Free Radical Biology of & Endocrine, Metabolic Immune Disorders

By Bentham Science Publishers

This reference explores the science of signaling mechanisms associated with diseases like endocrine, metabolic and immune disorders which are linked to oxidative stress mediated disease mechanisms. The common diseases in these categories include diabetes mellitus and arthritis and conditions defined by inflammation and autoimmunity. These diseases involve metabolic pathways mediated by reactive oxygen species or free radicals. The pathways are targets in the mechanism of drugs which aim to treat related disorders. The book covers key topics in free radical biology that help to understand the nature of the pathways and the pharmacology of the drugs that are designed to target them.5 chapters elucidate the free radical biology of the diseases.1. Role of free radical biology in diabetes mellitus2. Role of GSK3 in regulation of insulin release and glucose metabolism3. Regulatory role of NRF2 in rheumatoid arthritis4. Role of free radical biology in Alzheimer’s disease5. Regulatory role of immune cells mediated antibody on rheumatoid arthritis Key features:- Elucidates the key biochemical and pharmacological mechanisms that are mediated by free radicals in common endocrine, metabolic and immune diseases- Explains the activation of immunological factors (like RF and ACPA) that trigger inflammation and arthritis- Covers the role of free radicals in Alzheimer’s disease and new interventions that target mitochondrial mechanisms- Systematically explains the molecular biology of free radicals with the help of schematic diagrams - Includes references for further reading The accessible and structured text in this reference make it a suitable resource for all biomedical scientists, faculty and postgraduate students in academia and industry.

• Language: English
• Publisher: Bentham Science Publishers
• Release date: Aug 16, 2001
• ISBN: 9789815049220

Book preview

Free Radical Biology of Diabetes Mellitus

Bidita Khandelwal¹, *, Chamma Gupta², Levin Thomas³, Sonal Sekhar Miraj³, Mahadev Rao³, Rubi Dey⁴, Karma Gurmey Dolma⁵, Rinchen Doma Bhutia², Vijay Pratap Singh⁶

¹ Department of Medicine, Sikkim Manipal Institute of Medical Sciences, Sikkim Manipal University, Gangtok, Sikkim-737102, India

² Department of Biochemistry, Sikkim Manipal Institute of Medical Sciences, Sikkim Manipal University, Gangtok, Sikkim-737102, India

³ Department of Pharmacy Practice, Manipal College of Pharmaceutical Sciences, Manipal Academy of Higher Education, Manipal, Karnataka-576104, India

⁴ Department of Physiology, Sikkim Manipal Institute of Medical Sciences, Sikkim Manipal University, Gangtok, Sikkim-737102, India

⁵ Department of Microbiology, Sikkim Manipal Institute of Medical Sciences, Sikkim Manipal University, Gangtok, Sikkim-737102, India

⁶ Department of Physiotherapy, Kasturba Medical College, Manipal Academy of Higher Education, Manipal, 576104 India

Abstract

Free radicals play a pivotal role in the etiology of different diseases, including diabetes mellitus (DM). In the past three decades, the understanding of the fundamental role of free radicals in the etiology and disease progression of DM was studied broadly. This chapter aimed to enumerate the recent progress in the areas of free radical biology for the management of DM. Free radicals, as well as reactive oxygen species (ROS), having extra electrons in their outer orbitals, react with all biomolecules, including protein, lipids, and DNA, causing oxidative stress and damage. DM is also associated with oxidative stress induced by the elevated production of free radicals or reduced antioxidant activity. Recently, the importance of an antioxidant rich diet, yoga, and exercise has been well documented for the management of DM. Studies confirmed that exercise-induced ROS is an acute effect, while the chronic effect of exercise produces endogenous antioxidant defences and promotes a state of endogenous antioxidant defence mechanism. Therefore, regulating oxidative stress will lead to a significant future area of research for DM disease management.

Keywords: Antioxidant, Diabetes Mellitus, Oxidative damage, Oxidative stress, Reactive oxygen species.

---

* Corresponding author Bidita Khandelwal: Department of Medicine, Sikkim Manipal Institute of Medical Sciences, Sikkim Manipal University, Gangtok, Sikkim-737102, India; E-mail: [e-mail]

INTRODUCTION-FREE RADICALS

Free radicals have acquired a significant momentum in biology along with other oxidants because of their crucial functions in numerous physiological states and pathological implications in a wide range of disorders [1]. Chemical entities like atoms, molecules, and ions with one or more unpaired electrons in their outer orbitals, are referred to as free radicals, which generally display remarkable reactivities and also show independent existence ability [1, 2]. The odd number of electrons in free radical has been attributed to its instability, short-life, and higher reactivity [2]. These radicals have the ability to remove the electrons from another compound or biomolecules to become stable due to their greater reactivity. Thereby, the molecule undergoing an attack gives up an electron and is subsequently converted to a free radical. This process sets off a chain of reactions that causes injury or kills the cell which is alive [3].

Free radicals act as both harmful as well as valuable substances [4]. Therefore, they might be recognized as a necessary evil for signalling in the regular differentiation and migration process [5]. These species are generated during normal cellular metabolism or external sources such as radiation, pollution, smoking, etc. [4]. When cells utilize oxygen to produce energy, these species are released by the mitochondrial adenosine triphosphate (ATP) synthesis pathway. As a by-product of the cellular redox process, reactive nitrogen species (RNS), as well as reactive oxygen species (ROS), are released. The vital aspect of life is maintaining a subtle equilibrium between these two species’ antagonistic effects. An adequate quantity of RNS and ROS exerts favourable actions on different cellular responses, signalling pathways, mitogenic responses, redox regulation, and immune function [6, 7]. However, at high concentrations, both the species exert oxidative stress as well as nitrosative stress, producing potential deleterious effects on the biomolecules like lipids, proteins, nucleic acids, and others [8-13]. Overload of ROS/RNS, as well as a deficit of antioxidants (enzymatic as well as non-enzymatic), can produce oxidative and nitrosative stress. These stresses play a significant role in aging as well as developing chronic and degenerative disorders (Fig. 1). On the other hand, both endogenous and exogenous antioxidants serve as free radical scavengers and subsequently inhibit or repair injuries produced by free radicals [14-16].

Fig. (1))

Physiological and pathological role of free radicals in the biological system.

HISTORY ON THE CONCEPT OF FREE RADICALS IN BIOLOGICAL SYSTEM

The word ‘radical’ was first introduced in 1786 by French chemist Louis-Bernard Guyton de Morveau. Later, Gay-Lussac, Justus von Liebig, and Jöns Jakob Berzelius used the term to denote clusters of atoms that remained unaltered in several compounds. Early works involving free radicals were oxidative degradation of organic substances by hydrogen peroxide (H2O2) in the presence of ferrous (Fe²+) under acidic media, primarily discovered by British chemist Henry John Horstman Fenton in 1894. Subsequently, the reaction was applied to tartaric acid oxidation, with the Fenton reagent as the catalyst, and the process became known as Fenton reactions [17]. At that time, free radicals were unknown, and almost 30 years later, the hydroxyl radicals’ (OH•) role was suggested by Fritz Haber and Richard Willstätter [18]. Moses Gomberg, a chemistry professor at the University of Michigan, first prepared an organic radical, triphenylmethyl radical [(C6H5)3C•], and speculated the existence of the radical in the biological system in 1900 [19]. In 1922, Robert Williams Wood isolated and characterized atomic hydrogen for the first time in an electrical discharge tube [20]. Thereafter, atomic hydrogen’s chemical nature was experimented thoroughly by Karl Friedrich Bonhoeffer, a German physical chemist, in 1924 [21]. Methyl free radical (CH3•) was synthesized in 1929 by Friedrich Paneth, along with Wilhelm Hofeditz. To synthesize the CH3•, they followed pyrolysis of tetramethyl lead by adopting a system that Bonhoeffer used to investigate atomic hydrogen [22]. Elucidation of a free-radical mechanism and subsequent discovery of the peroxide effect has been credited to Morris Selig Kharasch and his fellow student, Frank Mayo, in 1933. Successively, they applied this mechanism to various other chemical systems too. They employed a free radical mechanism to add hydrogen bromide to olefins. Leonor Michaelis’s, a German biochemist, curiosity peaked when he found that free radicals were naturally occurring metabolic intermediates in a biological system and went on to observe the oxidation-reduction potential curves generated after enhancing the concentrations of oxidants added to hydroquinone in 1930. An electron loss at the onset was followed by the escape of an additional electron [23]. Michaelis believed that the initial electron loss led to the development of semiquinone radical while the subsequent loss of the later formed quinone, a wholly oxidized form.

Later in 1954, Rebeca Gershman and Daniel Gilbert proposed the free radical theory of oxygen toxicity, claiming that oxygen's toxicity is attributable to its capacity to produce free radicals [24]. At the same time, Barry Commoner, Jonathan Townsend, and George E. Pake conducted electronic spin resonance (ESR) studies and confirmed the presence of free radicals in enzyme-substrate systems [25]. In 1956, Denham Harman suggested the free radical theory of aging, which proposed free radicals’ biological implication in the aging process [26]. A new free radical research era was started by two American biochemists, Joe Milton McCord and Irwin Fridovich, in 1969. They were the first to describe superoxide dismutase’s (SOD) enzymatic activity. This is a primary enzymatic defense system for superoxide anion [27]. In 1971, G Loschen demonstrated that ROS are released in cellular metabolic respiration by indicating the production of H2O2 in mitochondria of the pigeon’s heart [28]. In 1977, F Mittal and C K Murad reported that guanylate cyclase enzyme activation by hydroxyl radicals induces second messenger cyclic guanosine monophosphate (cGMP) production [29]. In 1989, Barry Halliwell and John M.C. Gutteridge stated that ROS comprises free radical as well as non-radical oxygen derivatives [30]. In 1993, Reid et al. found the association of free radicals with muscle fatigue [31]. In 2000, V.J. Thannickal and B.L. Fanburg elucidated cellular signalling mechanisms initiated by ROS [32]. In the last few decades, there has been tremendous research on free radicals to understand their involvement in several disease pathogenesis and the protective effects of antioxidants.

CHARACTERISTICS OF FREE RADICALS AND OXIDANTS

Pro-oxidants and oxidants, in general, are referred to as ROS/NOS. Moreover, non-radical derivatives are included in oxidants. Although their reactivity is often more robust, the stability of radicals is relatively lower compared to non-radical entities. In the outer shell, free radicals have one or more unpaired electrons [4, 8-10]. The oxygen molecule is a radical (O2••) referred to as a biradical because of the presence of two unpaired electrons. Generally, free radicals are created when a chemical bond is broken, and each fragment retains an electron, when one radical is cleaved to yield a new radical, or when a redox reaction occurs [8, 9]. Free radicals include ROS such as OH•, superoxide (O2•-), alkoxy radical (RO•), peroxyl (ROO•), and lipid peroxyl (LOO•). Similarly, RNS includes nitric oxide (NO•) and nitrogen dioxide (NO2•). Whereas ROS such as singlet oxygen (¹O2), ozone (O3), H2O2, hypochlorous acid (HOCl), hypobromous acid (HOBr), lipid peroxide (LOOH), and RNS such as nitrous acid (HNO2), dinitrogen trioxide (N2O3), and peroxynitrite (ONOOH) are non- radicals. These are also referred to as oxidants and can readily involve free radical reactions in biological systems [33]. These free radicals are extremely unstable as they possess electrons to carry out reactions with many organic molecules like lipids, proteins, and nucleic acids.

Types of Free Radicals

Free radicals are categorized into the following three types [1, 5, 34]:

Reactive oxygen species (ROS).

Reactive nitrogen species (RNS).

Reactive chlorine species (RCS).

As discussed earlier, ROS and RNS are classified as ‘radicals’ as well as ‘non-radicals’ (Table 1).

Table 1 Classification of reactive oxygen and nitrogen species.

GENERATION/SOURCE OF FREE RADICALS AND OXIDANTS

These radicals can be formed from endogenous as well as exogenous sources. ROS’s endogenous sources comprise various cellular organelles like mitochondria, endoplasmic reticulum, and peroxisomes, where oxygen consumption is high. These free radicals are released from the activation of immune cells, phagocytic cells, prostaglandin synthesis, inflammation, auto-oxidation of adrenalin, mental distress, intensive exercise, ischemia, infection, neoplasia, and aging [35]. Whereas exogenous sources include water and air pollution, alcohol, tobacco smoke, high temperature, cooking (used oil, fat, smoked meat), pesticides, industrial effluents, heavy metals (Fe, Cu, Cr), transition metals (Pb, As, Hg), radiation (UV) and iatrogenic (acetaminophen, halothane, bleomycin, gentamycin, tacrolimus, etc.) [4, 36-38].

Intracellular production of the free radicals can happen in two types of reactions, either enzymatic or non-enzymatic ways. The former process of producing free radicals involves prostaglandin synthesis, phagocytosis, respiratory chain, and the cytochrome P450 system [3, 19, 24, 27-30]. Superoxide radicals are generated by various cellular oxidase systems, including xanthine oxidase, peroxidases, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, etc. Complex I, as well as complex III, are the major sites of superoxide formation in the electron transport chain (ETC) [39]. Once generated, the radical participates in multiple reactions yielding several ROS and RNS (Table 1). On the other hand, the non-enzymatic process of organic substances with oxygen and the ionizing radiations also produces free radicals in the mitochondria during oxidative phosphorylation [4, 33, 36].

BIOLOGICAL ROLE OF FREE RADICALS AND OXIDANTS

As previously mentioned, free radicals are considered a necessary evil since they have a vital role in the origin as well as the evolution of life. In adequate quantity, free radicals are essential for the physiological process, including cellular structures’ maturation, and serve as a biological weapon for the body’s defence system. Invading pathogenic organisms will be targeted by the free radicals released from phagocytes, thereby working as the host defence towards infections [36, 40]. The body’s immune system merely influences the ROS generation that is exemplified in individuals suffering from granulomatous disease [36]. Since these patients possess an impaired membrane-bound NADPH oxidase system, resulting in failure superoxide radical generation, consequently produce recurrent infections.

Another significant physiological role of the free radicals is that it is essential for modulating various intracellular signalling pathways. These contain mitogen-activated protein kinase (MAPK) (induces mitogenic responses) as well as extra-cellular-signal-regulated kinase (ERK) pathways, which regulates the expression of the gene and, together with SOD, initiates cellular necrosis [41]. For example, neurons generated RNS acts as neurotransmitters, and macrophages produce one act as immunity mediators. Besides, these species are involved in angiogenesis, leukocyte adhesion, and thrombosis and have implications in vascular tone. Likewise, ROS mediates gene transcription and single transduction activities and modulates several cellular processes [42]. The free radical formation by non-phagocytic NADPH oxidase isomers plays a pivotal role in modulating intracellular signalling cascade in different non-phagocytic cells such as thyroid tissue, fibroblasts, cardiac myocytes, endothelial cells, and smooth muscle cells of vasculatures. Indeed, NO is an intracellular messenger, which is involved in the regulation of blood circulation, thrombosis, and various neuronal activities [43]. Additionally, as a nonspecific defense of the host, NO is involved in terminating intracellular pathogenic microbes as well as tumor cells [33, 43].

OXIDATIVE DAMAGE TO DNA, LIPIDS, AND PROTEINS

The loss of homeostatic balance in free radical and ROS production and/ or consumption, in due course of time, damage the macromolecules (Lipids, Proteins and, Carbohydrates), including the nucleic acid [38, 44]. This course of the reaction is referred to as Oxidative stress (OS). Increased OS can either show the way towards disease development or can also be a consequence in the pathophysiology of various diseases such as metabolic syndrome [45], neurodegenerative diseases [46-48], cancers [49], and many more. For example, oxidative stress as an early event can be attributed to an individual’s lifestyle that influences health. Consumption of high calorie diet beyond the body’s energy requirement decrease physical activity, and environmental factors all put in towards an early event in developing OS [50]. Glucose tends to increase the concentration of ROS in the form of superoxide by increasing the activity of the enzyme NADPH oxidase in polymorphonuclear leucocytes and mononuclear cells in the blood. Increased enzyme activity is a function of increased p47 phox protein, an important component of NADPH oxidase [51, 52]. Likewise, excess protein and fat consumption also contribute towards an increase in ROS generation via the same pathway as glucose. They increase ROS generation in its own distinctive pattern [53], which, if continues to prevail, then causes damage to DNA, lipid, and protein. A brief mechanism of the reaction between the different types of ROS with these functional units is hereby underlined.

OXIDATIVE DAMAGE TO LIPID

Oxidative damage to lipid has been clearly explained. PUFA are highly susceptible to reaction with free radicals because of the chemical structure consisting of several double bonds [54], this damages the lipid molecule, and the process of deterioration of lipids by the free radicals is called lipid peroxidation (LPO). LPO proceeds in three stages: initiation, propagation, and termination. Initiation begins with the reaction of PUFA with a free radical, for example, a hydroxyl radical (OH•). The reaction results in the removal of the hydrogen atom from the methylene group adjacent to the C-C cis -double bond of the PUFA molecule. Due to