| Research background:Gastric cancer is a common malignant tumor in digestive tract,and its incidence rate ranks fifth among malignant tumors in the world,and it is also the third leading cause of cancer-related death[1].The incidence of gastric cancer in China is the highest in the world,accounting for 49.9%of global cases[2].Although the survival rate of gastric cancer patients has been greatly improved in the past 20 years due to the development of surgery,chemotherapy and targeted therapy,the prognosis is still not satisfactory[3].The 5-year survival rate of gastric cancer patients in China is 30.2%to35.9%,while that in European countries is only 10%to 30%[4].The early symptoms of gastric cancer are atypical,many patients are late when they are found,and often lose the chance of surgery,and nearly half of patients with gastric cancer have recurrence after operation.At present,there is no effective treatment for these patients[5].In the past few decades,although some risk factors related to the occurrence of gastric cancer have been clarified,the research on the molecular mechanism of the occurrence and development of gastric cancer is not completely clear.Therefore,it is particularly urgent to study the molecular mechanism of the occurrence and development of gastric cancer and find new biomarkers and therapeutic targets.MicroRNAs（miRNAs）are single-stranded non-small coding RNA（snc RNA）,about 18-25 nucleotides,transcribed from introns or non-coding DNA.Micrornas are small molecules with no coding function,which play an important role in the regulation of gene expression[6].They can negatively regulate the expression of target genes through m RNA degradation or translation inhibition by pairing with complementary mirnas of 3’-untranslated region（3’-UTR）,coding sequence（CDS）or 5’-untranslated region（5’-URT）[7],including regulating cell division,proliferation,migration and so on.In human beings,the genes encoding miRNA only account for a very small part（about 3%）of the genes,but they can regulate the synthesis of protein in more than 30%of human beings[8].Studies have found that miRNAs are abnormally expressed in different degrees in many tumors,and these abnormally expressed mirnas regulate the occurrence and development of tumors by regulating proto-oncogenes or tumor suppressor genes in tumor cells[9].MiR-330-3p was first discovered by weber[10].miR-330 is located in human chromosome 19q13.32[11].MiR-330 is composed of two auxiliary chains,namely miR-330-3p and miR-330-5p,and their smoothness differs only by one nucleotide[12].The role of miR-330 transcript in regulating metastasis has been shown in various cancers,including colorectal cancer,metastatic lung cancer,prostate cancer,ovarian cancer and gastric cancer[13,14].It was found that the expression of miR-330-3p had a positive effect on cell invasion,migration and angiogenesis[15,16].Purpose:In this study,we will explore the expression level of miR-330-3p in gastric cancer cell lines and tissues,and study the biological function and mechanism of miR-330-3p in the proliferation,invasion and metastasis of gastric cancer cells in vitro:miR-330-3p regulates the expression of CDC42 in a targeted manner,and inhibits the proliferation,invasion and metastasis of gastric cancer cells,thus providing a theoretical basis for miR-330-3p in targeted therapy of gastric cancer.Method:In this study,the expression levels of miR-330-3p and CDC42 in gastric cancer cell lines and normal cell lines,gastric cancer tissues and adjacent tissues were detected by qRT-PCR and Western Blot,and the correlation between miR-330-3p expression levels and clinicopathological features of gastric cancer patients was analyzed.The bioinformatics technology and double luciferase gene report experiment verified that miR-330-3p directly regulated the expression of CDC42,and the transfection,CCK8experiment,scratch experiment and Transwell experiment verified the ability of miR-330-3p targeting CDC42 to inhibit the proliferation,invasion and metastasis of gastric cancer cells.Finally,the molecular mechanism of miR-330-3p targeting CDC42inhibiting the proliferation,invasion and metastasis of gastric cancer cells was verified by animal model.Results:In gastric cancer cell lines and tissues,the expression level of miR-330-3p was significantly down-regulated（P<0.05）.The low expression of miR-330-3p was significantly correlated with the depth of tumor invasion,the number of lymph node metastasis and clinical stage（P<0.05）,and the low expression of miR-330-3p was significantly correlated with the poor prognosis of gastric cancer patients（P<0.05）.We confirmed that miR-330-3p directly regulated the expression of CDC42 through bioinformatics analysis.qRT-PCR and Western Blot showed that the expression level of CDC42 in gastric cancer tissues and gastric cancer cell lines was significantly increased（P<0.05）,and the expression level of CDC42 was down-regulated after transfection of miR-330-3p,and the double luciferase gene report also confirmed that miR-330-3p targeted regulated CDC42 and inhibited it.Finally,the results of CCK8experiment,scratch experiment and Transwell experiment all confirmed the ability of miR-330-3p targeting CDC42 to inhibit the proliferation,invasion and metastasis of gastric cancer cells,and the mouse animal model experiment also verified this result.Conclusion:In gastric cancer tissues and gastric cancer cell lines,the expression level of miR-330-3p is obviously down-regulated,and the low expression of miR-330-3p indicates the poor prognosis of gastric cancer patients.MiR-330-3p targeted down-regulated the expression of CDC42 and inhibited the proliferation,invasion and metastasis of gastric cancer cells.These results all prove the molecular mechanism of miR-330-3p targeting CDC42,which enriches the research on the occurrence and development of gastric cancer,and also shows that miR-330-3p is expected to become a new biomarker for diagnosis and treatment. |
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