About the author elena

Facts About Superoxide Dismutases: Role In Redox Signaling Revealed

Facts About Superoxide Dismutases: Role In Redox Signaling Revealed

The SODs represent the first enzymatic defense system against radical damage by oxygen: thus, this enzyme is essential for all aerobic organisms, but not for anaerobes. In support of this hypothesis, McCord believed that the existence of an aerobic organism depends mainly on its ability to produce SODs since its deficiency is responsible for oxygen sensitivity and allows survival only in an anaerobic environment.

In physiological conditions, the superoxide dismutases, together with the non-enzymatic ROS scavengers as vitamins E, A, and C maintain a steady state between oxidant and antioxidant systems (Russo et al., 2011). The dysregulation in redox homeostasis, determined by an imbalance between ROS production and scavenging capacity, determines considerable cellular damage as membrane lipoperoxidation, nucleic acid and structural alterations of proteins contributing to neurodegenerative and cardiovascular diseases.

In the last years, many data obtained in in vitro studies performed in many cellular lines, mainly neuroblastoma SK-N-BE cells, indicate that SOD1 is secreted and is able to activate, through muscarinic M1 receptor, cellular pathways involving ERK1/2 and AKT activation; these effects are associated with intracellular calcium increase that is further accentuated when these cells are stimulated with mutated SOD1G93A.

The intracellular cytosolic SOD1 localization has been a matter of debate; recent evidences, performed in transfected mouse neuroblastoma neuro2 cells, demonstrated that both wild type SOD1 (wt-SOD1) and SOD1 mutants are distributed into luminal structures of endoplasmic and Golgi apparatus (Urushitani et al., 2008). The first experimental evidence that some cellular lines could be able to secrete the Cu,Zn superoxide dismutase date back to many years ago when we, for the first time, showed the secretion of this protein by experiments performed in hepatocytes and fibroblasts (Mondola et al., 1996), neuroblastoma SK-N-BE cells (Mondola et al., 1998; Gomes et al., 2007; Polazzi et al., 2013) and in thymus derived epithelial cells (Cimini et al., 2002).

In addition, we demonstrated that in human neuroblastoma SK-N-BE cells, that show a greater sensitivity to glucose deprivation-induced cytotoxicity due to enhanced sensitivity to ROS (Shutt et al., 2010), SOD1 export takes place in normal conditions and is increased following oxidative stress (Mondola et al., 1996, 1998). Successively, we showed that SOD1 in human neuroblastoma SK-N-BE cells is exported through a microvesicular secretory pathway that is impaired by brefeldin-A (BFA), and by 2-deoxyglucose, and sodium azide, which reduces ATP intracellular pool (Mondola et al., 2003).

Indicators on The Role Of Manganese Superoxide Dismutase In … – Hindawi You Need To Know

The Best Guide To Superoxide Dismutase


The Ultimate Guide To Brenda - Information On Ec - Superoxide Dismutase


Another important aspect was the discovery that besides the constitutive SOD1 export, the secretion of this enzyme is also induced. To this respect, we showed (Santillo et al., 2007) that SOD1 is actively released from rat brain synaptosomes as well as from rat pituitary GH3 cells that represent a good model to study the inducible SOD1 release since they possess all the neuronal protein machinery involved in synaptic vesicle exocytosis.

In addition, in the attempt to evaluate the possible role carried out by SOD1 export, we recently demonstrated, in SK-N-BE neuroblastoma cell line, that this enzyme is able, through the involvement of muscarinic M1 receptor, to activate ERK1/2 and AKT in a dose and time-dependent manner. This effect was remarkably reduced by M1 receptor silencing as well as by using M1 antagonist pirenzepine (Damiano et al., 2013).

However, FGF-1 and the 18 kDa isoform of FGF-2 have been shown to be secreted by an alternative pathway being directly translocated from the cytoplasm into the extracellular space. Analogously, also interleukin 1β (IL-1β) has been reported to be secreted by a vesicular non-classical export pathway. Soluble proteins classically contain N-terminal signal peptides that direct them to the translocation apparatus of the Endoplasmic Reticulum (ER) (Walter et al., 1984).

In addition, non-classical protein secretion is both energy and temperature dependent and can be stimulated or inhibited by various treatments (Cleves, 1997; Hughes, 1999). The list of proteins that could be exported from cells in the absence of a functional ERG system (unconventional secretory pathway), as IL-1β and galectin-1 (also referred to as L-14), is continually growing; for further data see the review of Nickel (2003).

Superoxidedismutase (Sod) Fundamentals Explained


For this reason SOD1 secretion should bypass the canonical ERG secretory pathway. We previously demonstrated that BFA as well as 2-deoxyglucose and sodium azide (NaN3), impairs SOD1 export (Mondola et al., 2003). In our opinion, the treatment with BFA probably dysregulates not only the classical secretory ERG pathway but also the microvesicular membrane traffic of unconventional protein secretion or alternative protein export.

The 20-Second Trick For Superoxide Dismutase: Health Benefits, Uses, Side Effects …

Furthermore, these authors showed that SOD1A4V inhibits secretory protein transport from the ER to the Golgi apparatus. Amyotrophic lateral sclerosis (ALS) is an adult onset, neurodegenerative disease characterized by selective death of the upper and lower motor neurons of the brain and spinal cord. Symptoms include muscle atrophy, spasticity, paralysis and eventual death from respiratory failure within 3–5 years of diagnosis.

Clinical and pathological processes indicate that ER stress represents a key pathway involved in cell death. In the transgenic SOD1G93A ALS rat model unfolded protein response and ER stress-induced apoptosis has been observed (Atkin et al., 2006); an unfolded protein response, including induction of stress sensor kinases, chaperones, and apoptotic mediators, has been shown also in spinal cord motor neurons of human patients with the sporadic form of ALS (sALS) that is not restricted to SOD1 mutations (Atkin et al., 2008).

SOD1 and other proteins are misfolded in fALS and in sALS, but it is not clear how this triggers ER stress, fragmentation of the Golgi apparatus, disruption of axonal transport and apoptosis. Nearly 20% of fALS is caused by SOD1 gene mutations (Neumann et al., 2006). Indeed, the majority of SOD1 mutants maintain their enzymatic activity suggesting the occurrence of gain of toxic activity function rather than a simple loss of function (Strong et al., 2005; Dion et al., 2009).

(2005) demonstrated an impaired constitutive extracellular secretion of mutant SOD1 in NSC-34 cells that induces frequent cytoplasmic inclusions and protein insolubility. These data link the deficient secretion of mutant SOD1 with intracellular protein aggregates and toxicity in NSC-34 cells. In addition, these authors showed that in a transgenic rat model of ALS the chronic intraspinal infusion of exogenous human wt-SOD1 significantly delayed disease progression suggesting a novel extracellular role for SOD1 in ALS; therefore extracellular delivery of human wt-SOD1 could improve clinical disease in transgenic ALS rats supporting a novel extracellular role for mutant and wt-SOD1 in ALS pathogenesis and therapy, respectively.

In addition, in transgenic mice, carrying SOD1 mutations, toxic effects to motor neurons by microglia activation were observed (Urushitani et al., 2006; Zhao et al., 2010). The microglia cells can carry out an important role in ALS progression (Pramatarova et al., 2001) since microglia activation can be observed before neuron loss in transgenic mice expressing human SOD1 mutants (Alexianu et al., 2001).

DPPH Method


Cell damages are induced by Reactive Oxygen Species (ROS). ROS are free radicals, reactive anions containing oxygen atoms or oxygen containing molecules able to generate free radicals. Some examples are hydroxyl radical, superoxide and hydrogen peroxide.

Main source of ROS in vivo is aerobic respiration, but ROS are also produced during beta-oxidation of fatty acids, in the xenobiotic compounds metabolism trough cytochrome P450, in phagocytosis stimulation of pathogens or lipopolysaccharides, etc. ROS and oxidative stress in general are involved in some chronic conditions such as Alzheimer and Parkinson disease, cancer and aging.

Figure: Main oxigen radical species


Starting from an O2 molecule and adding one electron to the external orbital the reduction product of molecular oxygen: the superoxide anion (O2 .- ). It is produced during the oxidative phosphorylation, by enzymes (i.e. xanthine oxidase) and leukocytes. Due to its toxicity all aerobic organisms developed different isoforms of the antagonist enzyme: the superoxide dismutase (SOD). SOD is a very efficient enzyme able to combine the superoxide anion with two H+ catalyzing the dismutation reaction through a metal based co-factor yielding H2O2 and O2 as final products. If not properly and promptly inactivated the superoxide anion can create damages to membranes lipids, proteins and DNA.

Figure: Superoxide radical


In normal conditions, in our body, ROS are inactivated through enzymes such as superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx). SOD is a key enzyme able to inactivate the superoxide radical, one of the most reactive and therefore the most dangerous radical species.

Figure: In vivo generation of superoxide anion and its enzymatic inactivation paths


To reduces the harmful effects of ROS, cells have developed different defensive strategies including enzymatic and non-enzymatic systems. Considering the antioxidant enzymes, some of these are playing a preventative role eliminating directly ROS. Among these enzymes superoxide dismutase is the first line of defense removing the superoxide anion, the first and most reactive radical derived by molecular oxygen. SOD is therefore one of the main antioxidant defensive system present in almost all the cells exposed to oxygen. The SOD catalyzed reaction is a dismutation with a second-order kinetic based on the following half reactions:

DPPH Method

The antiradical capacity has been assessed using the DPPH method. The sample is placed in a concentrated solution of a standard free radical (1,1-diphenyl-2-picryl-hydrazyl) and its concentration is measured via spectrophotometry to assess the ability of the phytocomplex to quench the radicals. Superox-D has an high antiradical capacity due to quenching mechanisms.

16 folds more antiradical compared to melon

37 folds more antiradical compared to SOD from melon

Figure: Structure of the radical DPPH


Antiradical capacity (DPPH method) of carot, melon and commercial SOD from melon





Doddigarla Z Correlation of serum chromium, zinc, magnesium and SOD levels with HbA1c in type 2 diabetes: A cross sectional analysis. Diabet metab Syndrom. 2016 Jan-Mar;10(1 Suppl 1):S126-9. doi: 10.1.

Vouldoukis I, Conti M, Krauss P, et al. Supplementation with gliadin-combined plant superoxide dismutase extract promotes antioxidant defences and protects against oxidative stress. Phytother Res. 2004 Dec;18(12):957-62.

Vouldoukis I, Lacan D, Kamate C, et al. Antioxidant and anti-inflammatory properties of a Cucumis melo LC. extract rich in superoxide dismutase activity. J Ethnopharmacol. 2004 Sep;94(1):67-75.

Muth CM, Glenz Y, Klaus M, et al. Influence of an orally effective SOD on hyperbaric oxygen-related cell damage. Free Radic Res. 2004 Sep;38(9):927-32.

Barouki R. Ageing free radicals and cellular stress. Med Sci (Paris). 2006 Mar;22(3):266-72.

Faraci FM, Didion SP. Vascular protection: superoxide dismutase isoforms in the vessel wall. Arterioscler Thromb Vasc Biol. 2004 Aug;24(8):1367-73.

Fukai T, Folz RJ, Landmesser U, Harrison DG. Extracellular superoxide dismutase and cardiovascular disease. Cardiovasc Res. 2002 Aug 1;55(2):239-49.

Petersen SV, Oury TD, Ostergaard L, et al. Extracellular superoxide dismutase (EC-SOD) binds to type i collagen and protects against oxidative fragmentation. J Biol Chem. 2004 Apr 2;279(14):13705-10.

Maier CM, Chan PH. Role of superoxide dismutases in oxidative damage and neurodegenerative disorders. Neuroscientist. 2002 Aug;8(4):323-34.

Fattman CL, Schaefer LM, Oury TD. Extracellular superoxide dismutase in biology and medicine. Free Radic Biol Med. 2003 Aug 1;35(3):236-56.

Chung JM. The role of reactive oxygen species (ROS) in persistent pain. Mol Interv. 2004 Oct;4(5):248-50.

Bae SC, Kim SJ, Sung MK. Inadequate antioxidant nutrient intake and altered plasma antioxidant status of rheumatoid arthritis patients. J Am Coll Nutr. 2003 Aug;22(4):311-5.

Zawadzka-Bartczak E. Activities of red blood cell anti-oxidative enzymes (SOD, GPx) and total anti-oxidative capacity of serum (TAS) in men with coronary atherosclerosis and in healthy pilots. Med Sci Monit. 2005 Sep;11(9):CR440-4.

Gow A, Ischiropoulos H. Super-SOD: superoxide dismutase chimera fights off inflammation. Am J Physiol Lung Cell Mol Physiol. 2003 Jun;284(6):L915-6.

Flohe L. Superoxide dismutase for therapeutic use: clinical experience, dead ends and hopes. Mol Cell Biochem. 1988 Dec;84(2):123-31.

Carlo MD, Jr., Loeser RF. Increased oxidative stress with aging reduces chondrocyte survival: correlation with intracellular glutathione levels. Arthritis Rheum. 2003 Dec;48(12):3419-30.

Junqueira VB, Barros SB, Chan SS, et al. Aging and oxidative stress. Mol Aspects Med. 2004 Feb;25(1-2):5-16.

Vina J, Lloret A, Orti R, Alonso D. Molecular bases of the treatment of Alzheimer’s disease with antioxidants: prevention of oxidative stress. Mol Aspects Med. 2004 Feb;25(1-2):117-23.

Okada F, Shionoya H, Kobayashi M, et al. Prevention of inflammation-mediated acquisition of metastatic properties of benign mouse fibrosarcoma cells by administration of an orally available SOD. Br J Cancer. 2006 Mar 27;94(6):854-62.

Benedetti S, Lamorgese A, Piersantelli M, Pagliarani S, Benvenuti F, Canestrari F. Oxidative stress and antioxidant status in patients undergoing prolonged exposure to hyperbaric oxygen. Clin Biochem. 2004 Apr;37(4):312-7.

Dennog C, Radermacher P, Barnett YA, Speit G. Antioxidant status in humans after exposure to hyperbaric oxygen. Mutat Res. 1999 Jul 16;428(1-2):83-9.

Levin ED. Extracellular superoxide dismutase (EC-SOD) quenches free radicals and attenuates age-related cognitive decline: opportunities for novel drug development in aging. Curr Alzheimer Res. 2005 Apr;2(2):191-6. R

Superox-D: effects on intestine epithelium model

Superox-D has been tested for its ability of being up taken and able to provide protection at intestinal level. A human intestine model of enterocyte-like cell Caco-2 was selected been widely used as a model of the intestinal epithelial barrier. It has been well documented that Caco-2 monolayers represent a reliable correlate for studies on the absorption of drugs and other compounds after oral intake in humans.

Up take studies


To assess the ability of being efficiently absorbed in the intestine, the Caco-2 cell model were incubated for 2 hours with media containing different concentration of Superox-D. The ability of Superox-D of being up taken was assessed measuring the Total Antioxidant Activity (TAA) of cell cytosol. The data reported in Figure show that Superox-D is able to effectively penetrate the cell membrane and to induce an increased antioxidant capacity in cell cytosol. This effect is due to the ability of Superox-D of being promptly up taken by the intestine proving the high bioavailability of the product.

Antioxidant protection studies

The protective effect of Superox-D was assessed by measuring the ability of some enterocyte-like cell Caco-2 model culture of better resisting to intense antioxidant stress. The cell lines were pre treated for 2 hours with media containing different concentration of Superox-D. After the incubation with Superox-D a fresh control medium was used in all samples and the effect of an oxidative stress caused by tert-butyl hydroperoxide (t-BOOH) was assessed with a fluorometric assessment. As shown in Figure the cell lines pre treated with Superox-D proved to be far more resistant to the radical stress induced during the assay with a dose dependent behavior. Superox-D is therefore able of keeping the intestine protected from the harmful radical species.