| Objective:Gastric cancer is one of the top three malignant tumors in China.Metastasis is the most important cause of death in gastric cancer.Exploring the mechanism of gastric cancer metastasis is of great significance for monitoring the condition and prolonging the survival of patients.GRIM-19 expression is down-regulated in many malignancies.Previous studies have found that GRIM-19 is significantly down-regulated in gastric cancer tissues and chronic atrophic gastritis tissues of precancerous lesions,nevertheless,expression profile,clinical relevance and the mechanism underlying GRIM-19 regulates gastric cancer metastasis remain largely unknown.In this paper,we analyze the expression of GRIM-19 in gastric cancer tissues and demonstrate its specific role and mechanism in regulating gastric cancer metastasis.Methods:TCGA and NCBI GEO databases were used to analyze the relationship between GRIM-19 expression level,clinical relevance and survival rate in gastric cancer.CRISPR/Cas9 lentivirus was used to transfect gastric cancer cell lines SGC-7901 and MKN-45 to specifically knock out GRIM-19 expression,and flow cytometry was used to establish stably transfected cell lines.Cell migration and invasion experiments were performed to detect the effect of GRIM-19 deficiency on the migration and invasion ability of gastric cancer cells,and corresponding animal metastasis models were constructed.Flow cytometry was used to detect ROS and mROS in GRIM-19-deficient gastric cancer cell lines,and kits were used to detect changes in NADP~+/NADPH and GSH/GSSG.NRF2 and its downstream HO-1 and metastasis-related molecules in GRIM-19-deficient gastric cancer cells were detected by luciferase reporter gene,immunofluorescence,real-time PCR and western blot,and the corresponding specific inhibitors or activators were used to perform rescue experiments;HE and immunohistochemistry were used to detect the metastasis of the corresponding animal metastasis model and the expression of related proteins.Results:In gastric cancer clinical tissues,GRIM-19 was down-regulated in gastric cancer tissues compared with normal adjacent tissues and related to poor clinical outcomes,and DNA methylation was involved in its down-regulation.Deficiency of GRIM-19 in gastric cancer cell significantly promotes gastric cancer metastasis in vitro and in vivo.Clearance of ROS and inhibition of NRF2 and HO-1 expression can reverse GRIM-19 metastasis-promoting effect.Deletion of GRIM-19 in gastric cancer cells can increase mitochondrial ROS and intracellular ROS,which induce excessive oxidative stress,and activate NRF2 and its downstream HO-1 and other metastasis-related molecules promoting gastric cancer metastasis.Furthermore,HO-1 maintains NRF2 activation by NRF2-HO-1positive-feedback loop,resulting in metastasis-associated genes expression,thereby promoting gastric cancer metastasis.HO-1 acts as a key downstream molecule in GRIM-19 deletion via activating ROS-NRF2 to promote gastric cancer metastasis.Conclusion:Mitochondrial GRIM-19 deficiency promotes gastric cancer metastasis via ROS-NRF2-HO-1 axis,and HO-1 feedback maintains the oncogenic role of NRF2 in promoting metastasis,resulting in poor clinical outcomes.GRIM-19 and NRF2-HO-1 axis can be used as new therapeutic targets for clinical monitoring and interventional treatment of gastric cancer metastasis. |
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