Browsing by Author "Nagiah, Savania."

Now showing 1 - 13 of 13

A biochemical assessment of stress response following acute and prolonged exposure to antiretroviral drugs (nucleoside reverse transcriptase inhibitors) in vitro.
(2015) Nagiah, Savania.; Chuturgoon, Anil Amichund.; Phulukdaree, Alisa.
Nucleoside reverse transcriptase inhibitors (NRTIs) are the most extensively used antiretroviral (ARV) drugs in highly active antiretroviral therapy (HAART). The long term use of HAART is associated with changes to metabolic parameters leading to lipodystrophy and metabolic syndrome, as well as toxicity to high energy demand organs e.g. liver, kidney, heart, and nervous system. Underlying the myriad of NRTI-associated adverse health outcomes is mitochondrial (mt) toxicity. Although inhibition of mtDNA synthesis was one of the first identified mechanisms of toxicity, it did not provide a holistic explanation for all NRTIs. Furthermore, variations in adaptive stress responses were observed following acute and chronic exposure to NRTIs. Insight gained from the molecular changes induced by NRTIs will enable effective management and limit adverse health outcomes. The human hepatoma (HepG2) cell line was used as an in vitro model to investigate changes to mt function, cellular redox status, and antioxidant response following acute [24 hour (h)] and prolonged (120 h) exposure to NRTIs – Zidovudine (AZT, 7.1μM); Stavudine (d4T, 4μM); Tenofovir (TFV, 1.2μM). Long term exposure to AZT and d4T reduced mtDNA levels (120h, AZT: 76.1%; d4T:36.1%, p<0.05) and mt function was compromised as evidenced by reduced ATP levels (AZT: 38%; d4T: 56.4%) and increased mt membrane depolarisation (p<0.02). Tenofovir compromised mt function at 120 h independently of depleting mtDNA levels. Oxidative stress parameters were significantly elevated by AZT and TFV at 24h; and all NRTIs at 120 h (p<0.05). Endogenous antioxidant response was highest in TFV in both time periods (120h; p<0.05). Once NRTI induced oxidative stress in HepG2 cells was established, protein homeostatic response to oxidative stress was investigated. Lon protease expression and related endoplasmic reticulum (ER) stress was evaluated. The data showed that ATP-dependent protein homeostasis responses Lon, heat shock protein 60 (HSP60) and ER stress were significantly increased at 24 h (>2 fold); but significantly decreased at 120 h for all NRTIs (p<0.005). The compromised ATP-dependent stress response then led to the assessment of an ATP- dependent drug transporter responsible for efflux of xenobiotics in hepatocytes. The transporter, ATP-binding cassette C4 (ABCC4), is regulated by microRNA (miR-) 124a. Regulation of ABCC4 by miR-124a has implications for bio-accumulation and resultant toxicity. An inverse relationship between miR-124a and ABCC4 mRNA levels in all treatments at both time periods was observed. All NRTIs elevated miR-124a levels at 24 h (p=0.0009) and this observation was consistent in d4T and TFV treated HepG2 cells at 120 h (p<0.0001). This was accompanied with a concomitant decline in ABCC4 mRNA levels (p<0.0001) relative to the control. Prolonged exposure to AZT caused a decrease in miR-124a and elevated ABCC4 mRNA levels. Protein expression of multi-drug resistance protein 4 (MRP4), coded for by ABCC4, did not correlate to mRNA expression. At 120 h, all NRTIs caused significant depletion of MRP4 (possibly due to oxidative cell membrane damage or ATP depletion). In conclusion, all three NRTIs compromised mt function and induced oxidative damage in HepG2 cells, with greater toxicity over 120 h. Reduced ATP levels compromised the ATP-dependent stress response proteins and xenobiotic detoxification. Tenofovir could be considered a safer alternative as it elicited the highest antioxidant response in spite of reduced mt function.
Deoxynivalenol downregulates NRF2-induced cytoprotective response in human hepatocellular carcinoma (HepG2) cells.
(2017) Ndlovu, Siqiniseko Sinikiwe.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
Deoxynivalenol (DON) is a mycotoxin produced by Fusarium species that commonly infect agricultural foods. DON exhibits multiple toxic effects in both animals and humans, binding to the A site of the 28S ribosome and inhibits peptidyl transferase and protein elongation. It induces cytotoxicity through oxidative stress and inhibition of protein synthesis. Liver cells possess the antioxidant signalling mediator - Nuclear erythroid-2-Related factor (NRF2) that is activated in response to oxidative stress. There is no sufficient work done to show if the HepG2cells have an ability to withstand the molecular modifications induced by DON. The aim of the study was to investigate the cytotoxicity of DON and its effect on the NRF2 antioxidant response in HepG2 cells. The MTT assay was used to determine a dose response of DON (72 hr) on cell viability and to generate an IC50 value to use in subsequent assays. The intracellular concentration of GSH and ATP was determined using Luminometry. Lipid peroxidation and membrane damage were assessed by TBARS and LDH cytotoxicity assays respectively. Protein expression of NRF2, phosphorylated (p-)NRF2, catalase (CAT), superoxide dismutase (SOD)2, and Sirtuin (Sirt)3 was quantified by Western Blotting. The mRNA expressions of GPx, CAT and SOD2 were quantified using qPCR. DON decreased cell viability in a dose-dependent manner with an IC50 value of 26.17 μM. DON caused a significant decrease in the intracellular GSH concentration (1.77-fold, p= 0.0005). There was a significant decrease in the intracellular ATP content (1.92-fold, p= 0.0002).The study shows an induced lipid peroxidation and membrane damage in HepG2 cells by DON, as there was a significant increase in extracellular levels of both MDA (1.89-fold, p=0.0020) and LDH (1.35-fold, p=0.0207). DON reduced total NRF2 expression (0.30-fold, p= 0.0017), however activated p-NRF2 was significantly up-regulated (3.54-fold, p= 0.0085). There was a downregulation in the NRF2 target antioxidant proteins: CAT (0.33-fold, p= 0.005) with a concomitant decrease in CAT mRNA levels (0.02-fold, p= 0.0003), SOD2 (0.02-fold, p= 0.0137), with a parallel trend in the levels of SOD2 mRNA (0.06-fold, p= 0.0020) by DON. This toxin also significantly decreased the mRNA expression of GPx levels (0.03-fold, p= 0.0006). The expression of a mitochondrial stress response Sirt3 was significantly decreased (0.14-fold, p= 0.0058). Taken together, the data shows that DON causes oxidative stress and downregulates the NRF2-induced cytoprotection in HepG2 cells. Keywords: Deoxynivalenol Antioxidant response NRF-2
Fumonisin B1 induced antioxidant response in C57BL/6 male mice brain.
(2018) Sibiya, Thabani.; Chuturgoon, Anil Amichund.; Nagiah, Savania.; Ghazi, Terisha.
Background: Fumonisin B1 (FB1), a mycotoxin produced by the Fusarium species, contaminates maize. In South Africa maize is a dietary staple and FB1 endangers human and animal health. FB1 is known to have neurodegenerative effects; inhibits mitochondrial respiration, causes mitochondrial membrane depolarization and excessive ROS production. This study investigated the antioxidant response in mice brain after acute (24 hrs) and prolonged (10 days) exposure to FB1. Methods: Four groups (Control acute, FB1 acute, Control prolonged, FB1 prolonged) of C57BL/6 male mice (n=5 per group) were used. All controls were orally administered 0.1M PBS and FB1 groups were administered 5mg/kg of FB1. Following acute and prolonged exposure, the mice were euthanised by halothane anaesthesia. Brain tissues were harvested and stored in Qiazol and Cytobuster for RNA and protein isolation, respectively. Protein expression of CAT, pNrf2 and Nrf2 were determined using western blots. The mRNA expression of Nrf2, miR-141, SOD2, GPx, Tfam, LON, SIRT3 and Tau wwere determined using qPCR. Results: Protein expression of Nrf2 (Acute: *p=0,0144; prolonged: **p=0,0094) and pNrf2 (acute: *p=0,0132; prolonged: *p=0,0462) was significantly increased upon 24 hrs and significantly decreased upon 10 days in tissue exposed to FB1, while mRNA levels of Nrf2 were significantly reduced upon acute (***p=0,0001) and prolonged (**p=0,0013) exposure. FB1 induced a significant decrease in miR-141 levels in tissue following acute (**p=0,0019) and prolonged (***p=0,0004) exposure. FB1 increased the protein expression of CAT in tissue following acute (p=0,1206) and significantly increased expression upon prolonged (**p=0,0010) exposure. FB1 also significantly increased the mRNA expression of GPx in acute (***p=0,0001) and prolonged (**p=0,0024) exposure. FB1 significantly decreased the expression of SOD2 in mice brain following acute (**p=0,0070) and non-significantly decrease upon prolonged (p=0,2725) exposure. Tfam and LONP1 levels were significantly decreased upon acute (***p=0,0003, ***p=0,0005) and prolonged (*p=0,0196, *p=0,0117) exposure to FB1 respectively. However, SIRT3 expression was decreased upon acute (p=0,0594) and significantly increased upon prolonged (*p=0,0283) exposure to FB1.The mRNA expression of tau was significantly reduced upon acute (**p=0,0054) and prolonged (*p=0,0273) exposure to FB1. Conclusion: FB1 compromises antioxidant and mitochondrial survival responses in mice brain. This may have implications in FB1-induced neurodegeneration.
Fumonisin B1-induced oxidative stress in human liver (HepG2) cells – an alternate mechanism of carcinogenesis.
(2017) Arumugam, Thilona.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
Abstract available in pdf.
Fusaric acid dampens the Nrf2-mediated stress response in human embryonic kidney cells.
(2016) Govender, Melissa.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
Abstract available in PDF file.
Fusaric acid induces mitochondrial stress in human hepatocellular carcinoma (HepG2) cells.
(2015) Abdul, Naeem Sheik.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
Abstract available in PDF file.
Fusaric acid induces oxidative stress and apoptosis in human oesophageal cancer cells.
(2016) Devnarain, Nikita.; Chuturgoon, Anil Amichund.; Tiloke, Charlette.; Nagiah, Savania.
Abstract available in PDF file.
Fusaric acid-induced epigenetic modifications in vitro and in vivo: alternative mechanisms of hepatotoxicity.
(2019) Ghazi, Terisha.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
The Fusarium-produced mycotoxin, Fusaric acid (FA), is a frequent contaminant of agricultural foods that exhibits toxicity in plants and animals with little information on its molecular and epigenetic mechanisms. Epigenetic modifications including DNA methylation, histone methylation, N-6-methyladenosine (m6A) RNA methylation, and microRNAs are central mediators of cellular function and may constitute novel mechanisms of FA toxicity. This study aimed to determine epigenetic mechanisms of FA-induced hepatotoxicity in vitro and in vivo by specifically investigating DNA methylation, histone 3 lysine (K) 9 trimethylation (H3K9me3), and m6A-mediated regulation of p53 expression in human liver (HepG2) cells and C57BL/6 mice livers. FA induced global DNA hypomethylation in HepG2 cells; decreased the expression of DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B) by inducing promoter hypermethylation and upregulated expression of miR-29b. Further, FA decreased the ubiquitination of DNMT1, DNMT3A, and DNMT3B by decreasing the expression of the ubiquitination regulators, UHRF1 and USP7. FA induced promoter hypomethylation of the demethylase, MBD2 and increased MBD2 expression contributing to global DNA hypomethylation in HepG2 cells. DNA methylation and H3K9me3 function in concert to regulate genome integrity and gene transcription. Sirtuin (Sirt) 1 is a histone deacetylase and direct target of miR-200a that regulates the repressive H3K9me3 mark by post-translationally modifying both H3K9Ac and the histone methyltransferase, SUV39H1. FA upregulated miR-200a and decreased Sirt1 expression in HepG2 cells and C57BL/6 mice livers. FA decreased the expression of SUV39H1 and histone demethylase, KDM4B which led to a decrease in H3K9me3 and an increase in H3K9me1. FA also decreased cell viability via apoptosis as evidenced by the significant increase in the activity of the executioner caspase-3/7. The tumor suppressor protein, p53 regulates cell cycle arrest and apoptosis in response to cellular stress. The expression of p53 is regulated at the transcriptional and post-transcriptional level by promoter methylation and m6A RNA methylation. In HepG2 cells, FA induced p53 promoter hypermethylation and decreased p53 expression. FA also decreased m6A-p53 levels by decreasing the expression of the methyltransferases, METTL3 and METTL14, and the m6A readers, YTHDF1, YTHDF3, and YTHDC2, thereby, decreasing p53 translation. In C57BL/6 mice livers FA, however, induced p53 promoter hypomethylation and increased p53 expression. FA increased m6A-p53 levels by increasing the expression of METTL3 and METTL14; and increased expression of YTHDF1, YTHDF3, and YTHDC2 increased p53 translation. In conclusion, this study provides evidence for alternative mechanisms of FA-induced hepatotoxicity (in vitro and in vivo) by modulating DNA methylation, H3K9me3, m6A RNA methylation, and epigenetically regulating p53 expression ultimately leading to genome instability and apoptotic cell death. These results provide insight into a better understanding of FA induced hepatic toxicity at the epigenetic and cellular level and may assist in the development of preventative and therapeutic measures against FA toxicity. It also suggests that exposure to FA may lead to the onset of human diseases via epigenetic changes/modifications. This is particularly relevant in under privileged communities where the food supply and storage conditions are inadequate.
HMG-CoA reductase inhibitor, atorvastatin induces apoptasis in human lung adenocarcinoma cell (A549).
(2015) Ramharack, Pritika.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
Abstract available in PDF file.
An investigation into the mitochondrial toxicity of fusaric acid associated with aberrant energy metabolism and inflammatory responses.
(2019) Sheik-Abdul, Naeem.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
No abstract available.
An investigation of the anti-hyperglycaemic, biochemical and molecular effects of 4-hydroxyisoleucine and fenugreek seed extract in comparison to metformin in vitro and in vivo.
(2017) Naicker, Nikita.; Chuturgoon, Anil Amichund.; Nagiah, Savania.
Type two diabetes mellitus (T2D) is a significant cause of premature death and disability, accompanied with negative socio-economic impacts. This metabolic disorder is characterized by hyperglycaemia and defective insulin signalling. Long-term exposure to hyperglycaemia gives rise to altered fat metabolism and reactive oxygen species (ROS) generation. These precursors are central to the progression of dyslipidaemia and attenuated antioxidant (AO) response and detoxification system, respectively. Diabetic dyslipidaemia and oxidative stress (OS) are risk factors for the onset and progression of cardiovascular disease (CVD) and other diabetic complications. The treatment regimen for T2D comprises self-care and anti-diabetic drugs such as metformin. However, due to the lack of compliance to self-care recommendations and some undesirable side effects of metformin, there is the necessity for alternate therapy. Natural products have been used for the treatment of many disorders, including T2D. Trigonella foenum-graecum commonly known as fenugreek is a plant that possesses anti-diabetic effects. These effects are attributed to its bioactive compound – 4-hydroxyisoleucine (4-OH-lle), which constitutes approximately 80% of the bio-composition of the fenugreek seed. Despite these effects, biochemical and molecular effects of 4-OH-lle on insulin signalling, lipid metabolism, and ROS production is not well-documented. This study investigated the effects of 4-OH-lle in comparison to metformin and fenugreek seed extract (FSE) on hyperglycaemic human hepatoma (HepG2) cells and C57BL/6 male mice. Treatments were conducted under normoglycaemic and hyperglycaemic conditions as follows; control, 4-OH-lle (in vitro: 100ng/ml; in vivo: 100mg/kg Body weight) metformin (in vitro: 20mM; in vivo: 20mg/kg Body weight) and FSE (in vitro: 100ng/ml; in vivo: 100mg/kg Body weight) treatment groups. The experiments included; blood glucose measurements, lipid profile analysis, spectrophotometric assays (in vitro), western blotting for protein expression and qPCR for mRNA expression. First, to validate the effects on insulin signalling and glucose sensing, glucose levels were measured with completion of an oral glucose tolerance test. 4-OH-lle treatment attenuated glucose levels, and elevated the mRNA levels of glycogen synthase (GS) and glucokinase (Gck). This was followed by the investigation of the protein and gene expression of insulin signalling regulators: insulin receptor β (IRβ), insulin receptor substrate 1 (IRS1), phosphorylated protein kinase B (pAkt), phosphorylated glycogen synthase kinase 3α/β (pGSK3α/β) and glucose transport 2 (GLUT2). In in vivo hyperglycaemia, 4-OH-lle increased the expression of the investigated proteins and genes. The results showed that 4-OH-lle was just as potent as MF, and FSE in stimulating the insulin signalling cascade. Second, the effect of 4-OH-lle on dyslipidaemia was investigated by measuring mRNA levels of sterol regulatory binding element 1c (SREBP1c) and fatty acid synthase (FAS) – key factors in fatty acid metabolism. Both genes were up-regulated and correlated with the changes in triglyceride and cholesterol levels. Next the protein expression of proprotein convertase subtilisin-like/kexin type (PCSK9) - a regulator of low density lipoprotein cholesterol (LDLc) and peroxisome proliferator-activated receptor gamma (PPARG) – a regulator of high density lipoprotein (HDLc) was evaluated. The data showed that 4-OH-lle down-regulated protein and mRNA expression of PCSK9 and up-regulated protein expression of PPARG. The reduction in PCSK9 levels correlated with the changes observed in low density lipoprotein receptor (LDLr) and LDLc, whereas the increase in PPARG correlated with the elevated mRNA expression of apolipoprotein A1 (Apo A1) and HDLc. Together these results provide substantial evidence for the regulatory effect of 4-OH-lle, in comparison to metformin, and FSE on PCSK9, PPARG and related lipid factors. Finally, the effect of 4-OH-lle on redox status and AO response was assessed by measuring nuclear factor E2-related factor 2 (Nrf2). In both models, there was an increase in the protein expression of phosphorylated Nrf2 accompanied by an increase in mRNA levels of superoxide dismutase 2 (SOD2) and glutathione peroxidase (GPx), and GSH levels. Mitochondria play a central role in contributing to elevated ROS levels. While nuclear responses like Nrf2 regulate ROS, mitochondria possess their own maintenance proteins. These include mitochondrial Lon protease 1 (LonP1), Sirtuin 3 (SIRT3) and peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) which play an integral role in combatting OS and mitochondrial dysfunction. The results showed that 4-OH-lle displayed a potent effect in inducing the AO response and increasing mitochondrial regulatory proteins. In conclusion, 4-OH-lle improved the compromised insulin signalling and the altered lipid profile as well as induced the AO response and mitochondrial maintenance proteins, in the presence of elevated glucose. Furthermore, the effect of 4-OH-lle was greater than the first-line drug; metformin and FSE, albeit in cultured human liver cells and a mouse model. Also, the crude seed extract displayed promising effects on all investigated parameters. Considering the active role of chronic hyperglycaemia in the onset and progression of CVD and diabetic complications, 4-OH-lle poses as a highly favourable alternate therapy in the treatment of T2D. Moreover, this has great importance in socio-economically challenged communities where T2D is a common disorder, access to healthcare facilities is limited, and plants serve as sources of easily accessible treatments.
Involvement of MAPK and Akt in the immunotoxicity of fusaric acid on healthy peripheral blood mononuclear cells (PBMC) and acute monocytic leukemic (Thp-1) cells.
(2015) Dhani, Shanel.; Chuturgoon, Anil Amichund.; Nagiah, Savania.; Naidoo, D. B.
Fusaric acid is a divalent chelator with moderate toxicity in plant and animals. However, studies lack on its effect on human models and the immune system. This study investigated the immunotoxicity of FA on PBMCs and Thp-1 cells. Cell viability was determined using the WST-1 assay and the mode of cell death by flow cytometry using the annexin V-FITC stain. Caspase 8, 9 and 3/7 activities were determined using Caspase-Glo assay®. TNF-α levels were measured using the TNF-α ELISA kit. Oxidative damage (MDA) was determined using the TBARS assay. Flow cytometry was performed to determine mitochondrial function using the JC-1 stain. ATP levels were measured using the ATP CellTitre Glo reagent. Western blotting was performed to determine protein expressions of Bax, p- Bcl-2, p-Akt, p-ERK, p-JNK and p38. The immunotoxicity of FA was confirmed by the decreased cell viability of PBMCs and Thp-1 cells and was validated by the externalization of phosphatidylserine on both PBMCs (p<0.005) and Thp-1 cells (p<0.0001). In PBMCs, FA induced paraptosis, evidenced by the decreased caspase 8 (p<0.005), 9 (p<0.05) and 3/7 (p<0.005) activities. Whilst in Thp-1 cells, FA induced intrinsic apoptosis supported by a decrease in caspase 8 activity (p<0.05) and an increase in caspase 9 (p<0.05) and 3/7 (p<0.005) activities; corresponding with unchanged TNF-α levels in both PBMCs (p=0.3015) and Thp-1 cells (p=0.4540). In PBMCs, FA significantly decreased Bax (pro-apoptotic) protein expression (p<0.05) and increased p-Bcl-2 (antiapoptotic) protein expression (p<0.05) thereby maintaining mitochondrial membrane potential (p=0.5643). In Thp-1 cells, FA had no effect on the protein expression of Bax (p=0.6130) but significantly decreased the protein expression of p-Bcl-2 (p<0.005) with a corresponding increase in mitochondrial depolarization (p<0.005). In addition, FA increased oxidative stress (MDA levels) in both PBMCs (p<0.005) and Thp-1 cells (p<0.005) contributing to cellular damage and cellular signaling; and substantially decreased ATP levels in both PBMCs (p<0.005) and Thp-1 cells (p<0.005). Additionally, FA significantly increased phosphorylation of p-ERK (42kDa - p<0.05; 44kDa - p<0.005), p-JNK (46kDa - p<0.005; 54kDa - p<0.05) and p38 (p<0.05); and slightly increased the phosphorylation of p-Akt (p=0.1640) in PBMCs treated with FA. In Thp-1 cells, FA significantly up-regulated p-Akt (p<0.05) and p-ERK (42kDa - p<0.0001; 44kDa - p<0.005) expressions and significantly decreased p-JNK (46kDa - p<0.05; 54kDa - p<0.005) expression but had no effect on the expression of p38 (p=0.8446). This suggests the involvement of MAPK signaling in the induction of cell death in PBMCs and Thp-1 cells treated with FA. This study found that FA is immunotoxic to healthy human PBMCs and Thp-1 cells.
The role of the uncoupling protein2 -866G/A polymorphism in oxidative stress markers associated with air pollution exposure during pregnancy.
(2012) Nagiah, Savania.; Phulukdaree, Alisa.; Chuturgoon, Anil Amichund.
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.