Gene Expression Patterns of Colorectal Cancer Stem Cells  Following Ibuprofen and Hyperthermia Treatment

Behnaz  Valibeigi; Abbas  Behzad-Behbahani; Mohsen  Forouzanfar; Farzaneh  Zarghampoor; Mojtaba  Jaafarinia

doi:10.18502/ijph.v53i3.15155

Behnaz Valibeigi Department of Biology, Marvdasht Branch, Islamic Azad University, Marvdasht, Iran
Abbas Behzad-Behbahani Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sci-ences, Shiraz, Iran
Mohsen Forouzanfar Department of Biology, Marvdasht Branch, Islamic Azad University, Marvdasht, Iran
Farzaneh Zarghampoor Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sci-ences, Shiraz, Iran
Mojtaba Jaafarinia Department of Biology, Marvdasht Branch, Islamic Azad University, Marvdasht, Iran

DOI: https://doi.org/10.18502/ijph.v53i3.15155

Keywords: Colorectal cancer; Hyperthermia; Ibuprofen; Cancer stem cell

Abstract

Background: Cancer stem cells (CSCs) substantially influence the development of colorectal cancer (CRC), metastasis, relapse, and resistance to therapy. Ibuprofen and hyperthermia can be effective in the treatment of cancer. Herein, we evaluated the effects of hyperthermia and ibuprofen on the isolated-CSCs of CRC.

Methods: This experimental study was conducted between Sep 2020 and Jan 2022 at the Department of Pathology, School of Medicine, Shiraz University of Medical Sciences, Iran. A non-adhesive culture system was used to isolate CSCs from HT-29 cells. To confirm the stemness nature of isolated-CSCs, the expression of stemness genes and protein markers was evaluated by quantitative Real-time PCR (qRT-PCR) and flow cytometry assay. The isolated-CSCs were treated with hyperthermia and ibuprofen. The cell viability was determined by MTT assay and trypan blue staining. The expression of stemness, proliferation, Wnt signaling pathway and apoptosis genes was assessed by qRT-PCR.

Results: CSCs were isolated within 14 days. The expression of CD-133 marker and OCT3/4, C-MYC, KLF4, and NANOG genes in isolated-CSCs was higher than HT-29 cells (P<0.05). Cell viability of treated-CSCs were considerably reduced (P<0.05). Hperthermia reduced the expression of OCT3/4, NANOG, PCNA, WNT1 and CTNNB1 genes and increased the expression of P53, BAX, and KLF4 genes (P<0.05). Ibuprofen decreased the expression of OCT3/4, BCL2, NANOG, PCNA, WNT1, and CTNNB1 genes and increased the expression of P53, BAX, and KLF4 genes in treated-CSCs (P<0.05).

Conclusion: Hyperthermia and ibuprofen treatment demonstrate an inhibitory effect on colorectal CSCs. However, using combination therapy is remaining to be tested.