Study On The Mechanism Of Shuanghuanglian Component Traditional Chinese Medicine Intervening Colon Cancer Proliferation And Metastasis

Colon cancer(CC)is one of the digestive tract cancers that occurs in the colon with high incidence and mortality rate.In China,the incidence of CC is increasing year by year.Clinical observation shows that the early symptoms of CC are not obvious,and a significant proportion of patients are found to be at an advanced stage.The prognosis and effect are still poor under current therapies.Therefore,there is an urgent need to elucidate the molecular mechanism of CC and to find preventive and intervention measures.Numerous studies have shown that lysyl oxidase-like 2(LOXL2)is involved in tumor development and is closely related to clinical prognosis.LOXL2 was initially widely considered as a secretory protein distributed in extracellular matrix.In recent studies,it has also been found to be involved in intracellular responses and could regulate epithelial-mesenchymal transition of tumor cells,influence tumor angiogenesis,and induce tumor hypoxia.LOXL2 plays an extremely critical role in tumor progression.However,the effect of LOXL2 in CC remains unclear.In order to explore the specific mechanism of LOXL2 in CC,we plan to conduct the following study.Firstly,we found that LOXL2 was up-regulated in many types of cancers through the TCGA database.Immunohistochemistry(IHC)of CC tissue confirmed that LOXL2 was positively expressed in CC and the expression was associated with tumor size.Survival analysis revealed that CC patients with a higher LOXL2 expression always had a poor clinical prognosis,and its expression level could be used to grade patients in stage ??stage ?.Then,we discussed the relationship between LOXL2 and cancer-associated fibroblasts(CAFs)in tumor microenvironment.IHC results and R2 database analysis showed that LOXL2 expression was positively correlated with the distribution of CAFs in CC tissues.Western blotting and immunofluorescence results showed that NIH/3T3 cells were activated as CAFs after recombinant LOXL2(rLOXL2)stimulation.Transwell assay showed that CAFs activated by CC cells or rLOXL2 could further promote the migration and invasion of CT26 cells in vitroIn CC prevention and treatment,the methods of network pharmacology were adopted to screen a multi-component Chinese medicine(MCCM)with anti-tumor activity from Shuang-Huang-Lian.The results showed that baicalin,chlorogenic acid and forsythoside A were matched with anti-tumor activity.Then uniform design test,CCK-8,wound-healing,migration and invasion experiment were used to obtain the best combination.And then,a tumor model was established to confirm the antitumor activity of Shuang-Huang-Lian MCCM.In addition,we established tumor-bearing mouse model and lung injury combined with tumor blood metastasis mouse model to elucidate the anti-tumor mechanism of Shuang-lluang-Lian MCCM.The results showed that Shuang-Huang-Lian MCCM could reduce the activation of CAFs in the tumor microenvironment,and significantly inhibit the migration and invasion of CT26 cells induced by CAFs.Using autodock technology,we found that Shuang-Huang-Lian MCCM could integrated well with LOXL2.And it also decreased LOXL2 expression in vivo and in vitro.In summary,our research found that LOXL2 expression is up-regulated in CC,which was associated with tumor proliferation and poor prognosis.LOXL2 derived from tumor cells could activate CAFs,which promoted the migration and invasion of CC cells in turn.Shuang-Huang-Lian MCCM could not only directly inhibited the proliferation,migration and invasion of CC cells in vivo and in vitro,but also indirectly affected the biological behavior of CC cells by disrupting the activation of CAFs,which may be related to the inhibition of LOXL2 in tumor microenvironment.Therefore,the results of this study provide a new target and a new idea for the clinical treatment of CC.