1. WBCs Metabolism
By Dr. Samar Kassim

2. List the major metabolic pathways in neutrophils.
Understand the defensive role of WBCs.
Define the free radicals.
Know that ROS may be good, may be bad.
List the reactive oxygen species (ROS).

3. 6. Understand the oxygen burst. 7
6. Understand the oxygen burst. 7. Understand the function of different enzymes involved in production and scavenging of ROS. 8. Understand the regulation of proteinase activity. 9. Know how anti-proteinase deficiency cause disease.

4. Neutrophils Have An Active Metabolism
Major Biochemical Features of Neutrophils:
Active aerobic glycolysis
Active HMP
Moderately active oxidative phosphorylation (because mitochondria are relatively few)
High content of lysosomal enzymes
Unique enzymes as myeloperoxidase and NADPH oxidase

5. Acute Inflammatory Response
Neutrophils Contribute in
Acute Inflammatory Response

6. Acute Inflammatory Response
Increase of vascular permeability.
Entry of activated neutrophils into the tissues.
Activation of platelets.
Spontaneous subsidence (resolution) if the invading microorganisms have been dealt with.
One of the mechanisms of dealing with microorganisms is production of reactive oxygen species (ROS).

7. Reactive Oxygen Species ROS Trigger Inflammation

8. Reactive Oxygen Species ROS
Powerful oxidants produced during the course of metabolism,
Produced by blood cells and most other cells of the body.
Superoxide (O2—∙)
Hydrogen peroxide (H2O2)
Peroxyl radicals (ROO•)
Hydroxyl radicals (OH•) & OH-

9. Atoms or groups of atoms that have an Unpaired Electron
Free Radicals (ROS)
Atoms or groups of atoms that have an Unpaired Electron

10. ROS Play Important Role in Cellular Injury,
e.g., resulting from administration of various toxic chemicals or ischemia, some of which can result in cell death.
Several reactions are involved in their production and disposal.

11. Reaction 1 Superoxide (O2—∙) In RBCs: from Auto-oxidation of
Hb -> Met Hb
(about 3% daily).
In other tissues: Formed by cytochrome p450
and xanthine oxidase

12. Reaction 2 Respiratory Burst
Stimulated in neutrophils by contact with bacteria
They produce superoxide in a reaction catalyzed by NADPH oxidase
NADPH + 2O2 ↔ NADP+ + 2O2− + H+

13. Reaction 2 Respiratory Burst
The NADPH is generated mainly by HMP whose activity increases markedly during phagocytosis
NADPH + 2O2 ↔ NADP+ + 2O2− + H+

14. the rate of this same reaction is speeded up by
Dismutation
Spontaneous OR
the rate of this same reaction is speeded up by

15. Many Fates for H2O2 Catalase (R4) Myeloperoxidase (R5)
Glutathione peroxidase (R6)
The Fenton reaction (R7)
Haber-Weiss reaction (R8)

16. Reaction 4
Catalase

17. H2O2 and Halides produce Hypochlorus acid
Reaction 5
Myeloperoxidase
In Neutrophils
H2O2 and Halides produce Hypochlorus acid

18. Glutathione peroxidase (Se)
Reaction 6
Glutathione peroxidase (Se) Se

19. Reaction 7
The Fenton reaction

20. Reaction 8
Haber-Weiss reaction

21. Reaction 8
O2—∙ and Fe3+ are the substrates in the iron-catalyzed Haber-Weiss reaction, which also produces OH• and OH–

22. Reaction 8
O2—∙ can release iron ions from ferritin

23. Reaction 8
production of OH• may be one of the mechanisms involved in tissue injury due to iron overload in hemochromatosis

24. Any Questions

25. ROS Stress
Superoxide (O2—∙)
Hydrogen peroxide (H2O2)
Peroxyl radicals (ROO•)
Hydroxyl radicals (OH•)
Chemical compounds and reactions capable of generating potential toxic oxygen species can be referred to as
pro-oxidants.

26. Anti-oxidants
Enzymes: SOD, Catalase, GSH peroxidase
Non enzymes: NADPH, GSH, ascorbic acid, and vitamin E
Compounds and reactions disposing of these Toxic Oxidant species, scavenging them, suppressing their formation, or opposing their actions.

27. In a normal cell, there is an appropriate
pro-oxidant/antioxidant balance.
However, this balance can be shifted toward the pro-oxidants in the following conditions:
Increased production of oxygen species (e.g., following ingestion of certain chemicals or drugs).
The state of diminished levels of antioxidants by inactivation of enzymes involved in disposal of oxygen species.
This imbalance is called “oxidative stress” and can result in serious cell damage if the stress is massive or prolonged.

28. Oxidative stress
Imbalance between
production of toxic oxygen species & activity of different anti-oxidants

29. Neutrophil Reaction to OS
RESPIRATOY BURST

30. RESPIRATOY BURST
When neutrophils and other phagocytic cells engulf bacteria, they exhibit a rapid increase in oxygen consumption known as the respiratory burst.
This is the rapid utilization of oxygen (following a lag of 15–60 seconds) and production of large amounts of reactive oxygen derivatives, such as O2—∙ , H2O2, OH•, and OCl− (hypochlorite ion). Some of these products are potent microbicidal agents.

31. RESPIRATOY BURST
Neutrophils Contain Myeloperoxidase, Which Catalyzes the Production of Chlorinated Oxidants

32. RESPIRATOY BURST
The enzyme myeloperoxidase is present in large amounts in neutrophil granules.

33. RESPIRATOY BURST
It is responsible for the green color of pus, can act on H2O2 to produce hypohalous acids (hypochlorite)

34. Neutrophil Reaction to OS
HOCl, the active ingredient of household liquid bleach, is a powerful oxidant and is highly microbicidal.

35. Neutrophil Reaction to OS
When the Cl− is applied to normal tissues, its potential for causing damage is diminished.
Cl− reacts with some amines to produce chloramines, which are also oxidants, though less powerful than HOCl.

36. disposed of by the action of glutathione peroxidase or catalase.
Any superoxide that enters the cytosol of the phagocytic cell is converted to H2O2 by the action of superoxide dismutase,
In turn, H2O2 is used by myeloperoxidase OR
disposed of by the action of
glutathione peroxidase or catalase.

37. Any Questions

38. Reaction 2 Respiratory Burst
Stimulated in neutrophils by contact with bacteria
They produce superoxide in a reaction catalyzed by NADPH oxidase
NADPH + 2O2 ↔ NADP+ + 2O2− + H+

39. Reaction 2 Respiratory Burst
The NADPH is generated mainly by HMP whose activity increases markedly during phagocytosis
NADPH + 2O2 ↔ NADP+ + 2O2− + H+

40. Mutation in NADPH Oxidase

41. Mutation in NADPH Oxidase
Mutations in the Genes for Components of the NADPH Oxidase System Cause Chronic Granulomatous Disease

42. Neutrophils Have An Active Metabolism
Major Biochemical Features of Neutrophils:
Active aerobic glycolysis
Active HMP
Moderately active oxidative phosphorylation (because mitochondria are relatively few)
High content of lysosomal enzymes
Unique enzymes as myeloperoxidase and NADPH oxidase

43. Proteinases of Neutrophils
Proteinases of Neutrophils help getting rid of invading organisms by hydrolyzing elastin, various types of collagens, and other proteins.

44. Proteinases of Neutrophils
If uncontrolled, they can Cause Serious Tissue Damage.
Most of these proteinases are lysosomal enzymes and exist mainly as inactive precursors in normal neutrophils.

45. Proteinases of Neutrophils
The activities of elastase and other proteinases are normally kept in check by a number of anti-proteinases present in plasma and the extracellular fluid.

46. Proteinases of Neutrophils
Each of them can combine —usually forming a non-covalent complex— with one or more specific proteinases and thus cause inhibition.

47. Proteinases of Neutrophils
Genetic deficiency of α1-antiproteinase inhibitor (α1-antitrypsin) permits elastase to act unopposed and digest pulmonary tissue, thereby participating in the causation of emphysema

48. List the major metabolic pathways in neutrophils.
Understand the defensive role of WBCs.
Define the free radicals.
Know that ROS may be good, may be bad.
List the reactive oxygen species (ROS).

49. 6. Understand the oxygen burst. 7
6. Understand the oxygen burst. 7. Understand the function of different enzymes involved in production and scavenging of ROS. 8. Understand the regulation of proteinase activity. 9. Know how anti-proteinase deficiency cause disease.

50. Thx