Biochemistry

Respiratory system

Aoun batch

3

‫رانيا إياد‬ 1

Elana Ibdah

Biochemistry

Respiratory system

Aoun batch

Q: Oxygen is toxic or non toxic molecule? A: It depends on concentration That’s why all O2 concentration in atmosphere is lower than nitrogen concentration in order to be appropriate to our respiratory system For example the atmosphere has a high level of nitrogen which is non toxic which implies that we have certain tissues will be damaged if it found in high O2 concentration. Experimentally we found if there is a high concentration of oxygen especially in new and it’s it will be toxic especially for their respiratory system So we must be careful When we expose individual to high concentration of oxygen to supply him with it due to certain problems and respiratory system example COVID-19. In case of COVID-19:  The survival of patients is very low because we give them oxygen and immunosuppressant to prevent cytokine storm, which is the main cause of death in corona patients why? Because because of immune response the body secrets huge amounts of cytokines (immune modulators: signaling molecules that’s a secreted by immune cells ) then some complications occurs that lead to death  So that we give the patient immunosuppressant which may lead to secondary infections because his immune system is suppressed to avoid cytokine storm so that there’s challenge in dealing with COVID-19. History :  The term “oxidative stress” began to be used frequently in the 1970s.  Toxic effects of molecular oxygen (Gerschman et al., 1954)  Potential contribution of such processes to the phenomenon of aging (Harman, 1956).  The recognition in 1968 that biological systems could produce substantial quantities of the superoxide free radical, O2−, through normal metabolic pathways (McCord and Fridovich, 1968) and that enzymes, the superoxide dismutases (SOD), had evolved with the apparent sole purpose of protecting aerobic organisms from the presumed toxicity of this free radical (McCord et al., 1969 and McCord et al., 1971) spurred much interest.

2

Biochemistry

Respiratory system

Aoun batch

Reactive Oxygen Species (ROS): ROS : highly reactive chemical molecules formed due to the electron acceptability of O2 The study of these compounds is very difficult because they are unstable molecules they can convert from one molecule to another and from one compound to another or they can pass from one side to another very quickly But instead of studying them we study things that give indication about them( enzymes that interacts with them) >> example if you found Overexpression of enzyme that reacts with hydrogen peroxide that indicates that there is oxidative stress by hydrogen peroxide and so on. The terms “oxidative stress” and “free radical damage” are NOT synonymous and may not always be interchangeable. Free radical is generally represented by a superscript dot, (R•) e.g. Oxygen can accept single electrons, up to 2 unpaired electrons (O2) Therefore, “Free radicals (one or more unpaired electrons)” and “reactive oxygen species” (ROS) are also NOT synonymous, as many ROS (singlet oxygen 1O2, hydrogen peroxide H2O2)are NOT free radicals). .. : ً‫ٌعى‬ ROS : highly reactive chemical molecules formed due to the electron acceptability of O₂. Examples of ROS include peroxides, superoxide, hydroxyl radical, singlet oxygen, and alpha-oxygen. Free radicle : a radical is an atom, molecule, or ion that has an unpaired Valence electron.

Not all free radicals are ROS Therefore , “ Free radicals ( one or more unpaired electrons ) “ and “reactive oxygen species “ (ROS) are also NOT synonymous , as many ROS (singlet oxygen O2 , hydrogen peroxide H2O2) are NOT free radicals Free radical damage : there are radicals that are formed ( very active molecules) and have radical electrons that is able to reduce other agents.

3

Biochemistry

Respiratory system

Ros members: i. Superoxide anion radical (O2-• ). v. Lipid peroxide radical (ROO • ). ii. Hydroperoxyl radical (HOO • ). vi. Singlet oxygen (1 O2 ).(not free radical)

Aoun batch

ii. Hydrogen peroxide (H2 O2) (not free radical) vii. Nitric oxide (NO • ) iv. Hydroxyl radical (OH • ) viii. Peroxy nitrite (ONOO-• )

ROS formation:  They are formed naturally (physiologically ) by physiological process (metabolic process).  it caused mainly by mitochondria because it deals with electrons by electron transport chain so it always has electron leakage leads to reactive oxygen species formation (generate toxic molecules like these molecule :very active molecule and can destroy your DNA, proteins, lipids and all biological molecules ) ....‫…بس سبحان هللا اللطيف‬.  we have repairing mechanism in our bodies so that the body can deal with them. Formation : 1. The main source is oxidation process and electron transport chain in mitochondria (leakage of electrons) . ( All of us know that we have electrons have to be transport through the electron transport chain in the mitochondria and when we have more electrons some of these electrons are going to leak from mitochondria and which will result in the formation of free radicals ). 2. natural and physiological source such as the function of macrophages (WBC ) : reactive oxygen species will be generated by G6PD pathway which gives NADPH (reducing agents ) that can consumed to reduce the O2 to have oxygen free radical by NADPH oxidase. Q: Why we have this kind of reactions in our body? A: Simply to generate resource of NADP and the generation of oxygen radical . ‫طب هى بسمم و خطٍر نٍش بذي ٌاي؟‬ ‫ كمهىا قراءة عشان وفهم شى بصٍر‬،macrophage ‫عشان أخهص مه انباكخٍرٌا انهً حمج بهعمخها بال‬ Oxygen radical is toxic so we have SOD (super oxide dismutase) enzyme that can consume oxygen radical and covert it to H2O2 .  Then we have MPO (myeloperoxidase) can consume hydrogen peroxide then convert it two HCLO.  HCLO used to kill the bacteria so this mechanism is generated normally in macrophages in order to reduce defensive Ros. 4

Biochemistry

Respiratory system

Aoun batch

macrophages ‫ مه انمٍخىكىوذرٌا و مه ال‬: ‫ بطرٌقخٍه‬free radicals ‫ٌعىً باخخصار بخىخج عىذي ال‬ . ‫و وقخم انبكخٍرٌا‬HCLO ‫وكىن ال‬ ّ ‫ بىسخخذمها بانمىاعت عشان‬macrophages ‫و انهً بال‬

If the concentration of such kind of molecules increase , they will destroy our macromolecule . ‫ رح ٌضرووا بس‬macrophages ‫ٌعىً إرا صار حكىٌه زٌادة إلنهم سىاء عه طرٌق انمٍخىكىذرٌا أو ال‬ ‫سبحان هللا ربىا حطهىا آنٍاث فً جسمىا حخى وخعامم معهم ( ٌعىً هه بانىضع انطبٍعً مفٍذاث و‬ ) ‫بىحخاجهم بس إرا ارا زادوا رح ٌذمروا انخالٌا حذمٍر‬ …‫طٍب‬ Q: How the body is going to perform defense against these molecules ? A: we have defense mechanism in order to consume these molecules . Q: How we consume ROS ? A:by reduction steps of oxygen

In this figure we can see the gaining of electrons and protons in series of reactions and eventually it will be stable as water Characteristics of the ROS are: a. Extreme reactivity. b. Short life span. c. Generation of new ROS by chain reaction. d. Damage to various tissues.(because they can give the electron to other molecules such as DNA proteins and lipids so they can destroy the structure as well as causing mutations) 5

Biochemistry

Respiratory system

Aoun batch

Generation of ROS :  About 1-4% of oxygen taken up in the body is converted to free radicals.  Mitochondrial ROS production is modulated largely by the rate of ETC.  Xanthine oxidase(related to nucleotide formation) and aldehyde oxidase form superoxide anion radical or hydrogen peroxide.  NADPH oxidase  nitric oxide synthase (NO) will form No as ROS)  Lipooxygenase in platelets and leukocytes

   

ionizing radiation )physical factor Light of appropriate λ can produce singlet oxygen. (physical) Cigarette smoke and air pollutants. (chemical) (cause tissue aging) Under hypoxic conditions, the mitochondrial respiratory chain also produce NO

So all of these causes will lead to molecule damage and mainly DNA damage and if the DNA damaged this will lead to mutation, to cancer , metabolic disorder or genetic disorders. To confirm that oxidative stress is very dangerous in DNA :  there is difference between genomic DNA that found in nucleus and mitochondrial DNA and there and there are cells that depend totally mitochondria to stay alive and DNA damage could lead to total defect-in these cells, for example: the sperm 6

Biochemistry

Respiratory system

Aoun batch

 sperms are composed of (head neck and tail) -neck contain the mitochondria the mitochondria is very huge because it serve as an energy source in the sperm  and this energy source depends on ATP production inside it  and ATP production needs proteins to work  and proteins are translated from genes  so that one of the the infertility problems-is high rate of mutations-in the sperms DNA especially in asthenozoospermia and azospermia  this happens in these patients because they don’t have repairing mechanism and DNA doesn’t have introns.

7

Biochemistry

Polyphenols is also antioxidant which is found in some plants so it is very beneficial to eat and drin . ‫زي الشاي مثال‬

Respiratory system

Aoun batch

Fenton & Haber Weiss reaction both use iron to produce hydroxyl radical which can be neuro-toxic free radical

8

Biochemistry

Respiratory system

Aoun batch

‫و اسخخذاماحها فً مىاد انخجمٍم و مىاد انخىظٍف ( احها بخشخغم‬H2O2 ‫هىن انذكخىر حكا شغهت عه ال‬ ٠٣:٠٣ ‫كمبٍض) حسٍخها مش مهمت فما كخبخها بس بخقذروا حرجعىنها بانذقٍقت‬ Q: What that has to do with Oxygen ?! ** O2 is good: The oxygen is good for certain physiological functions :  O2 is an abundant element on the earth (49 %).  20 % of the atmosphere.  65 of Human body !  Cellular respiration.  Other metabolic reactions and defenses.  Drug metabolism ** O2 is bad : But when it’s high will cause oxidative stress so if you are enforcing O2 into the body it will be

9

Biochemistry

Respiratory system

Aoun batch

These are the molecules that will be affected, mainly the DNA is significant significant one

This figure shows the usage of self generated reactive oxygen species in order to kill infectious body HOCL one of the housekeeping materials in our body and we can re consume or deplete it. Cellular injury occurs through the production of ROS: 1. Superoxide Anion. 2. Hydoxyl Radicals 3. Hydrogen Peroxide There are antioxidant defenses within cells that reduce these species. Free Radical Scavenger Systems: 1. Superoxide dismutase (SOD) 3. Glutathione reductase

2. Glutathione peroxidase 4. Catalase 5. Polyphenols

We can use them to know if certain material increases or decrease oxidative stress. How? We will bring the cell that we want to test and expose it to chemicals and measure the activity of these enzymes. if the activity increases that means the material is very active , if activity decreases this means the material is safe and prevent reactive oxygen species formation.

10

Biochemistry

Respiratory system

Aoun batch

Hydrogen peroxide is less dangerous than reactive oxygen species it’s just toxic molecule that we can deal with by POD enzyme POD enzyme needs glutathione( which converted from reduced to oxidized form) , but in some reactions I need reduced form of glutathione so I have glutathione reductase enzyme convert it to reduced form. The reducing agent in this reaction is NADPH which I can take by glycolysis. This mechanism found in all cells with others mechanisms but in red blood cells we have only this mechanism. In red blood cells only have certain types of enzymes and in normal case the red blood cells are found in place with high oxidative stress so They have one mechanism (glutathione reaction ) that removes oxidative tress from inside the cell so this mechanism destroyed because of deficiency in one of the enzymes inside the cell (usually associated with glucose 6 phosphate dehydrogenase deficiency ) which will lead to Favosm( allergy against oxidants).  Because patient with glucose 6 phosphate deficiency can’t tolerate the oxidants that result from (infections, peans , anti biotics, salicylic acid) the RPC will breakdown.  In G6PD deficiency that in the blood cell will be distracted but other cells will still alive, why? By mal enzyme mechanism which converts malate to pyruvate And produce NADPH for cells to still alive. The other reaction is catalase enzyme reaction which is very simple reaction used to distinguish between bacterial types.

11

Biochemistry

Respiratory system

Aoun batch

Oxidized form

Reduced form

Non-enzymatic Antioxidant compounds: Glutathione ,Vitamin C , Vitamin E, Vitamin A , Uric acid

12

Biochemistry

Respiratory system

Aoun batch

Damage Produced by Reactive Oxygen Species:

Peroxidation of PUFA (poly unsaturated fatty acids)

All biological macromolecules are damaged

CLINICAL SIGNIFICANCE  Diseases will result if there is increase or less concentration ROS (less defense mechanism) 1. Chronic Inflammation (rheumatoid Arthritis, chronic glomerulonephritis) 2. Acute inflammation 3. Respiratory diseases A. bronchopulmonary dysplasia. Newborn O2 exposure B. Adult respiratory distress syndrome (ARDS), Neutrophils recruitment (In some cases we treat these patients with oxygen which lead to . bronchopulmonary displasia) 4. Diseases of the Eye A. Retrolental fibroplasia (retinopathy of prematurity), high O2 Change in the structure of tissues to be more fibrous B. Cataract 13

Biochemistry

Respiratory system

Aoun batch

5. Reperfusion Injury  During ischemia, the activity of xanthine oxidase is increased

6. Atherosclerosis and Myocardial Infarction  Low density lipoproteins (LDL)  Oxidation of this molecule stimulate the immune cells to attack and make foam cells in special mechanism which leads to balque information in blood vessels leads to MI  If there is injury that will lead to activation of oxidative stress 7. Shock-related Injury 8. Skin Diseases 9. Carcinogenesis and Treatment 10.Aging Process 1. 8-oxoguanine. 2. 5-hydroxyuracil. 3. Thymine glycol.

‫في هون رسمات بالسالدز‬ ‫ما عرفت احطها ارجعولها‬

Relieved by nucleotide hydrolase  So the mutation that make change in nucleotide will make bigger problem in the future Q: What will happen during pregnancy and birth? A: Higher activity of antioxidant enzymes Preterm babies have to face ROS stress ٠٤:٠٤ ‫ إلى‬٠٠:٤٤ ‫هون ما قدرت افهم فما عرفت أفرغ ًهبس ارجعوا للريكورد مه‬

14

Biochemistry

Respiratory system

Aoun batch

Proteins, DNA and Lipids will be affected by ROS e.g. Protein-SH, Oxo8dG and lipid peroxidation. Oxygen toxicity can cause damage at multiple levels 1. System wide cellular injury 2. Airway injury 3. Lung parenchymal injury Airway Injury:  Many healthy volunteers will experience substernal heaviness, pleuritic chest pain, cough and dyspnea within 24 hours of breathing pure oxygen.  This is a result of tracheobronchitis and absorptive atelectasis.  Erythema and edema can be observed on bronchoscopy in patients treated with 90% FiO2 for 6 hours.  Concentration of reactive oxygen species in exhaled gas increases after only 1 hour of breathing 28% oxygen, regardless of the presence of underlying lung disease Bronchopulmonary dysplasia or BPD  A disease in neonates following recovery from neonatal RDS.  Characterized by epithelial hyperplasia and squamous metaplasia in the large airways, thickened alveolar walls, and peribronchial and interstitial fibrosis. ‫*موضوع ال‬lipid peroxidation ‫للدكتور سحب عليه فما حطيته‬

Role of Anti-Oxidants • There are two types of anti-oxidants: 1. Preventive anti-oxidants: • Catalase • Glutathione peroxidase, • Ethylene diamine tetra-acetate (EDTA). 2. Chain breaking anti-oxidants • Superoxide dismutase • uric acid • vitamin E. Alpha tocopherol ‫و ختا ًما ٌا ع َْىن تذكزوا أنّ هللا عز و جل خلقىا لىكىن خلٍفته فً اَرض و لىع ّمز هذا الكىن كما ٌحب و‬ ‫ٌزضى و واجب الىقت علٍىا هأل هى دراستىا إلوه اإلحسان فٍها رح ٌخلٍىا األحسه فً مٍذان استخالفىا‬ ‫و تذكزوا‬ َ َ ْ ْ ْ ْ َ ُ ُ َ ُ ‫ع َم ًال ۚ َو ُه َى العَ ِز‬ )‫ىر‬ َ ُ‫سه‬ َ ْ‫الَّذِي َخلَقَ ال َم ْىتَ َوال َحٍَاة ِلٍَ ْبل َى ُك ْم أٌُّ ُك ْم أح‬ ُ ‫ٌز الغف‬ ‫و تذكزوا ان هللا ال ٌضٍع أجز المحسىٍه‬ ‫تم بحمذ هللا‬ 15