Study On The Mechanism Of Inhibiting Breast Cancer Metastasis By Sweet Potato Daucosterol Linoleate

In the past 10 years,cancer has been a major public health problem in the world,and has become the third largest cause of death in China and the second largest cause of death in the United States.In recent years,the incidence of breast cancer in China has shown an upward trend,reaching a peak at the age of 55,in which the incidence and mortality in urban areas are higher than those in rural areas.the metastasis of breast cancer in the later stage is the main cause of the death of breast cancer patients.However,so far,our basic theoretical research on metastatic breast cancer is not sufficient,and there is a lack of effective detection targets and therapeutic drugs in clinic.Metastatic breast cancer is a worldwide problem.At present,the treatment of breast cancer metastasis is radiotherapy and chemotherapy,but it is prone to drug resistance and side effects,so it is necessary to develop safe and effective breast cancer adjuvant drugs with low side effects.Sweet potato as the seventh largest economic crop in the world,its roots,stems and leaves are edible,and contains a variety of ingredients beneficial to human health,such as glycoprotein,carotene and so on.The previous study of our group found that Daucosterol Linoleate（DL）in sweet potato extract had a significant inhibitory effect on lung metastasis of breast cancer.The purpose of this study was to explore the mechanism of inhibitory effect of DL on breast cancer metastasis.This study mainly carries out experiments from the following aspects:（1）To investigate the inhibitory effect of DL on breast cancer cell metastasis in vitro.MTT assay was used to evaluate the effects of DL on the viability of breast cancer cells 4T1,MCF-7,and MDA-MB-231.MCF-7 cells were shown to be more sensitive to DL treatment than the other two types of breast cancer cells.The activity of 4T1 cells was weakly inhibited by DL（IC50:96.45μM）,followed by MDA-MB-231（IC50:93.00μM）and MCF-7（IC50:79.84μM）.The inhibitory effect of DL on the migration ability of three cell lines was investigated using a cell scratch test and a Transwell migration test.MCF-7 and MDA-MB-231 were reported to have a significant inhibitory effect on 4T1 migration,followed by DL.（2）To investigate the anti-metastatic effect of DL in vivo on breast cancer.The anti-metastatic impact of DL on MDA-MB-231 breast cancer in vivo was demonstrated using a tail vein injection lung metastasis model in nude mice with highly metastatic MDA-MB-231 breast cancer cells.The results demonstrated that a gavage dose of 87.8mg/kg of DL had no effect on the body weight of nude mice.In the tail vein injection lung metastasis model of MDA-MB-231 breast cancer nude mice,H&E staining of lung tissue sections suggested that the inhibition rate of DL could reach 63.16%.（3）DL’s potential targets for preventing breast cancer metastasis are being investigated.The GSEA gene set enrichment analysis of the preceding group of tumor-bearing nude mice’s lung metastasis proteomics revealed that epithelial mesenchymal-transformation and early estrogen response were significantly associated with the DL group.VAMP3,TAP2,CHK1,SCD1,and LOXL2 were identified as potential DL targets after screening,while SCD1 was screened as a potential target following q PCR validation and molecular docking to construct the DL binding model.（4）The goal of this study was to look at the interaction between DL and SCD1 in the prevention of breast cancer metastasis.q PCR and WB were used to detect the m RNA and protein expression levels of SD1 in three cell lines,4T1,MCF-7,and MDA-MB-231,and the MDA-MB-231 cells with the highest expression level were chosen for further investigations.MTT was used to estimate the IC50 of MDA-MB-231cells treated for 24 hours with the SCD1 protein inhibitor PluriSIn1.The effect of DL on the migration ability of MDA-MB-231 cells given after overexpression of SCD1 was investigated using a cell scratch assay and a Transwell migration assay.The results showed that the cells’migration ability was significantly increased after overexpression of SCD1,and that DL treatment effectively reduced the cells’migration rate to only 49.4%-67.5%of that of normal MDA-MB-231 cells,indicating that DL could reverse the enhanced ability.（5）The expression of EMT pathway marker proteins is regulated by DL.WB detection of protein expression of EMT pathway marker proteins N-Cadherin,MMP2,Vimentin,Snail,and ER showed a dose-dependent decrease,and SCD1 expression was similarly decreased in MDA-MB-231 treated with 1/2IC50_24h and IC50_24hconcentrations of DL and PluriSIn1,and WB detection of protein expression of EMT pathway marker proteins N-Cadherin,MMP2,Vimentin,Snail.Overexpression of a protein can be detected by Western blotting.WB was used to detect the expression of EMT pathway marker proteins in MDA-MB-231 overexpressing SCD1,and it was discovered that while all of these proteins appeared to be upregulated after overexpression of SCD1,their expression levels decreased to be similar to Victor after treatment with DL or PluriSIn1.（6）To see if DL reduces SCD1 expression and thereby suppresses breast cancer metastasis in vivo.The oe-SCD-MDA-MB-231 cell line was created using a lentiviral packaging system,and a lung metastasis model of breast cancer was established in nude mice by tail vein injection.The DL group was initiated by gavage one week after inoculation with oe-SCD-MDA-MB-231,and PluriSIn1 was administered by intraperitoneal injection as a positive medication targeting SCD1,and lung tissues of nude mice were taken after three weeks.When DL and PluriSIn1 were stained,it was discovered that they effectively decreased oe-SCD-MDA-MB-231 lung metastasis in nude mice,reduced the frequency of lung metastatic nodules,and suppressed the size of metastatic foci when compared to the oe-SCD group.In conclusion,this research concluded that DL has anti-metastatic effects on breast cancer and that it is involved in reducing breast cancer cell migration via decreasing SCD1 expression.The expression of SCD1 was positively correlated with the expression of EMT pathway marker protein in a series of experiments using oe-SCD-MDA-MB-231,and DL may suppress EMT and breast cancer metastasis by decreasing SCD1 expression.