Dataset Information

Minichromosome maintenance 3 promotes hepatocellular carcinoma radioresistance by activating the NF-?B pathway.

ABSTRACT:

Background

Hepatocellular carcinoma (HCC) is the most common tumors in the worldwide, it develops resistance to radiotherapy during treatment, understanding the regulatory mechanisms of radioresistance generation is the urgent need for HCC therapy.

Methods

qRT-PCR, western blot and immunohistochemistry were used to examine MCM3 expression. MTT assay, colony formation assay, terminal deoxynucleotidyl transferase nick end labeling assay and In vivo xenograft assay were used to determine the effect of MCM3 on radioresistance. Gene set enrichment analysis, luciferase reporter assay, western blot and qRT-PCR were used to examine the effect of MCM3 on NF-?B pathway.

Results

We found DNA replication initiation protein Minichromosome Maintenance 3 (MCM3) was upregulated in HCC tissues and cells, patients with high MCM3 expression had poor outcome, it was an independent prognostic factor for HCC. Cells with high MCM3 expression or MCM3 overexpression increased the radioresistance determined by MTT assay, colony formation assay, TUNEL assay and orthotopic transplantation mouse model, while cells with low MCM3 expression or MCM3 knockdown reduced the radioresistance. Mechanism analysis showed MCM3 activated NF-?B pathway, characterized by increasing the nuclear translocation of p65, the expression of the downstream genes NF-?B pathway and the phosphorylation of IKK-? and I?B?. Inhibition of NF-?B in MCM3 overexpressing cells using small molecular inhibitor reduced the radioresistance, suggesting MCM3 increased radioresistance through activating NF-?B pathway. Moreover, we found MCM3 expression positively correlated with NF-?B pathway in clinic.

Conclusions

Our findings revealed that MCM3 promoted radioresistance through activating NF-?B pathway, strengthening the role of MCM subunits in the tumor progression and providing a new target for HCC therapy.

SUBMITTER: Yang Q

PROVIDER: S-EPMC6580494 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Publications

Minichromosome maintenance 3 promotes hepatocellular carcinoma radioresistance by activating the NF-κB pathway.

Yang Qing Q Xie Binhui B Tang Hui H Meng Wei W Jia Changchang C Zhang Xiaomei X Zhang Yi Y Zhang Jianwen J Li Heping H Fu Binsheng B

Journal of experimental & clinical cancer research : CR 20190617 1

<h4>Background</h4>Hepatocellular carcinoma (HCC) is the most common tumors in the worldwide, it develops resistance to radiotherapy during treatment, understanding the regulatory mechanisms of radioresistance generation is the urgent need for HCC therapy.<h4>Methods</h4>qRT-PCR, western blot and immunohistochemistry were used to examine MCM3 expression. MTT assay, colony formation assay, terminal deoxynucleotidyl transferase nick end labeling assay and In vivo xenograft assay were used to deter ...[more]

PMID: 31208444

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Project description:BackgroundHepatocellular carcinoma (HCC) is the sixth most common cancer and the second leading cause of cancer-related deaths worldwide. Despite new technologies in diagnosis and treatment, the incidence and mortality of HCC continue rising. And its pathogenesis is still unclear. As a highly conserved protein of the Golgi apparatus, Golgi phosphoprotein 3 (GOLPH3) has been shown to be involved in tumorigenesis of HCC. This study aimed to explore the exact oncogenic mechanism of GOLPH3 and provide a novel diagnose biomarker and therapeutic strategy for patients with HCC.MethodsFirstly, the expression of GOLPH3 was detected in the HCC tissue specimens and HCC cell lines. Secondly, RNA interference was used for GOLPH3 gene inhibition. Thirdly, cell proliferation was analyzed by MTT; cell apoptosis was analyzed by Annexin-V/PI staining, Hoechst 33,342 staining and caspase 3/7 activity assay. Fourthly, xenograft tumor model was used to study the function of GOLPH3 in tumor growth in vivo. Finally, western blotting and immunohistochemistry were used to investigate the role of GOLHP3 in the mTOR signaling pathway.ResultsData showed that the mRNA and protein expression of GOLPH3 were up-regulated in HCC tumor tissue and cell lines compared with those of control (P < 0.05). Correlation analyses showed that GOLPH3 expression was positively correlated with serum alpha-fetoprotein level (AFP, P = 0.006). Knockdown GOLPH3 expression inhibited proliferation and promoted apoptosis in HCC cell lines. What's more, knockdown GOLPH3 expression led to tumor growth restriction in xenograft tumor model. The expression of phosphorylated mTOR, AKT and S6 K1 were significantly higher in HCC tumor tissue and cell lines compared with those in normal liver tissues (p < 0.05). While the phosphorylated mTOR, AKT and S6 K1 were much lower when diminished GOLPH3 expression in HCC cell lines both in vitro and in vivo.ConclusionThe current study suggests that GOLPH3 contributes to the tumorigenesis of HCC by activating mTOR signaling pathway. GOLPH3 is a promising diagnose biomarker and therapeutic target for HCC. Our study may provide a scientific basis for developing effective approaches to treat the HCC patients with GOLPH3 overexpression.