Study On The Mechanisms Of SRPX2 Responsible For Invasion And Metastasis In Pancreatic Cancer

Bankground: Pancreatic ductal adenocarcinoma(PDAC)is a major cause of cancer-associated mortality,with a dismal overall prognosis that has remained virtually unchanged for many decades.Currently prevention or early diagnosis at a curable stages is exceedingly difficult;patients rarely exhibits symptoms and tumours do not display sensitive and specific markers to aid detection.Pancreatic cancers also have few prevalent genetic mutations;the most commonly mutated genes are KRAS,CDKN2 A,TP53 and SMAD4—none of which are currently druggable.Indeed therapeutic options are limited and progress in drug development is impeded because most pancreatic cancers are complex at the genomic,epigenetic and metabolic levels,with multiple activated pathways and crosstalk evident.Fewer than 20% of patients have surgically resectable disease,however,neoadjuvant therapies might shift tumours towards respectability.Although newer drug combinations and multimodel regimens in this setting,as well as the adjuvant setting,appreciably extend survival.Microscopically,the tumor is characterized by thick desmoplastic stroma that surrounds islands of pancreatic cancer cells.The tumor microenvironment has been found to be of importance in carcinogenesis,the development of drug resistance,and mediating immunosuppression.The understanding the tumor-stroma interaction has led to the development of novel therapeutic approaches.Sushi repeat-containing protein linked 2(SRPX2),is a chondroitin sulfate proteoglycan with the feature of extracellular matrix proteins and a candidate downstream target protein for E2A-HLF.It has been discovered that SRPX2 can promote cell migration,adhesion and mediating angiogenesis.Recent studies suggest that SRPX2 may play an important role in the development of gastrointestinal tumors,especially in the migration and invasion.However,its function on the invasion and metastasis of pancreatic cancer isn't clear by now.Therefore,it is important to investigate the action of SRPX2 in the invasion and metastasis of pancreatic cancer and to master the molecular mechanism of pancreatic cancer.Objective: The aims of current study were to investigate the expression of SRPX2 in PDAC and the correlations between relative expression of SRPX2 and clinic-pathologic information of these pancreatic patients.As well as to explore the role of SRPX2 on cell migration,invasion,viability,apoptosis in pancreatic cancer in vitro.Materials and methods:(1)RT-PCR and Western blotting were used to detect expression of SRPX2 gene in pancreatic cancer cell lines,AsPC1?Capan2?CFPAC1?HPAC?PANC1?SW1990 and human pancreatic ductal epithelial cells(hTERT-HPNE).(2)RT-PCR was employed to detect expression of SRPX2 gene in 12 pairs of pancreatic cancer tissues and the tumor-adjacent non-malignant tissues.Immunohistochemistry(IHC)analysis was performed that were made from 81 cases of pancreatic cancer tissue specimens to analyse the expression of molecular markers including SRPX2 gene and phosphorylation levels of FAK as well as SRPX2 gene was detected in 44 cases of the tumor-adjacent non-malignant tissues.Then correlations between relative expression of SRPX2 and clinic-pathologic information of these pancreatic patients were analyzed by different statistical methods.At the same time the correlation was analyzed between relative expression of SRPX2 and the phosphorylation levels of FAK.(3)PANC1 and SW1990,were transfected with two siRNAs(si-1 and si-2)target SRPX2.We performed western blot to study the expression of SRPX2 on the knockdown of pancreatic cancer cells in vitro.Transwell cell migration assay and Matrigel cell invasion assay were applied to study the role of SRPX2 on metastasis of pancreatic cancer cells in vitro.CCK-8 assay was used to assess the role of SRPX2 on cell viability of pancreatic cancer in vitro.Result: The expression of SRPX2 at mRNA level was up-regulated in all of six(1)PDAC cell lines compared with hTERT-HPNE.This trend was also confirmed by protein expression as shown by western blotting.(2)The expression of SRPX2 at the mRNA level was significantly up-regulated in PDAC tumor tissues compared with corresponding non-tumor tissues.SRPX2 protein was significantly up-regulated in PDAC tumor tissues.Besides,high expression of SRPX2 was remarkably associated with advanced TNM stage(P = 0.001),no significant difference was found in age,gender,tumor size,tumor location,chronic pancreatits,smoking,diabetes and neuronal invasion using the Chi-square test.SRPX2 expression levels in PDAC tissues significantly correlated with the levels of phosphor-FAK.(3).PANC1 and SW1990,were transfected with two siRNAs(si-1 and si-2)target SRPX2;the protein expression of SRPX2 was markedly decreased after both siRNAs treatment.Migration assay showed that the number of migratory cells in siRNA treated group was dramatically decreased compared with those in the negative control group in PANC1 cells.the invaded cells were also decreased significantly after SRPX2 knockdown in PANC1 cells.No significant difference was found in the cell viability or cell apoptotic ratio between siRNA treated group and negative control group.Conclusions: SRPX2 is highly expressed in pancreatic cancer cell lines and tissues,and is associated with TNM staging of pancreatic cancer.SRPX2 enhances the migration and invasion of cancer cells and has nothing to do with cell proliferation and apoptosis.SRPX2 raised the level of phosphorylated FAK.SRPX2 and FAK signaling pathway may play an important role in the occurrence and development of pancreatic cancer.This study may targeted treatment for pancreatic cancer to provide new clues.