Determination of Endogenous Radioresistance Biomarkers on HeLa Cells

Mamadou Soumboundou,Macoura Gadji,Marie Thérèse Aloy,Mounibe Diarra,Gora Mbaye,Ahmadou Dem,Celine Malesys,Claire Rodriguez-Lafrasse

doi:10.11648/j.ejb.20200802.18

Abstract

The Genes such as carbonic anhydrase IX (CAIX), hypoxia inducible factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) have been suggested as hypoxic biomarkers in cancer. Indeed, these endogenous biomarkers have been shown to have stronger prognostic value response after treatment by irradiation. However, inconsistent results suggest that factors other than oxygen influence their expression. This present study deciphers the level of expression of different radioresistance biomarkers in both normoxia and hypoxia conditions followed by irradiation of human ovarian tumor cell lines (uterine cervix squamous cell carcinoma (HeLa). HeLa cells were submitted to hypoxia (1% O2) conditions in a Thermo Scientific Heracell i CO2 incubator. The cells were subjected to two doses 4-10 Gy irradiation and re-incubate in their starting conditions for 4 hours, then fixed in 4% paraformaldehyde for 20 min. Protein expressions were assessed by immunocytochemistry staining and fluorescent images were captured by a Axio Imager Z1 fluorescence microscope with oil immersion lens at 63× magnification. In normoxia conditions there was no modification of the level of expression of the CAIX after irradiation. However, an increasing expression level of VEGF was noted. The level of expression of HIF-1 in normoxia was low compared to the other two proteins (CAIX and VEGF). Hypoxia conditions at 2% resulted in a low expression of CAIX and VEGF before and after irradiation at 10 Gy in HeLa cells. HIF-1 had a maximum expression level compared to CAIX and VEGF at 2% oxygen after irradiation in HeLa cells. As tumor hypoxia occurs in a deprived microenvironment, other environmental factors such as irradiation might interact with the effect of low oxygen concentration on gene expression. This study shows that irradiation of HeLa cells has a profound influence on the oxygen dependent induction of certain endogenous hypoxic markers as HIF-1, CAIX, and VEGF.

Highlights

IntroductionCervical cancer is the third most commonly diagnosed cancer and the fourth leading cause of cancer death in women worldwide [1]
Cervical cancer is the third most commonly diagnosed cancer and the fourth leading cause of cancer death in women worldwide [1].With over 528,000 new cases and more than 266,000 deaths in 2012 alone, cervical cancer is the fourth most common cancer in women worldwide, and second for women ages 15 to 44 [2]
Cells were fixed in 4% paraformaldehyde for 20 min, and immunodetection was performed as described by Hanot et al [17]

Summary

Introduction

Cervical cancer is the third most commonly diagnosed cancer and the fourth leading cause of cancer death in women worldwide [1]. With over 528,000 new cases and more than 266,000 deaths in 2012 alone, cervical cancer is the fourth most common cancer in women worldwide, and second for women ages 15 to 44 [2]. 740 deaths per day occur due to cervical cancer, making it the second most common cause of cancer death in women [2]. Radiotherapy (RT) is the first treatment option offered to a majority of patient cases, since most of them present in clinical stages IIB, III or IV [4]. Clinical prognostic factors include tumor stage and nodal involvement, metabolic response after treatment has been shown to have even stronger prognostic value [5]

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