Oxidative Stress and X-ray Exposure Levels-Dependent Survival and Metabolic Changes in Murine HSPCs.

Melis Karabulutoglu,Rosemary Finnon,Mark A Hill,Lourdes Cruz-Garcia,Christophe Badie

doi:10.3390/antiox11010011

Abstract

Haematopoietic bone marrow cells are amongst the most sensitive to ionizing radiation (IR), initially resulting in cell death or genotoxicity that may later lead to leukaemia development, most frequently Acute Myeloid Leukaemia (AML). The target cells for radiation-induced Acute Myeloid Leukaemia (rAML) are believed to lie in the haematopoietic stem and progenitor cell (HSPC) compartment. Using the inbred strain CBA/Ca as a murine model of rAML, progress has been made in understanding the underlying mechanisms, characterisation of target cell population and responses to IR. Complex regulatory systems maintain haematopoietic homeostasis which may act to modulate the risk of rAML. However, little is currently known about the role of metabolic factors and diet in these regulatory systems and modification of the risk of AML development. This study characterises cellular proliferative and clonogenic potential as well as metabolic changes within murine HSPCs under oxidative stress and X-ray exposure. Ambient oxygen (normoxia; 20.8% O2) levels were found to increase irradiated HSPC-stress, stimulating proliferative activity compared to low oxygen (3% O2) levels. IR exposure has a negative influence on the proliferative capability of HSPCs in a dose-dependent manner (0–2 Gy) and this is more pronounced under a normoxic state. One Gy x-irradiated HSPCs cultured under normoxic conditions displayed a significant increase in oxygen consumption compared to those cultured under low O2 conditions and to unirradiated HSPCs. Furthermore, mitochondrial analyses revealed a significant increase in mitochondrial DNA (mtDNA) content, mitochondrial mass and membrane potential in a dose-dependent manner under normoxic conditions. Our results demonstrate that both IR and normoxia act as stressors for HSPCs, leading to significant metabolic deregulation and mitochondrial dysfunctionality which may affect long term risks such as leukaemia.

Highlights

O2 environment of the bone marrow (BM) endosteal stem cell niche supports the maintenance of long-term hematopoietic stem cells (LT-haematopoietic stem cell (HSC)) in a protective quiescent state, whereas the actively differentiating/proliferating short-term HSCs (ST-HSCs) and progenitor cells are in the higher O2 environment of the perivascular niches [3,5,6,7,8,9]
In this study we describe using the Seahorse XFp Analyzer to assess the effect of ionizing radiation (IR) exposure on mitochondrial function and O2 metabolism in primary haematopoietic stem and progenitor cell (HSPC) cultures obtained from a radiation-induced Acute Myeloid Leukaemia (rAML)-sensitive CBA/Ca mouse model under normoxic (20.8%) and low (3%) O2 conditions
In this paper we have presented an analysis of the changes in cellular O2 metabolism, mitochondrial activity and function and proliferation potential induced by exposure to ionising radiation in HSPC obtained from CBA/Ca rAML model mice in short-term primary cell culture

Summary

Introduction

Haematopoiesis relies on the fine regulation of bone marrow (BM) haematopoietic stem cell (HSC) biology in response to continuously changing internal and external cues from the niche microenvironment [1,2]. O2 availability has been stated in multiple stem cell contexts and has been extensively studied in the haematopoietic stem and progenitor cell (HSPC) population [3,4]. O2 environment of the BM endosteal stem cell niche supports the maintenance of long-term hematopoietic stem cells (LT-HSCs) in a protective quiescent state, whereas the actively differentiating/proliferating short-term HSCs (ST-HSCs) and progenitor cells are in the higher O2 environment of the perivascular niches [3,5,6,7,8,9].

Methods

Discussion

Conclusion