Effect Of Linc-pint On Proliferation Of Gastric Cancer Cells And Its Mechanism Of Action

Abnormal accelerated growth of cancer cells is the basis for the development and progression of different types of cancer.Inhibition of cancer cell growth is currently considered to be the most promising therapeutic target for the treatment of cancer.Gastric cancer is one of the most common malignancies.In China,gastric cancer is the third most common malignant tumor,and its incidence is after lung cancer and breast cancer.Early symptoms of gastric cancer are concealed,and most of them are diagnosed at mid-to-late stage,which seriously affects the health of patients.Although a series of chemical drugs have been developed to inhibit the growth of gastric cancer cells,the overall therapeutic effect is still unsatisfactory due to the unclear tumor growth mechanism.Therefore,the identification of novel molecular targets capable of inhibiting the growth of gastric cancer is of great significance.Signal transduction of hypoxia-inducible factor-1??HIF-1??plays a key role in the development,progression and progression of cancer and is considered to be a target for cancer therapy.Hypoxia is a core regulator of tumor growth,and the role of HIF-1?in the growth of gastric cancer cells has been reported.Long intergenic non-protein encoding long-chain RNA p53-induced transcript?Linc-pint?is a newly discovered long-chain non-coding RNA?lncRNA?that has potential tumor suppressor function in different types of cancer.It is well known that HIF-1?interacts with lncRNAs in some cases and participates in physiological or pathological processes.In this study,we demonstrate that Linc-pint may inhibit the growth of gastric cancer by down-regulating HIF-1?.Part one Correlation study and survival analysis on Linc-pint and gastric cancer patientsObjective:1.T o analyze the correlation between the expression level of Linc-pint and clinical data of patients with gastric cancer.2.To clarify the relationship between Linc-pint expression and survival of patients with gastric cancer.Methods:1.According to the median expression level of Linc-pint,52 patients with gastric cancer were divided into Linc-pint high expression group?n=26?and Linc-pint low expression group?n=26?.2.ROC curve analysis was used to evaluate the diagnostic value of Linc-pint expression in gastric cancer.3.Chi-square test was used to analyze the correlation between Linc-pint expression and clinical data of patients with gastric cancer.4.Following-up the PFS of 42 patients received first-line chemo-therapy and make survival analysis based on clinical data.Results:1.Linc-pint expression was significantly associated with tumor size?P=0.03?and pathological grades?P=0.007?,but it was independent of whether the tumor had distant metastases,lymph node metastasis,age and gender.2.The area under the ROC curve is 0.8910,the standard error is 0.03153,and the 95%confidence interval is 0.8291-0.9528.It shows that Linc-pint has a high value in the diagnosis of gastric cancer.3.The median PFS of high Linc-pint expression group was 8.25±1.11months,while that of low expression group was 6.37±0.65 months,P=0.163.4.PFS in the high expression group of Linc-pint was better than that in the low expression group for female patients?P=0.042?Part two Correlation between Linc-pint and HIF-1?in tumor tissues of gastric cancerObjective:1.To detect the expression of Linc-pint and HIF-1?in tumor tissues of gastric cancer patients,and to analyze the relationship between expression levels.2.Clarify the correlation between HIF-1?and clinical data and make survival analysis.Methods:1.qRT-PCR analysis of the expression of Linc-pint and HIF-1?mRNA in gastric cancer tissues of gastric cancer patients and gastric tissues of healthy controls.2.Correlation analysis of Linc-pint and HIF-1?mRNA expression was analyzed by Pearson correlation coefficient.3.Chi-square test was used to analyze the correlation between HIF-1?expression and clinical data of patients with gastric cancer.4.Following-up the PFS of 42 patients received first-line chemo-therapy and make survival analysis based on clinical data.Results:1.The expression levels of HIF-1?mRNA in gastric cancer tissues and normal gastric tissues were?3.32±0.87?and?2.12±0.63?,respectively.P<0.05.2.HIF-1?expression was significantly associated with tumor size,P=0.01.3.HIF-1?expression was significantly associated with grading of lymph node metastasis,P=0.007.4.There was no correlation between HIF-1?expression and PFS in patients with gastric cancer.5.The higher the expression level of Linc-pint in gastric cancer patients,the lower the HIF-1?mRNA level;the lower the expression level of Linc-pint,the higher the HIF-1?mRNA level,there is a significant inverse correlation bet-ween them?P<0.0001?.In contrast to this result,no significant correlation between Linc-pint and HIF-1?mRNA was found in healthy controls?P=0.05484?.Part three The mechanism of action of Linc-pint on the growth of gastric cancer cellsObjective:To examine the role of Linc-pint in the proliferation of gastric cancer cells and to conduct further research on its mechanism of action.Methods:1.Linc-pint overexpressing SNU-1 and AGS cell lines were established by cell transfection technique;2.Western blotting was used to detect the expression of HIF-1?under normal conditions and the expression of HIF-1?under hypoxia?1%O₂??3.The effects of HIF-1?on the expression of Linc-pint were detected by SMN-1 and AGS cells treated with exogenous human recombinant HIF-1??Sigma-Aldrich?at 1,10,50 and 100 ng/ml,respectively;4.The effects of Linc-pint overexpression,HIF-1?and both on the growth of gastric cancer cells were detected by CCK-8 kit.Results:1.Compared with control cells?C?and negative control cells,SNU-1 and AGS cells with Linc-pint overexpression showed significant down-regulation of HIF-1?protein levels under normal conditions?P<0.05??2.Under hypoxic conditions?1%O₂?,HIF-1?protein levels in Linc-pint overexpressing SNU-1 and AGS cells were still lower than those in control and negative cells?P<0.05??3.Different concentrations of HIF-1?had no significant effect on Linc-pint expression under normal conditions or under hypoxic conditions?4.Compared with the control cells and the negative control cells,the proliferation of Linc-pint overexpressing gastric cancer cells decreased;after the addition of exogenous HIF-1?,the proliferation of gastric cancer cells increased.Compared with Linc-pint overexpressing gastric cancer cells,Linc-pint overexpressing gastric cancer cells were inhibited by proliferative ability after addition of exogenous HIF-1?,but still lower than control cells and negative control cells.Conclusion:1.The expression of Linc-pint in gastric cancer tissue was lower.The decrease of Linc-pint expression is a sensitive index in gastric cancer tissues,which may be of great significance for the prognosis of gastric cancer.2.The expression of Linc-pint in gastric cancer was negatively correlated with the tumor size and pathological grade of primary tumors.3.The median PFS was tends to prolonged in the Linc-pint high expression group.4.In female subgroup,PFS was tends to prolonged in the Linc-pint high expression group.it was suggested that gender differences may influence the expression of Linc-pint.5 Compared with healthy tissues,HIF-1?expression was higher in gastric cancer tissues.HIF-1?expression was positively correlated with tumor size and the grade of lymph node metastasis,and it was an important prognostic indicator.6.Survival analysis showed no correlation between the expression of HIF-1?and PFS in gastric cancer patients.7.There was a significant negative correlation between Linc-pint and HIF-1?.These results suggested that overexpression of Linc-pint can inhibit the growth of gastric cancer cells by down-regulating HIF-1?.