Effect Of SLC1A5 On The Biological Behavior Of Gastric Cancer And Its Relation With The Clinicopathological Features Of Gastric Cancer

BackgroundGastric cancer(GC)is a common malignant tumor in the digestive system.GC is still the second major cause of cancer-related death in the world.In view of the high mortality and poor prognosis of advanced gastric cancer patients,the research on the mechanism of the occurrence and development of gastric cancer still leaves us some problems to be solved.We should find new targets from gastric cancer tissues and cells,further elaborate the mechanism of the occurrence and development of gastric cancer,and provide new ideas for clinical diagnosis and treatment of gastric cancer.The abnormal energy metabolism of malignant tumor is also considered as one of the ten major features of malignant tumor.The new view is that malignant tumors are also a metabolic disease.Exuberant glutamine metabolism is an important feature of malignant tumor.The tumor requires more glutamine intake to promote its rapid proliferation.Extracellular glutamine must be transported by a series of amino acid transporters on cell membrane to enter cells.Therefore,amino acid transporters are likely to be upregulated in tumor tissues.The 1 member of the solute carrier family 5(Solute Carrier Family 1 Member SLC1A5),is an important member of the amino acid transport system,which transport a variety of small molecular neutral amino acids,including glutamine.Many studies have shown that SLC1A5 is highly expressed in many malignant tumor tissues and cells,and is related to tumor proliferation,invasion and poor prognosis.In vivo and in vitro studies indicate that inhibition of SLC1A5 can inhibit tumor cell growth.There is no research on SLC1A5 and gastric cancer at present.Our study was divided into three parts.Part one aimed to explore the expression of SLC1A5 in GC and analyze its relation with the prognosis and the clinic features of GC.Part two aimed to investigate the effect on the malignant biological behavior of gastric cancer cells and its molecular mechanism in vitro.Part three aimed to study the effects of SLC1A5 on the growth of GC in vivo.Part one The expression of SLC1A5 in gastric cancer and its relation with the clinicopathological features of gastric cancerObject:To observe the expression of SLC1A5 in GC and analyze its relation with the clinicopathological features of GC.Method: 70 cases of advanced gastric cancer tissues were selected as experiment group,and corresponding paracancerous tissues were showed as control group.Immunohistochemistry were used to observe the protein expression levels of SLC1A5 in two groups.We analyzed the relationship between SLC1A5 and the expression level of clinicopathological features.By using gene database,we also analyze the expression level of SLC1A5 in gastric cancer tissues and adjacent tissues,and its relation with prognosis of GC.Results: Compared to the control group,the expression levels of SLC1A5 significantly increased in advanced GC(P<0.0001).statistic analysis by using GSE database also confirmed that the expression levels of SLC1A5 significantly increased in GC tissues compared to the paracancerous tissues(GSE 65801,P =0.0046;GSE 63809,P<0.0001;GSE 27342,P=0.0147).The expression level of SLC1A5 in gastric cancer tissues was correlated with tumor size(P<0.05),depth of invasion(P<0.001),lymph node metastasis(P<0.05),TNM stage(P<0.05)and Ki-67 index(P<0.01).It was not related to age,sex,tumor location and degree of differentiation.Higher expression level of SLC1A5 in GC is associated with poor prognosis,which can significantly shorten the overall survival time(GSE 14210,P=0.0.011,GSE 22377,P=0.0.0015),and the progression free survival time of GC patients(GSE 14210,P=0.0.0095;GSE 22377,P=0.0.0012).Conclusion:SLC1A5 protein expression in GC tissue increased,and the expression of SLC1A5 is associated with tumor size,depth of invasion,lymph node metastasis,TNM stage in advanced GC.SLC1A5 would be a promising target for diagnosis and prognosis of gastric cancer.Part two Effect of SLC1A5 on the malignant biological behavior of gastric cancer cells and its molecular mechanismObjective: To study the effect of SLC1A5 on the proliferation,cell cycle,cell invasion and cell migration of human gastric cancer cells.And to explore the relationship between SLC1A5 and mTOR /p70S6 K signaling pathway.Methods: Western blot were performed to detect expression levels of SLC1A5 in five different gastric cancer cell lines.p GU6/GFR/Neo vectors containing sh RNA suppressing SLC1A5 m RNA were used to transfect GC cells.CCK-8 assay were used to detect cell viability in GC cells.Flow cytometry were used to analyze cell cycle.Cell invasion and migration assay were performed to observe the cell invasion by transwell.Western blot were performed to determine the expression levels of mTOR,p-mTOR,p70S6 K,p-p70S6 K and Cyclin D1.Results: Higher expression levels of SLC1A5 in gastric cancer cell lines HGC-27 and MGC-803 were identified by Western blot.Knockdown SLC1A5 inhibited the proliferation of GC cells,lead to G0/G1 phase arrest,reduced cell invasion and migration,inhibited m TOR /p70S6 K signaling pathway,and downregulated the expression level of Cyclin D1.Conclusion: SLC1A5 can promote the growth and invasion of GC cells.The mechanism is related to the promotion of the m TOR /p70S6 K signaling pathway.SLC1A5 would be a new therapeutic target of GC.Part three Effect of SLC1A5 on tumor growth in vivoObjective: To study the effect of SLC1A5 on the growth of subcutaneous tumor in mice.Methods: 4-6 week old male BALB/c nude mice were randomly divided into two groups,knockdown-SLC1A5 HGC-27 cell suspension(experimental group)and Negative control cell suspension(control group)were injected into the lower edge of the scapula on the back side of the mice.The length and diameter of tumor in nude mice were regularly measured,and tumor volume was calculated at the end.Tumor tissues were isolated after Nude mice sacrificed 20 days later.The expression levels of SLC1A5 in tumor tissues was detected by Western blotting.Results: SLC1A5 obviously promoted the growth of tumor in mice(P<0.0001),and the size of the tumor was significantly different at tenth days after the tumor was successfully loaded(P<0.0001).Compared to the control group,the expression levels of SLC1A5 in the tumor tissue of the experimental group was significantly downregulated.Conclusion: SLC1A5 can promote the growth of GC in vivo.