Rbc structure and metabolism

description about RBC membrane and its structural peculiarities,how it differs from other cells of our body. How this specialized cell manage homeostasis and function in a well defined manner. This presentation will also help in understanding various RBC storage lesions ,an important aspect of blood banking.

1. DR AKSHAYA TOMAR DEPARTMENT OF IMMUNOHEMATOLOGY AND BLOOD TRANSFUSION AFMC,PUNE RBC STRUCTURE AND METABOLISM

2. INTRODUCTION RBCs are complex ,metabolically active cells using glucose to make ATP and reducing equivalents to ensure flexibility and O2 delivery Life span – 120 days Neither use O2 for extraction of energy nor synthesizes protein Proteomics has identified 2200 separate proteins in RBC that are the product of 5% of all human genes

3. RBC MEMBRANE Erythrocyte membrane that is normal in structure and function is essential for survival of red cell Accounts for the cell's antigenic characteristics Maintains stability and normal discoid shape of cell Preserve cell deformability Retain selective permeability

4. RBC MEMBRANE Semipermeable lipid bilayer supported by a meshlike protein cytoskeleton Proteins and phospholipids are organized asymmeterically . Biochemical composition includes 52% proteins,40% lipids and 8% carbohydrate.

5. MEMBRANE LIPIDS Choline containing, uncharged phospholipids, outer layer: – Phosphatidylcholine (PC) (30%) – Sphingomyelin (SM) (25%) Charged phospholipids, inner layer: – Phosphatidylethanolamine (PE) (28%) – Phosphatidylserine (PS) (14%)- Apoptotic marker of RBC

6. Asymmetric phospholipids distribution is maintained by: –Differential rate of diffusion through membrane bilayer of choline containing phospholipids (PC and SM diffuse slowly) –Charged phospholipids interaction with membrane skeletal protein. –Active transport of amino phospholipids (PC and PE) from outer to inner layer.

7. This asymmetric phospholipid distribution among the bilayer is the result of the function of several energy-dependent and energy-independent  phospholipid  transport proteins.

8. MAINTAINENCE OF ASSYMETRY

9. MEMBRANE PROTEINS Integral proteins –Embedded in membrane via hydrophobic interactions with lipids. Peripheral proteins –Located on cytoplasmic surface of lipid bilayer, constitute membrane skeleton. –Anchored via integral proteins –Responsible for membrane elasticity and stability.

10. INTEGRAL PROTEINS Band 3 Glycophorin Aquaporin

11. BAND 3 •Functions: –Anion transport Exchanges bicarbonate for chloride –Structural: Linkage of lipid bilayer to underlying membrane skeleton. – Interaction with ankyrin and protein 4.2, secondarily through binding to protein 4.1. Important for prevention of surface loss.

12. GLYCOPHORINS Comprise 2% of RBC membrane proteins. – Sialic acid rich glycoproteins (A,B,C) 3 domains: – Cytoplasmic – Transmembrane : single spanning alpha helix –Extracellular: glycosylated

13. GLYCOPHORINS Human RBCs glycophorins are integral membrane proteins rich in sialic acids that carry blood group antigenic determinants and serve as ligands for viruses and parasites

14. AQUAPORINS Aquaporins selectively conduct water molecules in and out of the cell, while preventing the passage of ions and other solutes Allow RBC to remain in osmotic equilibrium with extracellular fluid.

15. Sphingomyelin (SM) Phosphatidylethanolamine (PE)

16. PERIPHERAL MEMBRANE PROTEINS Spectrin Actin Protein 4.1 Pallidin (band 4.2) Ankyrin Adducin Tropomycin Tropomodulin

17. SPECTRIN Flexible, rod like molecule, 100nm length. Responsible for biconcave shape of RBC Two subunits: –Alpha and beta, entwined to form dimers . –Associate head to head to form tetramers End-to-end association of these tetramers with short actin filaments produces the hexagonal complexes observed

18. ACTIN Short, uniform filaments 35nm in length Length modulated by tropomyosin / tropomodulin Spectrintail associated with actin filaments Approx 6 spectrin ends interface with one actin filament, stabilized by protein 4.1

19. ANKYRIN Interacts with band 3 and spectrin to achieve linkage between bilayer and skeleton. Augmented by protein 4.2.

20. OTHER PERIPHERAL PROTEINS Protein 4.1 –Stabilizes actin-spectrin interactions Adducin –Also stabilizes interaction of spectrin with actin . –Influenced by calmodulin (thus can promote spectrin-actin interactions as regulated by intracellular Ca concentration)

21. CHARACTERISTICS OF RBCs DEFORMABILITY : It is controlled by ATP driven membrane cytoskeleton. Loss of ATP leads to decrease in phosphorylation of spectrin . Increase in deposition of membrane calcium. Rigid cells are removed from the circulation.

22. PERMEABILITY : Permeability properties of the RBC membrane and the active RBC cation transport prevent colloid hemolysis and control the volume of RBC. Freely permeable to water and anions; relatively impermeable to cations . When RBC are ATP depleted Ca and Na are allowed to accumulate intracellularly and K and water are lost.

23. METABOLIC PATHWAYS Mainly anaerobic, RBC have to deliver not consume O2. No nucleus/No mitochondria. RBC metabolism may be divided into anaerobic glycolysis and 3 ancillary pathways

24. RBCs contain no mitochondria, so there is no respiratory chain, no citric acid cycle, and no oxidation of fatty acids or ketone bodies. The RBC is highly dependent upon glucose as its energy source. Energy in the form of ATP is obtained ONLY from the glycolytic breakdown of glucose with the production of lactate (anaerobic glycolysis ).

25. RBC METABOLISM Glucose transport through RBC membrane: Glucose is transported through RBC membrane by facilitated diffusion through glucose transporters (GLUT-1).

26. GLYCOLYSIS Importance of glycolysis in red cells: Energy production: It is the only pathway that supplies the red cells with ATP. Reduction of methemoglobin : Glycolysis provides NADH for reduction of metHb by NADH-cytob5 reductase In red cells 2,3 bisphosphoglycerate binds to Hb , decreasing its affinity for O2, and helps its availability to tissues.

27. UTILIZATION OF ATP Phosphorylation of sugars and proteins ATPase driven ion pumps Maintenance of membrane asymmetry Maintenance of red cell shape and deformability using ATP dependent cytoskeleton

28. METH Hb REDUCTASE PATHWAY Maintains Iron in reduced state for effective transport of O2 Protect SH group of Hb and membrane proteins from oxidation

29. LEUBERING RAPOPORT SHUNT The hydrogen ion concentration is the most important physiological modulator

30. LEUBERING RAPOPORT SHUNT Binding of 2,3 DPG to DeoxyHb stabilise the tense state of Hb and favours release of O2

31. LEUBERING RAPOPORT SHUNT Free 2,3 DPG also binds with Band 3 and causes partial detachment of membrane from cytoskeleton allowing lateral movement of membrane structure.

32. PENTOSE PHOSPHATE PATHWAY Production of NADPH – ‘reducing power’ Glutathione is needed in reduced form for: Elimination of peroxide Protection of proteins SH groups This shunt also provide ribose 5 phosphate needed for PRPP ( substrate for adenine nucleotides reqd for continuing ATP synthesis)

33. PENTOSE PHOSPHATE PATHWAY

34. PATHWAYS

35. HEMOGLOBIN OXYGEN DISSOCIATION Dissociation and binding of oxygen by hemoglobin are not directly proportional to pO2. Sigmoid-curve . It permits a considerable amount of O2 to be delivered to the tissues with a small drop in O2 tension.

36. THANK YOU