Erythrocytes Metabolism

• Erythrocytes metabolism
  

1. Introduction

2. Cytoskeletal Proteins

3. Metabolism and clinical correlation

4 Synthesis of hemoglobin

5. Porphyrias

6. Catabolism of hemoglobin

 

• Introduction
  

Although devoid of nucleus and intracellular

organelles, the red cell:

- squeezed through small capillary beds, buffeted by high shear forces in heart,

- subjected to sequestration,

- static

- substrate deprivation in spleen,

- has a life-span approximating four month

- maintains its shape, pH, ionic equilibrium, protects hemoglobin integrity and function by virtue of its membrane- cytoskeletal structure and its metabolic activity

• Cytoskeletal proteins membrane
  

- Inner surface of the membrane consist 5 principal proteins:

1. Spectrins 2. Actin 3. Ankyrin 4. Bands 4.1

5. Bands 4.2

- Other membrane proteins serve transport functions

1. Non-polar substances enter RBCs by diffusion

2. Polar solutes enter at special transport sites

• Glucose
  
  

- enters the cell by facilitated diffusion mediated by the glucose transporter (bands 4.5) because the glucose transporter can supply much more glucose than is required by the glycolytic pathway

- glucose is not rate limiting in normal red cell

metabolism

Na⁺ and K⁺

- The concentrations Na⁺ and K⁺ within the red cell

are regulted by the Na⁺/K⁺ ATPase pump

- The intracellular conc. of these cations approximate

130 mM of K⁺ and 8mM of Na⁺ and in the plasma about 140 mM of Na⁺ and 4 mM of K⁺

• continued
  

- Passive diffusion of the Na⁺ and K⁺ by the active transport mechanism, the cell must continuously move Na⁺ out and K⁺ in by way of the Na⁺/K⁺ ATPase powered pump

Ca²⁺ and Mg²⁺

- Ca²⁺ and Mg²⁺ con. are regulated by the Ca²⁺/Mg²⁺ ATPase pump.

- Band 3 both links the lipid bilayer to the cytoskeleton and functions as an anion exchanger for chloride-bicarbonate exchange

Glycosylphosphatidylinositol (GPI)

- embedded in the outer leaflet of the bilayer is a phospholipid anchor exposed hydrophilic protein

 

• continued
  

- There are two complement - regulatory proteins

1. Decay accelerating factor (DAF): to protects the cell against complement- induced lysis

2. Membrane inhibitor of reactive lysis (MIRL = CD59): to protects against complement-mediated by binding to C8

- Defect in GPI synthesis, the red cell becomes vulnerable to complement-induced lysis

- Defect in GPI synthesis can be caused by somatic mutation in a totipotent hematopoietic stem cell

• Erythrocytes Metabolism
  

- red cell is dependent on a continuous of glucose for it energy requirement

- is supplied by glycolysis, the pathway begins with facilitated insulin-dependent, uptake glucose and the production of lactate

- the role glycolysis in supporting the energy-requiring housekeeping duties of membrane maintenance and osmotic stability, methemoglobin reduction, regulation of hemoglobin function,

- In normal red cell, 90% of glucose follow the an aerobic pathway to lactate and 10% flows through the aerobic HMPS.

• The function energy in red cell
  

- is required to maintain

* cationic balance. shape and membrane integrity of cells

* prevent oxidative damage

* maintain hemoglobin in its un oxidized functional deoxy-form

* provide 2,3 BPG - a facilitator of hemoglobin function

- The function are made possible through:

1. Generation of energy in the form of ATP

2. Generation of NADPH and reduced glutathione (GSH) in

HMPS

3. The methemoglobin reduction NADH system

4. The Rapoport-Luebering shunt that produces 2,3

BPG

 

 

• Anaerobic glycolysis
  

- The final product is lactate because the latter reaction, by the enzyme lactic dehydrogenase (LDH) converts pyruvat to lactate

- In this reaction only two molecules ATP are produced per mol of glucose consumed

The Hexose Monophosphate Shunt

(aerobic glycolysis)

- The role of the reactions to product NADPH and reduced glutathione (GSH) under condition stress, to protect the cell membrane and cell contents against oxidative damage

- The key enzyme of HMPS is glucose 6-PO4 dehydrogenase (G6PD) in the production of NADPH

- NADPH is use to reduction of oxidized

glutathione (GSSG) to be reduced form (GSH)

- GSH is an essential intracellular reducing agent

that protects cell against oxidant injury

 

Rapoport-Luebering Shunt

- The role of the shunt reaction

* produces 2,3-BPG which is a regulator of hemoglobin oxygenation-deoxygenation and modulates ATP synthesis

 

- The BPG mutase converts 1,3-BPG to 2,3-BPG, while BPG phosphatase removes the phosphate at the 2 position of 2,3-BPG, and return 3-PG to glycolytic pathway has two primary roles:

* when 1,3-BPG converted directly to 3-PG

the high energy phosphate from the 1

position generates ATP

* In the convertion of 1.3-BPG to the lower

energy 2.3-BPG through the action of

BPG mutase, no ATP is form.

• Methemoglobin reduction
  

- is formed under condition the oxidation of heme Fe2⁺ to Fe3⁺

- can not transport O2

- deoxyhemoglobin is essential for transport in O2

- methemoglobin is converted to deoxyhemoglobin

is mediated through the transfer of an electron

from NADH to the iron atom of heme via cyt. B5 and cyt. B5 reductase

- NADH is generated in convertion of G-3P to 1,3-BPG by G 3-P dehydrogenase

• Clinical correlation
  

Defect in red cell metabolism

- changes in lipid content or cholesterol/phospholipid ratios with expansion of the lipid bilayer to cell volume is reflected in changes of red cell morphology frequently in patients with liver diseases

- hereditary defects, manifest by structurally altered or unstable glycolytic enzymes, occur in both the anaerobic and aerobic pathways.

- produce varying degrees of shortened red cell life-span - congenital jaundice, splenomegaly and cholithiasis. The most common hereditary enzyme defect of red cell are PK and G6PD deficiencies

• G6PD deficiency
  

- result s in decreased NADPH synthesis with inability to maintain GSH levels the oxidants damage the red cell membrane, oxidize heme to methemoglobin, and denature globin which precipitates on the membrane as Heinz bodies

- the membrane changes reduce deformability, increase cation leakage, and result in a fragile cell

hemolysis with hemoglobinuria

- Methemoglobinemia

- due to a deficiency of NADH-cytochrome b5

reductase (methemoglobin reductase)

• Hemoglobin
  

- hemoglobin is protein constitutes 90% of the dry weigh of erythrocytes serves as oxygen transporting

- also plays a vital role in the transport of carbon dioxide and hydrogen ion

- the capacity of hemoglobin to bind oxygen depend on the presence of non polypeptide unit a heme group

- is composed by heme and globin

- globin is polypeptides synthesized in erythropoitic

cells

- heme, a cyclic tetrapyrrole that imparts a red color

• Synthesis of heme
  

- is synthesized in living cell , most mammalia cell with the exception of nature erythrocytes, from from succinyl-CoA & glicin

- succinyl-CoA is dirived from reaction:

1. The citric acid cycle in mitochondria

2. Methyl malonyl to succinyl-CoA by coenzyme deoxyadenosylcobalamin (active form vit. B12)

- in the reaction to "active" glicin, is pyridoxal phosphate necessary

- there are 7 enzymes for heme synthesis of heme

1. ALA synthase (d-amino levulinic acid synthethase)

- is key regulatory enzyme in heme biosynthesis, act as a negative regulator of the synthesis of ALA synthase

- to be the role-controlling enzyme in porphyrin biosynthesis in mammalia liver

- the reaction by this enzyme occur in the mitichondria

- many xenobiotics, when administrated to humans

hepatic ALA synthase (xenobiotics is metabolized by a system in liver that utilizes a specific hemoprotein: cytochrome P-450)

- during the metabolism of xenobiotics utilization of heme in turn diminishes the intracellular heme concentration ALA-synthase heme synyhesis

- the factor affect the induction ALA-synthase in the liver:

a. Glucose: prevent the induction of ALA-sinthase

b. Iron : in chelated form exerts a synergistic

effect on induction of hepatic ALA-synthase

c. Steroid : permissive role in the drug-mediated

derepression of ALA-synthase in vivo

d. Hematin: prevent the drug-mediated derepression

of ALA-synthase in vivo

2. ALA-dehytratase (ALA-dehydrase)

- is a Zn -containing enzymes and is senzitive to inhibition by lead, because lead combine with SH groups in enzyme ALA-dehydrase

- the reaction occurs in the cytosol

3. Uroporohyrinogen I synthase (porphobilinogen deaminase

- the reaction occurs in the cytosol

4. Uroporphyrinogen III cosynthase

- the reaction occurs in cytosol

5. Uroporphirinogen decarboxylase

- the reaction occurs in cytosol

6. Coproporphirinogen oxidase

- the reaction occurs in mitochondria

7. Protoporphyrinogen oxidase

- the reaction occurs in mitochondria

8. Ferrochelatase = heme synthase

- the reaction occurs in mitochondria, can be inhibited by lead, because lead combine with SH-group in ferrochelatase

- regulation of heme synthesis occurs at ALA-synthase by a repression-dereprission mechanism mediated by heme and its hypothetical aporepressor

- lesions of enzymes 2 - 8 cause the porphyrias

• Pophyrias:
  - group of inborn errors of metabolism due to
  mutation in genes direcly the synthesis of the
  enzymes involved in the biosynthesis of heme
  - in general, the porphyrias describe are inherited in
  an autosomal manner
  - can be classified on the basis the organs or cells
  that are most affected
  - these are generally organs or cells in synthesis of
  heme is particularly active:
  * the bone marrow: synthesis hemoglobin
  * liver is active in the synthesis of another hemo-
  protein: cytochrome P-450 classification
  of the porphyrias is to designate as erythropoietic,
  hepatic, and erythrohepatic
  
• Globin synthesis
  - globin polipeptides are synthesized in erythropoietic cells, defects in globin chain synthesis in thalassemias
  

Catabolismm of hemoglobin

- under physiologic condition in human adult, 1-2 X 10 degree 8 erythrocyte are detroyed per hour

in 1 day turns over approximately 6 gram of hemoglobin

- globin, may be reutilized oer in the form of its constituent amino acids

- iron of heme enters the iron pool, the iron-free porphyrin is degraded mainly in the reticuloendothelial cells of the liver, spleen etc.

• + NADPH + NADPH
  Heme (Fe2⁺) hemin (Fe³⁺) + O2
  - NADP - NADP
  Fe3+ (reutilized) + CO(exhaled) + biliverdin IX-a
  
  
  + NADPH
  - NADP
  yellow pigment bilirubin IX-a
  

- the further metabolism of bilirubin occurs in primarily in liver and then divided into 3 process

1. Uptake of bilirubin by liver parenchymal cells

2. Conjugation of bilirubin in the smooth e. r.

3. Secretion of conjugated bilirubin into bile

• 1. Uptake bilirubin by liver parenchymal cell.
  

- bilirubin sparingly soluble in plasma, in plasma it is protein-bound (albumin)

- Each mol. albumin have one high-affinity site and one low-affinity site for bilirubin

- such antibiotic and othe drugs compete with bilirubin for the high-affinity binding site on albumin these compounds can displace bilirubin from albumin significant clinicalseffect

2 In the liver

- bilirubin are removed from the albumin and take up at the sinusoidal surface of the hepatocytes

• 3. Conjugation in the liver
  

- bilirubin is converted to a water soluble form that can be secreted into the bile

- in the smooth endoplasmic reticulum, bilirubin is excreted in the bile in the form of a bilirubin diglucuronide which is catalysis by UDP-glucuronyltransferase

- UDP-glucuronyltransferase can be induced by phenobarbital

- Hyperbilirubinemia causes jaudice

Reference: lecture from:

Wiryatun Lestariana

Biochemistry Department

Fac. of Medicine GMU

Yogyakarta