and mass number,retention time and fragment ions.Finally,Metabo Analyst was used to conduct pathway analysis of metabolites that could potentially promote tumor vascular normalization.Research results:(1)By examining the physiological and pathological status of mice,it was found that the average tumor inhibition rate of paclitaxel group was 42.15%,and the tumor inhibition rate of combined administration group was increased to 56.80%;The mean tumor weight,tumor growth rate and expression levels of TGF-β and CSF-1 related factors related to tumor metastasis in the combined administration group were lower than those in the paclitaxel monotherapeutic group.Further characterization of tumor vascular structure and function showed that,compared with paclitaxel monotherapy group,the combined administration group could improve the disordered arrangement of vascular cells in tumor tissues,increase the coverage of pericellular cells,and alleviate the hypoxia of tumor tissues.(2)Construct a lactic-stimulated tumor angiogenesis model in vitro.After concentration and time screening,it was found that lactic acid treated with 10 m M for 48 h had the most significant effect on the proliferation of endothelial cells,while salvianolic acid A treated with 0.01 m M and 0.02 m M could significantly interfere with the activation of lactic acid on the proliferation of endothelial cells,and reduce the expression of the proliferation-related protein Ki-67.(3)The results of cell metabolomics analysis showed that there were metabolic differences between the cells of the control group and the cells of the lactate-induced model group,and 13 different metabolites were identified and compared.Salvianolic acid A can regulate the expression of c AMP,AMP,niacinamide,glucose-6-phosphate and other differential metabolites.Further metabolic pathway analysis indicated that purine metabolism may be A potential key metabolic pathway for salvianolic acid A to promote endothelial cell homeostasis and regulate tumor vascular normalization.Conclusion: Salvianolic acid A may stabilize vascular structure and endothelial cell proliferation by regulating purine metabolism,thereby promoting tumor vascular normalization and improving the efficiency of paclitaxel chemotherapy. |
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