Hydrogen Peroxide and Amyotrophic Lateral Sclerosis: From Biochemistry to Pathophysiology

Free radicals are unstable chemical reactive species produced during Redox dyshomeostasis (RDH) inside living cells and are implicated in the pathogenesis of various neurodegenerative diseases. One of the most complicated and life-threatening motor neurodegenerative diseases (MND) is amyotrophic lateral sclerosis (ALS) because of the poor understanding of its pathophysiology and absence of an effective treatment for its cure. During the last 25 years, researchers around the globe have focused their interest on copper/zinc superoxide dismutase (Cu/Zn SOD, SOD1) protein after the landmark discovery of mutant SOD1 (mSOD1) gene as a risk factor for ALS. Substantial evidence suggests that toxic gain of function due to redox disturbance caused by reactive oxygen species (ROS) changes the biophysical properties of native SOD1 protein thus, instigating its fibrillization and misfolding. These abnormal misfolding aggregates or inclusions of SOD1 play a role in the pathogenesis of both forms of ALS, i.e., Sporadic ALS (sALS) and familial ALS (fALS). However, what leads to a decrease in the stability and misfolding of SOD1 is still in question and our scientific knowledge is scarce. A large number of studies have been conducted in this area to explore the biochemical mechanistic pathway of SOD1 aggregation. Several studies, over the past two decades, have shown that the SOD1-catalyzed biochemical reaction product hydrogen peroxide (H2O2) at a pathological concentration act as a substrate to trigger the misfolding trajectories and toxicity of SOD1 in the pathogenesis of ALS. These toxic aggregates of SOD1 also cause aberrant localization of TAR-DNA binding protein 43 (TDP-43), which is characteristic of neuronal cytoplasmic inclusions (NCI) found in ALS. Here in this review, we present the evidence implicating the pivotal role of H2O2 in modulating the toxicity of SOD1 in the pathophysiology of the incurable and highly complex disease ALS. Also, highlighting the role of H2O2 in ALS, we believe will encourage scientists to target pathological concentrations of H2O2 thereby halting the misfolding of SOD1.

Molecular Mechanism of Changes in Gene Expression of Cytosolic Cu/Zn SOD, Lipid Peroxidation and Hepatic Function Impairments during Experimental Fluorosis

Background: Long term intake of high amount of fluoride leads to fluorosis causing metabolic, functional and structural damages affecting many tissues and organs including dental and skeletal manifestations. The liver is the most susceptible organ to fluoride toxicity because of its active and major role in digestion and detoxification. Aim: The present study aims to elucidate the effects of sodium fluoride on hepatic function biomarkers, lipid peroxidation and gene expression of Cu/Zn SOD in albino rats. Materials and Methods: Wistar albino rats were randomly assigned to three groups. The control rats were given 1 ml deionized water orally for 40 days. Groups II and III were administered 300 and 600 mg NaF/kg b.w. /day for the same period. Animals were sacrificed under anaesthesia, liver tissue was excised and used for biochemical and molecular analysis. The level of fluoride and lipid peroxidation (MDA) as well as reduced glutathione (GSH) content was determined. The activities of cytosolic copper/zinc superoxide dismutase (Cu/Zn SOD), aminotransferases (ALT and AST), lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) in the hepatic tissue were determined. The analysis of gene expression of Cu/Zn SOD in the liver was determined using Real-time PCR. Results: The results revealed significantly (P<0.0001) higher concentration of fluoride and MDA in the liver of rats exposed to fluoride when compared to control. The GSH content reduced significantly (P<0.0001) in fluorotic rats. The activities of hepatic function biomarkers viz; ALT, AST, LDH and ALP elevated significantly (P<0.0001) compared to the control. An elevation of 108.60% (ALP), 121.45% (AST), 33.77% (LDH) and least 24.40% (ALT) was found in rats treated with 300 mg/L fluoride and maximum elevation of 226.20% (ALP), 211.52% (AST), 57.75% (LDH) and least 56.79% (ALT) was registered in rats exposed to 600 mg/L fluoride in drinking water. The activity of cytosolic Cu/Zn SOD decreased significantly (P<0.0001) in fluorotic rats. Pearson’s bivariate correlation and simple linear regression analysis exhibited strong positive correlation between level of hepatic tissue fluoride and activities of ALT (Pearson r= 0.85), AST (Pearson r= 0.94), LDH (Pearson r= 0.89), ALP (Pearson r= 0.86) and in MDA (Pearson r=0.984) while negative correlations existed between levels of fluoride and GSH (Pearson r=-0.93) as well as activity of Cu/Zn SOD (Pearson r= -0.99). The gene expression of cytosolic Cu/Zn SOD was significantly (P<0.0001) reduced in fluorotic rats. Conclusion: The present study revealed that fluoride declined the antioxidant activity of hepatic Cu/Zn SOD at biochemical as well as molecular level which leads to oxidative stress and tissue damage. This further affirms by increased activities of hepatic function biomarkers in correlation with high fluoride level during experimental fluorosis.

Coping with oxidative stress in extreme environments: the distinctive roles played by Acinetobacter sp. Ver3 superoxide dismutases

ABSTRACTAcinetobacter sp. Ver3 is a polyextremophilic strain characterized by a high tolerance to radiation and pro-oxidants. The Ver3 genome comprises the sodB and sodC genes encoding an iron (AV3SodB) and a copper/zinc superoxide dismutase (AV3SodC), respectively; however, the specific role(s) of these genes has remained elusive. We show that the expression of sodB remained unaltered in different oxidative stress conditions whereas sodC was up-regulated in the presence of blue light. Besides, we studied the changes in the in vitro activity of each SOD enzyme in response to diverse agents and solved the crystal structure of AV3SodB at 1.34 Å, one of the highest resolutions achieved for a SOD. Cell fractionation studies interestingly revealed that AV3SodB is located in the cytosol whereas AV3SodC is also found in the periplasm. Consistently, a bioinformatic analysis of the genomes of 53 Acinetobacter species pointed out the presence of at least one SOD type in each compartment, suggesting that these enzymes are separately required to cope with oxidative stress. Surprisingly, AV3SodC was found in an active state also in outer membrane vesicles, probably exerting a protective role. Overall, our multidisciplinary approach highlights the relevance of SOD enzymes when Acinetobacter spp. are confronted with oxidizing agents.

Activity of CdTe Quantum-Dot-Tagged Superoxide Dismutase and Its Analysis in Capillary Electrophoresis

Quantum dots (QDs) have a broad range of applications in cell biolabeling, cancer treatment, metastasis imaging, and therapeutic drug monitoring. Despite their wide use, relatively little is known about their influence on other molecules. Interactions between QDs and proteins can influence the properties of both nanoparticles and proteins. The effect of mercaptosuccinic acid-capped CdTe QDs on intercellular copper–zinc superoxide dismutase (SOD1)—one of the main enzymatic antioxidants—was investigated. Incubation of SOD1 with QDs caused an increase in SOD1 activity, unlike in the case of CdCl2, which inhibited SOD1. Moreover, this effect on SOD1 increased with the size and potential of QDs, although the effect became clearly visible in higher concentrations of QDs. The intensity of QD-SOD1 fluorescence, analyzed with the use of capillary electrophoresis with laser-induced fluorescence detection, was dependent on SOD1 concentration. In the case of green QDs, the fluorescence signal decreased with increasing SOD1 concentration. In contrast, the signal strength for Y-QD complexes was not dependent on SOD1 dilutions. The migration time of QDs and their complexes with SOD1 varied depending on the type of QD used. The migration time of G-QD complexes with SOD1 differed slightly. However, in the case of Y-QD complexes with SOD1, the differences in the migration time were not dependent on SOD concentration. This research shows that QDs interact with SOD1 and the influence of QDs on SOD activity is size-dependent. With this knowledge, one might be able to control the activation/inhibition of specific enzymes, such as SOD1.

Antioxidant Fusion Protein SOD1-Tat Increases the Engraftment Efficiency of Total Bone Marrow Cells in Irradiated Mice

Total body irradiation is a standard procedure of bone marrow transplantation (BMT) which causes a rapid increase in reactive oxygen species (ROS) in the bone marrow microenvironment during BMT. The increase in ROS reduces the engraftment ability of donor cells, thereby affecting the bone marrow recovery of recipients after BMT. In the early weeks following transplantation, recipients are at high risk of severe infection due to weakened hematopoiesis. Thus, it is imperative to improve engraftment capacity and accelerate bone marrow recovery in BMT recipients. In this study, we constructed recombinant copper/zinc superoxide dismutase 1 (SOD1) fused with the cell-penetrating peptide (CPP), the trans-activator of transcription (Tat), and showed that this fusion protein has penetrating ability and antioxidant activity in both RAW264.7 cells and bone marrow cells in vitro. Furthermore, irradiated mice transplanted with SOD1-Tat-treated total bone marrow donor cells showed an increase in total bone marrow engraftment capacity two weeks after transplantation. This study explored an innovative method for enhancing engraftment efficiency and highlights the potential of CPP-SOD1 in ROS manipulation during BMT.