The Analysis Of Hsa＿circ＿0035445 Diagnostic Value In Gastric Cancer

ObjectivesGastric cancer（GC）is one of the most common gastrointestinal malignant tumors in China.According to statistics,the incidence and mortality of gastric cancer in China ranks 3rd in the incidence and mortality of malignant tumors,and has obvious regional characteristics.However,the sensitivity and specificity of carcinoembryonic antigen（CEA）,a commonly used clinical diagnostic marker for gastric cancer,are only 0.6 and 0.3,respectively.It can be seen that it is particularly important to find ideal diagnostic markers for gastric cancer,especially early diagnosis.Circular RNA（circ RNA）is a special type of RNA that is covalently closed in a circular shape.The study found that circ RNA is closely related to the occurrence and development of gastric cancer,and can be stable in gastric cancer tissues and plasma,and is expected to become a new marker for clinical diagnosis of gastric cancer.Methods1.Screen the target circ RNA（hsa＿circ＿0035445）according to the circ RNA microarray and the circ RNA database（circ Base）.2.Collect 104 pairs of gastric cancer tissues and paired non-cancer tissues,also collect fasting plasma of 42 normal people and 42 gastric cancer patients 1 day before and 10 days after operation.3.Using q RT-PCR technology,that is,quantitative reverse transcription-polymerase chain reaction,to detect the expression level of hsa＿circ＿0035445 in the above samples and gastric cancer cells,respectively.4.Apply small interfering RNA（si RNA）technology to down-regulate the expression of hsa＿circ＿0035445;design an over-expression vector for hsa＿circ＿0035445 to up-regulate its expression,and use the transfection reagent Lipofectamine 2000 to transfer into cells.Therefore,the effect of hsa＿circ＿0035445 on the occurrence of gastric cancer was studied at the cellular level.5.After cell transfection,changes in gastric cancer cells proliferation after up-regulated and down-regulated circ RNA hsa＿circ＿0035445 were detected by Cell Counting Kit-8（CCK-8）and colony formation assays;The effects of up-and down-regulated circ RNA hsa＿circ＿0035445 on gastric cancer cell migration ability was examined by Trans-well assay and scratch assay.Flow cytometry was used to detect changes in gastric cancer cell cycle and apoptosis after up-and down-regulated hsa＿circ＿0035445.Results1.The results of q RT-PCR showed that the expression level of hsa＿circ＿0035445 in gastric cancer tissues was lower than that of the corresponding paracancer tissues.The plasma of patients with gastric cancer compared with the plasma of 1 day before surgery was low compared with 10days after surgery,1 day of gastric cancer patients with preoperative plasma compared with healthy volunteer plasma,and expression level of hsa＿circ＿0035445 in the gastric cancer cells were also low compared with normal gastric mucosal epithelial cell（GES-1）.2.Further analysis found that the area under the ROC curve（AUC）of receiver operating characteristic（ROC）of hsa＿circ＿0035445 in the tissues of gastric cancer patients was 0.68;the AUC in the plasma of gastric cancer patients was 0.86,after the combination of the two,the AUC can increase to 0.88,It can be seen that hsa＿circ＿0035445 has a certain clinical diagnostic value and is hopeful to become a new type of biomarker for gastric cancer diagnosis.3.After down-regulation the expression of hsa＿circ＿0035445,the results of CCK-8 and cell clone formation assays demonstrated that the proliferation ability of gastric cancer cells（AGS and BGC）was enhanced;the Trans-well assay and scratch assay results found that the migration ability of gastric cancer cells（AGS and BGC）was increased;the results of flow cytometry revealed that the apoptosis of gastric cancer cells（AGS and BGC）could be reduced and the cells were blocked in the S phase.4.After the up-regulation the expression of hsa＿circ＿0035445,the results of CCK-8 and cell clone formation assays found that the proliferation ability of gastric cancer cells（AGS and BGC）was weakened;the Trans-well assay and scratch assay results showed that the migration ability of gastric cancer cells（AGS and BGC）was reduced;the results of flow cytometry indicated that it could promote the apoptosis of gastric cancer cells（AGS and BGC）and block the cells in the G₀/G₁ phase.Conclusions1.hsa＿circ＿0035445 showed low expression in gastric cancer tissues,plasma and gastric cancer cells.2.The ROC curves showed that hsa＿circ＿0035445 has certain diagnostic value in gastric cancer,and it is hopeful to become a new marker for clinical diagnosis of gastric cancer.3.In vitro cell level experiments showed that down-regulation the expression of hsa＿circ＿0035445 can enhance the proliferation and migration of gastric cancer cells and can reduce the apoptosis of gastric cancer cells and gastric cancer cells are blocked in S phase;while up-regulation the expression of hsa＿circ＿0035445 can reduce the proliferation of gastric cancer cells and migration ability can promote gastric cancer cell apoptosis and gastric cancer cells are blocked in G₀/G₁ phase.This indicates that hsa＿circ＿0035445 may become a new target for clinical treatment of gastric cancer.In summary,hsa＿circ＿0035445 has the potential for clinical diagnosis of gastric cancer,it may be used as a novel marker molecule,or it may become a new target for guiding clinical treatment of gastric cancer.