The Regulation Of Long Non-Coding RNA HULC To PI3K/AKT Signaling Pathway Mediate The Malignant Progression Of Glioblastoma

BackgroundGlioblastoma multiforme?GBM?is a highly malignant intracranial tumor.Because of the rapid growth of tumors,and their recurrence,metastasis,and drug resistance,theprognosis is poor.The occurrence and development of tumors involve the regulation of a large number of signaling pathways and the abnormal expression of genes.Studies have shown that long non-coding RNAs?LncRNAs?are closely related to the occurrence and development of malignant tumors.Highly upregulated in liver cancer?HULC?,a member of LncRNA,was initially identified as being highly up-regulated in hepatocellularcarcinoma and gradually found in other malignancies.Our previous experiments showed that HULC levels in the GBM cell lines were significantly higher than normal brain tissue and could promote cell growth.Several studies have confirmed that the PI3K/AKTsignaling pathway contributes to the malignant progression of GBM.Therefore,whether the PI3K/AKT pathway that dominates cell growth is related to HULC is worth further studying.ObjectiveThe purpose of this study is to clarify the expression and clinical significance ofHULC in GBM,and to explore the mechanism of HULC regulating PI3K/AKT signaling pathway in GBM,in order to provide theoretical basis for subsequent targeted therapy and prognosis assessment.Methods?1?A total of 263 paraffin-embedded tissue specimens with pathological diagnosis of GBM were collected,and the tumor tissue was matched with normal brain tissue to make tissue microarrays.At the same time,the corresponding clinical data and follow-up data were collected.?2?The tissue microarrays were used to detect the expression of LncRNA HULC using the BaseScope^TM in situ hybridization method.The relationship between the expression level of HULC and the clinicopathological features of GBM and overall survival?OS?wasanalyzed.?3?The bioinformatics website was used to query the sequence information of HULC.The RNA-RNA relationship prediction algorithm was combined with the website data toanalyze the target genes that might be regulated by HULC.A network map of HULC and target genes was constructed,and the genes that may be related to the PI3K/AKT signaling pathway were screened out in combination with the TCGA signaling pathway database.?4?Using tissue microarrays,the expression of the target gene protein was detected by an automatic immunohistochemistry.At the same time,the protein levels of p-FAK?Tyr397?,PI3K,p-AKT?Ser473?and PTEN were detected.The correlation between the expression of this target gene protein and HULC and p-FAK?Tyr397?in GBM tissues was analyzed,as well as the correlation between p-FAK?Tyr397?and PI3K,p-AKT?Ser473?expression levels.Results?1?In the 263 pairs of GBM tumors and normal tissues adjacent to tumors,BaseScope^TMM in situ hybridization showed that there were 60 cases?22.8%?of high expression of HULC in normal brain tissues,and 173 cases?65.8%?in GBM tissues.There was a statisticallysignificant difference in the expression of HULC in tumors and normal tissues?P<0.01?.?2?Analysis of the relationship between HULC and clinicopathological features showed that the expression level of HULC was positively correlated with the age of patient and the length of recurrence??6 months or>6 months??P<0.01?,but there was no significant correlation with the patient's gender and tumor diameter?P>0.05?.?3?The relationship between HULC expression levels and overall survival?OS?in 263cases of GBM patients was statistically analyzed.The results showed that patients withhigh HULC expression had significantly shorter OS than those with low HULC expression?P<0.01?.Cox proportional hazards regression model showed that the age of patient and HULC expression level were independent risk factors affecting the prognosis of GBMpatients?P<0.01?.?4?According to the results of bioinformatics prediction,tenascin-R?TNR?was the target of this study,which may be regulated by HULC and also has a relationship withPI3K/AKT pathway.?5?In the 263 pairs of GBM tumors and corresponding normal tissues adjacent to thetumor,the immunohistochemistry results showed that the positive expression rate of TNR protein in tumor tissues was 23.6%?62/263 cases?,and the positive expression rate ofparatumor normal brain tissues was 81.7%?215/263 cases?,the difference between the two expressions was statistically significant?P<0.01?.?6?The immunohistochemistry results of p-FAK?Tyr397?,PI3K,and p-AKT?Ser473?showed that the expression levels in tumor tissues were significantly higher than those in corresponding normal brain tissues?P<0.01?.PTEN protein expression was significantly reduced in tumor tissues?P<0.01?.There was a positive correlation between p-FAK?Tyr397?and PI3K and p-AKT?Ser473?in tumor tissues?P<0.01?.?7?Spearman correlation analysis showed that TNR was significantly negatively correlated with the expression of HULC and p-FAK in GBM tissues?P<0.05?.Conclusions?1?LncRNA HULC promotes GBM progression and is positively correlated with the poor prognosis of patients,can be used as a new prognostic marker.?2?The PI3K/AKT signaling pathway plays a role in GBM tissues and may be associated with the activation of FAK.?3?The increase of HULC in GBM may down-regulate the expression of extracellular TNR,which stimulates the phosphorylation of FAK,thereby inducing the activation of downstream PI3K/AKT signaling pathways and mediating tumor biological processes.