Gastroenterology

Gastroenterology

Volume 149, Issue 4, October 2015, Pages 1006-1016.e9
Gastroenterology

Original Research
Full Report: Basic and Translational—Alimentary Tract
FOXM1-Induced PRX3 Regulates Stemness and Survival of Colon Cancer Cells via Maintenance of Mitochondrial Function

https://doi.org/10.1053/j.gastro.2015.06.007Get rights and content

Background & Aims

Reagents designed to target cancer stem cells (CSCs) could reduce tumor growth, recurrence, and metastasis. We investigated the mitochondrial features of CSCs.

Methods

Colon adenocarcinoma fragments were obtained from 8 patients during surgery at Busan Paik Hospital in Korea. We used immunohistochemistry and quantitative polymerase chain reaction to compare expression of mitochondrial peroxiredoxin 3 (PRX3) in CD133+CD44+ Lgr5+cells (CSCs) vs CD133CD44Lgr5 colon tumor cells (non-CSCs). Cell survival and expression of mitochondrial-related genes were analyzed in the presence of 5-fluorouracil and/or antimycin A. We used small-interfering and short-hairpin RNAs and an overexpression vector to study PRX3, which functions in the mitochondria. CD133+ cells with PRX3 knockdown or overexpressing PRX3 were grown as xenograft tumors in immunocompromised mice. Metastasis was studied after injection of tumor cells in spleens of mice. We used chromatin immunoprecipitation and reporter assays to characterize transcriptional regulation of PRX3 by forkhead box protein 1.

Results

CSCs had a higher mitochondrial membrane potential and increased levels of adenosine triphosphate, Ca2+, reactive oxygen species, and oxygen consumption than non-CSCs. Levels of PRX3 were increased in colon CSCs compared with non-CSCs. PRX3 knockdown reduced the viability of CSCs, but non non-CSCs, by inducing mitochondrial dysfunction. PRX3 knockdown reduced growth of CSCs as xenograft tumors or metastases in mice. The expression of FOXM1 activated transcription of PRX3 and expression of CD133 in colon CSCs.

Conclusions

Human colon CSCs have increased mitochondrial function compared with colon tumor cells without stem cell properties. Colon CSCs overexpress the mitochondrial gene PRX3, which is required for maintenance of mitochondrial function and tumorigenesis, and is regulated by forkhead box protein 1, which also regulates expression of CD133 in these cells. These proteins might be therapeutic targets for colorectal cancer.

Section snippets

Materials and Methods

Detailed Materials and Methods are available in the Supplementary Material.

Colon Cancer Stem Cells Utilize Primary Mitochondrial Oxidative Phosphorylation Rather Than Glycolysis to Produce Adenosine Triphosphate

We assessed potential differences in mitochondrial function between colon CSCs and non-CSCs using antibody-based detection of CD133, CD44, and Lgr5 (Figure 1). The CD133+CD44+Lgr5+ cells (CSCs) had a higher mitochondrial membrane potential (ΔΨm), and increased levels of Ca2+ and ROS compared with CD133CD44Lgr5- cells (non-CSCs) (Figure 1A−C). In addition, the oxygen consumption rates and mitochondrial ATP levels of CSCs were much higher than non-CSCs, and lactate production was higher in

Discussion

In the current study, we found that highly expressed mitochondrial Prx3 plays a critical role in maintaining mitochondrial function via elimination of ROS produced by ATP production in the OXPHOS system in CSCs. This role of Prx3 is important for promoting stemness and survival of CSCs. We further showed that Prx3 and CD133 expression is up-regulated by the transcription factor FoxM1 in colon CSCs. We also showed that the mitochondrial function of colon CSCs is improved in comparison with that

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    Conflicts of interest The authors disclose no conflicts.

    Funding This study was supported by a grant from a Priority Research Centers Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science, and Technology (2010-0020224), and a Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science, and Technology (2012R1A1A2041700 and 2012R1A2A1A03007595).

    Author names in bold designate shared co-first authorship.

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