The Mechanism Of Drug Resistance In Pancreatic Cancer Caused By SMAD4 Gene Deletion

Objectives To detect the expression level of SMAD4 gene in pancreatic cancer AsPC-1and MIA PaCa-2 cells,and to explore the relationship between SMAD4 gene and pancreatic cancer resistance to gemcitabine,in an attempt to solve the clinical problem of ineffectiveness for drug-resistant pancreatic cancer.In order to provide new research ideas for the personalized medicine of pancreatic cancer.Methods 1 Bioinformatics Analysis of SMAD4 Gene expression in Pancreatic Cancer cells.2 CCK8 method was used to detect the toxic effect of different concentrations of gemcitabine on pancreatic cancer cells,and then the best concentration was selected.3Trypan blue exclusion test was used to detect the effect of gemcitabine on the survival rate of pancreatic cancer cells under different times,and then select the best action time.4 The m RNA and protein levels of SMAD4 gene were detected by real-time fluorescence quantitative PCR and Western blotting.5 The effects of silenced or overexpressed SMAD4genes on cell migration and invasion were detected by scratch test and Transwell test.Results 1 According to the cBio Portal database,33%of pancreatic cancer patients have SMAD4 gene mutations,Moreover,mutations were mostly deletions,missense mutations,and truncation mutations;the expression of SMAD4 gene in MIA PaCa-2 cells was significantly higher than that in AsPC-1 cells by high-throughput sequencing,and the difference was more than two-fold;meanwhile,the expression levels of SMAD4 gene in AsPC-1 cells m RNA and protein expression levels in AsPC-1 cells were also significantly lower than those in MIA PaCa-2 cells.2 The optimal concentration of gemcitabine acting on AsPC-1 cells was 10^-5 M,The optimal duration of action was 48 h;and the optimal concentration of gemcitabine acting on MIA PaCa-2 cells was 10^-7 M,the optimal duration of action was 48 h.3 Compared with the control group,the survival rate of gemcitabine-activated MIA PaCa-2 cells was significantly increased after silencing the SMAD4 gene;the number of migrating and invading cells was significantly lower in the spiked group compared with the PBS group.However,the number of migrating and invading cells was significantly upregulated in the transfected group compared with the non-transfected group.4 After overexpression of the SMAD4 gene,the survival rate of gemcitabineacting on cells in the transfected group was significantly lower than that in the control group,and the cell scratch area of the transfected plasmid-plus group was the slowest to decrease.The number of invaded cells passing through the small chamber wassignificantly reduced.Conclusions 1 SMAD4 gene was significantly different in AsPC-1 cells and MIA PaCa-2cells.2 SMAD4 gene deletion is associated with GEM resistance in pancreatic cancer cells.3 As a tumor suppressor gene,the SMAD4 gene can enhance the ability of GEM to inhibit the proliferation,migration and invasion of pancreatic cancer cells,thus providing a new method for solving the drug resistance of pancreatic cancer.Figure 12;Table 8;Reference 112...