Oxygen and differentiation status modulate the effect of X-ray irradiation on physiology and mitochondrial proteome of human neuroblastoma cells

Abstract

Cytotoxic effects, including oxidative stress, of low linear energy transfer (LET)-ionizing radiation are often underestimated and studies of their mechanisms using cell culture models are widely conducted with cells cultivated at atmospheric oxygen that does not match its physiological levels in body tissues. Also, cell differentiation status plays a role in the outcome of experiments. We compared effects of 2 Gy X-ray irradiation on the physiology and mitochondrial proteome of nondifferentiated and human neuroblastoma (SH-SY5Y) cells treated with retinoic acid cultivated at 21% and 5% O. Irradiation did not affect the amount of subunits of OxPhos complexes and other non-OxPhos mitochondrial proteins, except for heat shock protein 70, which was increased depending on oxygen level and differentiation status. These two factors were proven to modulate mitochondrial membrane potential and the bioenergetic status of cells. We suggest, moreover, that oxygen plays a role in the differentiation of human SH-SY5Y cells.

I-Shaped Dimers of a Plant Chloroplast FOF1-ATP Synthase in Response to Changes in Ionic Strength

“…BN-PAGE was performed as described [ 55 , 56 ]. Briefly, the samples of cF O F 1 were analyzed in linear gradient polyacrylamide gel (from 4 to 14% ( w / v ) BN-PAGE, the separation gel was overlayed with a 3% ( w / v ) sample gel).…”

Oxygen Concentration and Oxidative Stress Modulate the Influence of Alzheimer’s Disease Aβ1–42 Peptide on Human Cells

“…Human neuroblastoma (SH-SY5Y) cells were grown in DMEM (Gibco, Life Technologies, Paisley, UK) supplemented with 10% FBS (PAA Laboratories GmbH, Pasching, Austria), 2% L-glutamine, and 5 U/ml penicillin/5 μ g/ml streptomycin (Gibco, Life Technologies, Paisley, UK) at 37°C, 5% CO 2 , at two different oxygen conditions (standard 21% O 2 and 5% O 2 , resp.) as described previously [ 23 ]. Simple home-made incubators for cultivation of cells at 5% O 2 were set up according to the protocol by Wright and Shay [ 27 ].…”

“…Total cellular ATP concentration of SH-SY5Y cells was determined using a luminescent ATP detection assay (ab113849, Abcam, Cambridge, UK) according to the manufacturer's protocol and as described previously [ 23 ].…”

“…On the list of other possible causative agents and factors for the development of AD is ionizing radiation (IR), particularly dental X-rays and related IR capable of destroying dividing microglial cells that support neurons [ 21 ], by damaging microglia telomeres causing premature death as proposed by Rodgers [ 22 ]. Furthermore, mitochondria are very important targets of ionizing radiation [ 23 ] and their direct damage leads to further nuclear DNA damage [ 24 ]. Accumulation of a common deletion in mtDNA (Δ-mtDNA 4977 ) occurs after mitochondrial degeneration in diseases and aging and is induced by ionizing radiation as well [ 25 , 26 ].…”

“…Since oxygen in the cell culture modulates cellular response to stress [ 23 ], we studied effects of ionizing radiation or A β 1–42 peptide disaggregated to monomers and oligomers on human cells pretreated with retinoic acid for induction of differentiation at two different oxygen concentrations (21% and 5% O 2 ). For the first time, to our knowledge, the combined effects of the A β peptide and ionizing radiation on cellular parameters and survival were investigated.…”