6. Red Blood Cell Metabolism

Blood Components

55% Plasma, 45% RBCs, <1% WBC’s and platelets

General characteristics of RBCs

• Function

• Abundance in blood

• Organelles

• Function: transport oxygen and contribute to buffering of the blood

• The most abundant cell in blood —> compared to WBCs and platelets

• Has no organelles, as they are lost during differentiation

Red Blood Cell Metabolism General Overview

• _________ RBCs contain no ______________ ____________, thus metabolic pathways are forced to occur in the ___________

• Because they lack ________,the only way RBC’s can generate ATP is through _________, as they convert ________ into ___________

• ATP of RBCs is used for what 3 things?

• RBCs _________ contains enzymes for _____________ and _________ of damage done by __________ _________ _________ , they get these enzymes from the ______ pathway

• Mature RBCs contain no intracellular organelles, thus metabolic pathways are forced to occur in the cytoplasm

• Because they lack organelles ,the only way RBC’s can generate ATP is through glycolysis, as they convert pyruvate into lactate

• ATP of RBCs is used for ion transport, phosphorylation of membrane proteins and priming of glycolysis reactions

• The RBC’s cytosol contains enzymes for prevention and repair of damage done by reactive oxygen species , they get these enzymes from the PPP pathway

Pentose Phosphate Pathway/ Hexose Monophosphate (HMP) Shunt —> Quick review

• What route does it bypass?

• What glycolysis precursor does it start and which one does it end with?

• What electron carrier does PPP generate and what is it used for?

• PPP breaks down what type of biomolecule?

• What are the two phases? Which one generates the electron carrier molecule mentioned before

• Which stage is irreversible

• What molecule does the oxidative phase start and end with, how many electron carriers are formed

• What is the crucial enzyme needed for the oxidative phase, what happens when it is deficient?

• What molecule does the Non-Oxidative phase start and end with?

• Bypasses the first stage of glycolysis

• Starts with glucose-6-phosphate and ends with fructose-6-phosphate

• PPP generates NADPH, which is used in nucleotide biosynthesis, Fatty acid synthesis and cholesterol biosynthesis

• PPP breaks down carbohydrates/sugars

• The two phases are oxidative and non-oxidative; NADPH is generated in the Oxidative phase only

• The irreversible stage is the Oxidative one

• Oxidative phase starts with glucose-6-phosphate and ends with ribulose-5-phosphate; 2 NADPH are produced

• The enzyme is glucose-6-dehydrogenase, if it is deficient it can cause hemolysis/hemolytic anemia because there is no NADPH for glutathione to detoxify ROS

• The non-oxidative phase starts with ribulose-5-phosphate and ends with fructose-6-phosphate

Rapoport- Luebering Shunt —> occurs only in RBC’s

• Steps

• Why is the function of this in metabolism

• What pathway is the initial molecule shunted from

• Because of this shunt, what happens to net ATP produced from glyolysis?

• 1,3-bisphosphoglycerate (1,3BPG) — mutase → 2,3BPG

  —phosphatase→ 3-phosphoglycerate

• The function is to produce 2,3GPG to: moderate oxygen binding, stabilize the deoxy form of hemoglobin and facilitate oxygen release to tissues

• 1,3-BPG is shunted from glycolysis

• Because a step is skipped (would generate 2 ATP), the second stage of glycolysis only generates 2 ATP, which brings the net ATP of glycolysis to 0 when you subtract the 2 ATP from the preparative stage

If RBC’s can only produce ATP from glycolysis, they cannot undergo The e’ Transport system or TCA….. how do they use the NADH produced by glycolysis?

NADH reduces Cytochrome b5, which converts Fe³⁺ (ferric, deoxy) of hemoglobin into Fe²⁺ (ferrous, oxy)

What is the lactate produced by RBC’s used for?

The Cori cycle in the liver turns lactate into glucose through gluconeogenesis

Heme structure: describe groups, order and main structure

Heme is the main ____________ found in the body and forms _____________, ____________ and the ____________

4 Pyrrole rings connected by 4 methylene bridges

8 side chains on pyrrole rings: 4 methyl (M) groups, 2 vinyl (V) groups, 2 propionate (P) groups

Order: MVMVMPPM

Heme is the main porphyrin found in the body and forms hemoglobin, myoglobin and the cytochromes

• Heme synthesis:

  • 3 steps

  • Requirements of the step 1 enzyme

• Allosteric inhibitors

1. glycine + succinyl-CoA — delta aminolaevulinic acid synthase→ delta aminolaevulinic acid (delta-ALA)

   • Condensation reaction where glycine is decarboxylated

   • Enzyme requires pyridoxal phosphate (PLP) —> Pyridoxine (Vit B₆) —> deficiencies cause microcytic hypochromic anemia (RBCs that are smaller and thus have less heme)

   • Allosterically inhibitor: Heme

2. 2 molecules of delta-ALA —delta ALA dehydratase→ porphobilinogen (a pyrrole)

3. 4 pyrrole rings condense to form a chain and then a series of porphyrinogens

   • Deficiencies in the enzymes used in all these steps are all different types of Porphyrias

4. Iron is added

• Heme degradation occurs when _____ reach the end of their life at approximately _____ days and they are _____________ by the cells of the ___________________ __________

Steps of Heme Degradation

• Heme degradation occurs when RBC’s reach the end of their life at approximately 120 days and they are phagocytosed by the cells of the reticuloendothelial system

• Steps of Heme Degradation

  1. Hemoglobin → Heme + Globin

     • Globin —> Amino acids

  2. Heme → Bilirubin

     1. Heme → Biliverdin

        • methylene bridge is broken

        • Catalyzed by heme oxygenase

        • CO₂ and iron released

     2. Biliverdin → Bilirubin

        • Center methylene bridge reduced (double bond to single bond)

        • Catalyzed by Biliverdin reductase

        • NADPH needed from PPP

  3. Bilirubin → Bilirubin-albumin

     • Bilirubin is being carried in the blood by albumin

  4. Bilirubin-Albumin → Bilirubin diglucuronide → bile

     • Conversion to bilirubin-diglucuronide allows excretion by the liver as bile

Sources of Iron

• Iron is obtained from the _____, the daily amount for men and post-menopausal women is ____, while for pre-menopausal women it is _________ due to menstruation

• The average US diet contains _____, but only ______% is normally absorbed

• Are iron deficiencies common or uncommon?

• Iron in meats is in the ______ form, which is _______ absorbed, and iron in plants is in the _______ form, which is _____ absorbed

• Vitamin ___ can ________ the uptake of non-heme iron from the digestive tract

• Iron is obtained from the diet, the daily amount for men and post-menopausal women is 10mg, while for pre-menopausal women it is 15mg due to menstruation

• The average US diet contains 10-50mg, but only 10-15% is normally absorbed

• Are iron deficiencies are common

• Iron in meats is in the heme form, which is readily absorbed, and iron in plants is in the non-heme form, which is not readily absorbed

• Vitamin C can increase the uptake of non-heme iron from the digestive tractF

Iron transport

• Iron is absorbed in the __________ state

• Iron is then ________ to the __________ state by the enzyme ____________

• Iron is carried in the blood as _______ by the protein ______________

• _____ + Apotransferrin= ____________ _________

• Transferrin binds to __________ _________ on the cell surface and is _________, then it is __________ to _____ and released into the __________ by iron transporter __ ( ____ )

• Deficiency in ________ leads to __________ _________ ________ ________

• Once in the cytoplasm, iron is ________ to necessary ________

• Excess iron is ______ back into ______ and binds to __________ for _____-term storage in the _______, _______ and bone __________.

• _______ + Apoferritin = _________ complex

• Iron can be drawn from ________ stores for cells that require iron such as __________ that use it to synthesize ____________

• About __mg of iron is lost per day in feces, urine, sweat, skin and menstruation

• Iron is absorbed in the ferrous (Fe²⁺) state

• Iron is then oxidized to the ferric (Fe³⁺) state by the enzyme ferroxidase (ceruloplasmin)

• Iron is carried in the blood as Fe³⁺ by the protein Apotransferrin

• Iron + Apotransferrin= Transferrin complex

• Transferrin binds to transferrin receptors on the cell surface and is internalized, then it is reduced to Fe²⁺ and released into the cytoplasm by iron transporter I ( DMT-I)

• Deficiency in DMT-I leads to refractory hypochromic microcytic anemia

• Once in the cytoplasm, iron is shunted to necessary enzymes

• Excess iron is oxidized back into Fe³⁺ and binds to Ferritin for long-term storage in the liver, spleen and bone marrow.

• Iron + Apoferritin = Ferritin complex

• Iron can be drawn from ferritin stores for cells that require iron such as reticulocytes that use it to synthesize hemoglobin

• About 1mg of iron is lost per day in feces, urine, sweat, skin and menstruation

Red Blood Cells

• Under a microscope, RBC’s appear as a red ____ with ______ central area

• The shape facilitates _____ _________ across cell membrane

• The shape facilitates RBC’s to ________ across capillaries with small __________ to deliver oxygen to tissues

• They can ______ and ________ to travel restricted spaces

• The _____ determines the viability of RBC’s because they have to pass its small _____ diameter, thus the RBC must be highly _________ to pass through

• Damaged cells become _________ in the _____ and destroyed by ___________

• Under a microscope, RBC’s appear as a red discs with pale central area

• The shape facilitates gas exchange across cell membrane

• The shape facilitates RBC’s to travel across capillaries with small diameter to deliver oxygen to tissues

• They can shrink and expand to travel restricted spaces

• The Spleen determines the viability of RBC’s because they have to pass its small 3 micrometer diameter, thus the RBC must be highly deformable to pass through

• Damaged cells become trapped in the spleen and destroyed by macrophages

Red Blood Cells

• RBC deformability lies in its shape, but also the _________ of the proteins that make up the membrane

• These proteins are on the ___________ side of the membrane

• List the proteins and their function

• Spectrin is the ________ of the membrane and are the ones responsible for altering ____ when the cell is subjected to _________ stress

• _______ RBC’s cannot synthesize new membrane proteins or lipids to use for ___________ detoxification

• Defect in these membrane proteins can result in _________ ________

• When RBC’s lose their deformability, they can ____ in response to stress, get trapped and ________ or be malformed and become a ________

• RBC deformability lies in its shape, but also the organization of the proteins that make up the membrane

• These proteins are on the cytoplasmic side of the membrane

• List the proteins and their function:

  • Spectrin: heterodimer alpha or beta subunits that wound around each other → looks like strings

  • Actin and Band 4:1: bind at the ends of Spectrin dimers and can connect multiple of them

  • Ankyrin and Band 4.2 anchor Spectrin to the membrane

• Spectrin is the cytoskeleton of the membrane and are the ones responsible for altering shape when the cell is subjected to mechanical stress

• Mature RBC’s cannot synthesize new membrane proteins or lipids to use for glutathione detoxification

• Defect in these membrane proteins can result in hemolytic anemia

• When RBC’s lose their deformability, they can lyse in response to stress, get trapped and destroyed or be malformed and become a spherocytes

Hematopoesis

• All cell lineages descended from _____________ _______ cells found in ______ __________

• The population of hematopoietic stem cells is ___ per ____ bone marrow cells

• RBC’s come from the same lineage as ________ and their shared characteristic is that they lack a _________

• All cell lineages descended from pluripotent stem cells found in bone marrow

• The population of hematopoietic stem cells is 1 per 10⁵ bone marrow cells

• RBC’s come from the same lineage as platelets and their shared characteristic is that they lack a nucleus

Hematopoiesis Cytokines

• What cells support the development of progenitor cells?

• How do they support?

• Stromal cells: fibroblasts, endothelial cells, adipocytes and macrophages

• Things done for support:

  • Secrete Growth Factors

    • Growth Factors are recognized by Cytokine receptors and cause receptor aggregation

    • JAK’s phosphorylate cytokine receptor, making it active

    • STAT proteins bind to activated receptor and are phosphorylated to initatiate gene transcription

    • SOCS (silencer) inhibits this mechanism

Erythropoiesis

• RBC production is regulated by the ______ of oxygen delivery to tissues

• In response to reduced tissue oxygenation, the _________ releases hormone called ___________ that stimulates ____________ and replication of ___________ progenitors

• Describe the order of maturation of RBCs

• RBC production is regulated by the demand of oxygen delivery to tissues

• In response to reduced tissue oxygenation, the _________ releases hormone called ___________ that stimulates ____________ and replication of ___________ progenitors

• Describe the order of maturation of RBCs:

  1. Stem cell

  2. Mixed Myeloid Progenitor Cell (CFU-GEMM) → gives rise to granulocytes megakaryotes and monocytes

  3. BFU-E (burst forming unit-erythroid)

  4. CFU-E (colony-forming unit-erythroid)

  5. Pronormoblast → first recognizable RBC precursor

  6. Reticulocyte → forms after 4 rounds of division and still has organelles

  7. RBC → no organelles

Anemia

• When hemoglobin concentration falls below normal values, the patient is classified as _______

• Anemias can be classified by cell sizes: what are the size classes?

• Anemias can also be classified in hemoglobin concentration, what are the concentration classes?

• When hemoglobin concentration falls below normal values, the patient is classified as anemic

• Anemias can be classified by cell sizes: what are the size classes?

  • Normal → Normocytic

  • Small → Microcytic

  • Large → Macrocytic

• Anemias can also be classified in hemoglobin concentration, what are the concentration classes?

  • Normal → Normochromic

  • Low → Hypochromic

• Types of Anemia

• Types of Nutritional Anemias

• Causes of Hemolytic Anemia

Regular Anemias

• Microcytic and Hypochromic → impaired hemoglobin synthesis → iron deficiency, thalassemia, lead poisoning

• Macrocytic, normochromic → Impaired DNA synthesis → B12 deficiency or folic acid deficiency, erythroleukemia

• Normocytic. normochromic → RBC loss → acute bleeding, sickle cell disease

Nutritional Anemias

• Iron deficiency → Smaller (Microcytic) and Paler (Hypochromic)

• Vitamin B12 or Folate deficiency → Macrocytic/Megaloblastic Anemia

  • These nutrients are needed for DNA synthesis

  • Causes bigger and lesser blood cells produced because they cant divide

Hemolytic Anemias are due to deficiency in the following enzymes:

• Pyruvate kinase

• G6P dehydrogenase