Orexin A induces autophagy in HCT-116 human colon cancer cells through the ERK signaling pathway

Wen,Jing; Zhao,Yuyan; Guo,Lei

doi:10.3892/ijmm.2015.2409

Orexin A induces autophagy in HCT-116 human colon cancer cells through the ERK signaling pathway

Authors:
- Jing Wen
- Yuyan Zhao
- Lei Guo
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Affiliations: Department of Endocrinology, First Affiliated Hospital, China Medical University, Shenyang, Liaoning 110001, P.R. China, Department of Orthopedic Surgery, First Affiliated Hospital, China Medical University, Shenyang, Liaoning 110001, P.R. China
Published online on: November 13, 2015 https://doi.org/10.3892/ijmm.2015.2409
Pages: 126-132

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Abstract

Orexins are a class of peptides which have a potent influence on a broad variety of cancer cells. Autophagy is closely associated with tumors; however, its function is not yet completely understood. In this study, we aimed to determine whether orexin A induces autophagy in HCT‑116 human colon cancer cells and to elucidate the molecular mechanisms involved. For this purpose, HCT‑116 cells were treated with orexin A, and cell viability was then measured by MTT assay, and apoptosis was determined by flow cytometry. The expression levels of autophagy‑related proteins were measured by western blot analysis. Quantitative analysis of autophagy following acridine orange (AO) staining was performed using fluorescence microscopy, and cellular morphology was observed under a transmission electron microscope. In addition, the HCT‑116 cells were treated with the extracellular signal‑regulated kinase (ERK) inhibitor, U0126, or the autophagy inhibitor, chloroquine, in combination with orexin A in order to examine the activation of ERK. We found that orexin A significantly inhibited the viability of the HCT‑116 cells. Both autophagy and apoptosis were activated during the orexin A‑induced death of HCT‑116 cells. When the HCT‑116 cells were treated with orexin A for 24 h, an accumulation of punctate microtubule-associated protein-1 light chain 3 (LC3) and an increase in LC3‑Ⅱ protein levels were also detected, indicating the activation of autophagy. Moreover, orexin A upregulated ERK phosphorylation; however, U0126 or chloroquine abrogated ERK phosphorylation and decreased autophagy, compared to treatment with orexin A alone. Therefore, our findings demonstratedm that orexin A induced autophagy through the ERK pathway in HCT‑116 human colon cancer cells. The inhibition of autophagy may thus prove to be an effective strategy for enhancing the antitumor potential of orexin A as a treatment for colon cancer.

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