The role of the uncoupling protein2 -866G/A polymorphism in oxidative stress markers associated with air pollution exposure during pregnancy.
Consistently high levels of air pollutants such as sulphur dioxide, particle matter and nitric oxides have been observed in the Durban South (DS) industrial basin. The adverse health outcomes associated with ambient air pollution (AAP) exposure have underlying molecular mechanisms. Oxidative stress is a known outcome of AAP exposure and contributes to the exacerbation of adverse AAP related outcomes such as chronic obstructive pulmonary disorder (COPD) and asthma. Pregnant women are at increased risk of developing oxidative stress due to increased energy expenditure. Oxidative stress during pregnancy is linked to adverse birth outcomes such as intrauterine growth retardation and low birth weight. The mitochondria are the most abundant source of endogenous reactive oxygen species (ROS), making these organelles extremely susceptible to oxidative damage. Alterations in mitochondrial function by air pollutants can contribute to oxidative stress. Uncoupling protein2 (UCP2) is an anion carrier located on the inner mitochondrial membrane that regulates mitochondrial ROS production by reducing mitochondrial membrane potential (Δψm) through mild uncoupling. Genetic variation in genes that play a role in oxidative stress response is likely to influence susceptibility to oxidative stress related health outcomes. The aim of this study was to evaluate air pollution associated oxidative stress response in women from the DS industrial basin and determine the functional relevance of a common -866G/A promoter polymorphism in the UCP2 gene. Fifty pregnant women from DS and 50 from north Durban (DN; control) were recruited. The thiobarbituric acid assay (TBARS) and comet assay were performed to measure oxidative stress and DNA fragmentation. Mitochondrial function was evaluated by JC-1 Mitoscreen and ATP luminometry. Quantitative PCR (qPCR) was performed to measure mitochondrial DNA (mtDNA) damage. Antioxidant response was determined by qPCR to measure mRNA expression of superoxide dismutase 2 (SOD2), nuclear factor erythroid 2-related factor 2 (Nrf2) and UCP2 mRNA expression. Western blots were performed to quantify UCP2 and Nrf2 protein expression. The samples were genotyped using PCR - restriction fragment length polymorphism. Results from the TBARS assay showed women from DS displayed elevated levels of MDA, a marker for oxidative stress (0.07±0.06μM; p = 0.56). ATP (1.89 fold) and Δψm (45.3±17.2%; p = 0.8) were also elevated in women from DS, favouring free radical production. DNA fragmentation, as indicated by comet tail length was also higher in DS when compared to the control group (0.57±0.16μm; p = 0.037). Analysis of mtDNA viability showed a 0.49 fold change in mtDNA amplification in women from the industrialized DS. All antioxidant genes, i.e. Nrf2 (0.73 fold), UCP2 (1.58 fold), SOD2 (1.23 fold), were up regulated in women from DS. Analysis of protein expression showed a significant increase in UCP2 expression (0.08±0.03RBI; p = 0.049) and a significant decline in Nrf2 levels (1.68±0.84RBI; p = 0.03). The homozygous G genotype was significantly more frequent in DS (37.5%) than in DN (18.6%; p = 0.047; OR: 2.57; 95% CI: 1.353 to 4.885). This genotype exhibited higher MDA levels, comet tail length, Δψm, SOD2, Nrf2, and UCP2 expression than the AA/GA in genotype in women from DS (p > 0.05). This study found that pregnant women from a more industrialized area exhibit higher markers for oxidative stress and conditions that favour mitochondrial free radical production.