MEH session 5 Flashcards
Where does haemopoiesis occur?
• In an early embryo, begins in the vasculature of the yolk sac
• Week 5-8 gestation, shifts to the embryonic liver
• After birth, sole site of haemopoiesis is in the bone marrow. This is extensive throughout the skeleton in an infant
• There is more limited distribution in adulthood. Main sites:
◦ Sternum
◦ Skull
◦ Ribs
◦ Vertebrae
From which cells do all blood cells originate from?
Haemopoietic stem cells residing in bone marrow which have the unique ability to give rise to all of the different mature blood cell types.
What is haematopoiesis controlled by?
Hormones or cytokines determine which blood cells develop from the haemopoietic stem cells.
What is the reticuloendothelial system?
Network of phagocytic cells throughout the body which is part of the larger immune system.
Function:
- removal of dead/damaged cells
- identify and destroy foreign antigens in blood and tissues
Cells:
- monocytes in blood
- different types of macrophages eg. Kupffer cells, tissue histiocytes, microglial cells in CNS
Which organ has a particularly prominent role in the reticuloendothelial system?
Spleen- all blood passes through the spleen so the reticuloendothelial cells in the spleen are important in filtering blood to remove deformed and old cells from the circulation particularly erythrocytes
Liver also has a role
Why is it important that white blood cell count and platelet count is considered in addition to red blood cell count and haemoglobin count in a suspected anaemia?
To rule out pancytopenia
What is the function of red blood cells?
- Carry haemoglobin
- Maintain haemoglobin in its reduced ferrous state
- Generate ATP to maintain osmotic equilibrium
- Maintain osmotic equilibrium to maintain cell structure
In order to deliver oxygen to tissues (and transport CO2)
What is the purpose of the biconcave structure of red blood cells?
- Optimises laminar flow properties of blood in large vessels
- Allows them to deform and squeeze through small capillaries
- Increases surface area for oxygen exchange
What causes the shape of the red blood cell to change?
Changes in the components of the cell membrane (congenital or acquired)
What is the function of the globin chains in haemoglobin?
- protect haem molecule from oxidation
- confer solubility
- permits variation in oxygen affinity
How long do red blood cells live for?
120 days - therefore, erythropoiesis must be a continual process.
What happens to the haemoglobin removed from senescent erythroyctes?
The haemoglobin removed from senescent erythrocytes is recycled by the spleen with the:
• globin portion being degraded to its constitutive amino acids
• haem portion metabolised to bilrubin which is conjugated in the liver and secreted in bile
What does excess red cell destruction (eg. haemolytic anaemia) and hence excess haemoglobin catabolism present as?
Jaundice
How is bilirubin metabolised and excreted?
- Bilirubin is conjugated by the liver
- Secreted in bile
- Bacteria in the colon deconjugate and metabolise the bilirubin not colourless urobillinogen
- Urobillinogen is oxidised to form urobilin and stercobilin (these are responsible for the brown colour of stool)
- A small amount of the urobulinogen is reabsorbed and processed by the kidneys which gives urine its yellow colour
What are the two main metabolic pathways in red blood cells?
1) glycolysis
Glucose metabolised to lactate
ATP generated
2) pentose phosphate pathway
Glucose-6-phosphate metabolised
Generates NADPH
Which organ is important in regulating red blood cell synthesis?
Erythropoietin is produced by interstitial fibroblasts in the kidney and its production is under negative feedback. Erythropoietin is an essential hormone for red blood cell production, its primary effect is on red blood cell progenitors in the bone marrow promoting their survival.
Erythropoietin production increases in response to a decrease in the oxygen level in the bloodstream. Reduced pO2 is detected in interstitial peritubular cells in kidney.
Explain the significance of the reticulocyte count
provides a good diagnostic estimate of the amount of erythropoiesis occurring in a patient’s bone marrow.
How does Glucose-6-phosphate dehydrogenase deficiency affect red blood cells?
this enzyme catalyses the first step in the pentose phosphate pathway
Mature blood cells lack nuclei so they are unable to replace damaged proteins by re-synthesis making them particularly susceptible to oxidative damage in diseases such as glucose-6-phosphate dehydrogenase deficiency
How does pyruvate kinase deficiency affect red blood cells?
this enzyme catalyses the last step of glycolysis
Mature red blood cells have a lack of mitochondria and therefore a reliance on glycolysis for energy production.
What happens in hereditary spherocytosis?
Red blood cells lose their biconcave shape due to gene mutations in these membrane associated proteins.
What is iron required for?
The function of many enzymes and proteins such as:
• Haemoglobin
• Cytochromes in the electron transport chan
• Catalase involved in the protection against oxidative stress
Why is free iron toxic to cells?
It acts as a catalyst in the formation of free radicals from reactive oxygen species.
How is iron lost from the body?
There is no method of excretion. Approximately 1-2mg is lost from the skin and GI mucosa
Where and how is iron stored and how can this be tested for?
Iron is stored in two forms:
• Ferritin - protein-iron complex which can be incorporated by phagolysosomes to from haemosiderin granules (can have a biochemical blood test for ferritin)
• Haemosiderin - insoluble derivative of ferritin in macrophages (stain tissue and view by microscopy for haemosiderin)
All cells have the ability to sequester iron as either ferritin or haemosiderin. The highest concentrations of stored iron are in the liver, spleen and bone marrow.
Where is most available iron found?
In haemoglobin
On a daily basis, does most of the iron come from the diet?
No, only a small fraction of total iron requirement is gained from the diet.
Most of the iron requirement comes from the recycling of old red blood cells taken up by macrophages in the reticuloendothelial system and is returned to the storage pool.
What food sources are good sources of iron?
Haem sources (animal sources) are more readily absorbed than inorganic iron which consists of both ferric (Fe3+) and ferrous iron (Fe2+). Ferric iron must first be reduced to the ferrous form before it is absorbed.
Where is iron absorbed?
In the duodenum and upper jejunum
In what form must iron be to be absorbed in the GI tract?
Ferrous (Fe2+)
Ferric iron is reduced to ferrous iron by duodenal cytochrome B reductase (DcytB) before uptake by DMT1.
The mechanism by which haem iron is absorbed remains unclear but once in the enterocyte haem is degraded to release ferric iron.
How is iron stored in enterocytes?
Ferritin- this is a protein-iron complex
How does iron leave the enterocytes to be absorbed into the bloodstream?
Ferroportin
Once in the blood, how is iron transported?
It is bound to the transport protein transferrin.
How does the foetus absorb iron
Fetal enterocytes have receptors for Lactoferrin –primary source of iron in infants
How is iron taken up by cells from the bloodstream?
Binding of iron-transferrin complex to transferrin receptor. Transferrin is a plasma glycoprotein.
Erythroid cells contain the highest number of transferrin receptors.
How is absorption of iron regulated?
- regulation of transporters eg. Transferrin
- expression of receptors eg. HFE and transferrin receptor
- hepcidin and cytokines
- crosstalk between the epithelial cells and other cells like macrophages
How does hepcidin regulate the absorption of iron?
- Hepcidin is produced by the liver.
- It binds to ferroportin and results in its degradation.
- This prevents iron from leaving the enterocytes and from stores in macrophages
- Hepcidin inhibits transcription of the DMT1 gene. This transporter is located on the apical surface of enterocytes and facilitates the uptake of ferrous iron.
If iron levels in the blood are high, will hepcidin production by the liver be high or low?
High
When there is a high rate of erythropoiesis, is hepcidin production increased or decreased?
Decreased
What should a clinician do if iron deficiency is found?
It is a symptom, not a diagnosis.
Deficiency can result from:
• Insufficient intake
• Poor absorption
• Increased use due to physiological reasons eg. Pregnancy
• Pathological reasons eg. Excessive bleeding
What are the symptoms of iron deficency anaemia?
Tiredness
Reduced oxygen carrying capacity- pallor and exercise intolerance
Cardiac symptoms - angina, palpitations, development of heart failure
What are the signs of iron deficiency anaemia?
Pallor
Tachycardia
Increased respiratory rate
Epithelial changes - large painful swollen tongue, spooning of nails
What are the blood film features observed in iron deficiency?
Hypochromic - low haemoglobin content
Microcytic - small red blood cells, low mean cell volume
Anisopoikilocytosis - change in size and shape eg. Pencil cells and target cells
What would biochemical tests of blood show in an iron deficient patient?
Low haemoglobin levels
Low reticulocyte haemoglobin content (CHR) - but this is also low in patients with thalassaemia
Low ferritin
A patient has a low reticulocyte haemoglobin content (CHR). Do they definitely have iron deficiency?
No.
It is low in thalassaemias
A patient has a high ferritin count. Does this mean they do not have iron deficiency?
No
It is low in inflammatory responses.
Why is excess iron dangerous?
Iron concentration can get so high that it exceeds binding capacity of transferrin.
Free Fe2+ acts as a catalyst for free radical formation. (Hydroxyl and lipid radicals)
This damaged lipid membranes, DNA and proteins
Excess iron is deposited in tissues as haemosiderin causing organ damage
What is haemochromatosis?
Disorder of iron excess resulting in end organ damage due to iron deposition.
Causes liver cirrhosis, diabetes mellitus, hypogonadism, cardiomyopathy, arthropathy, skin pigmentation
What are the two types of haemochromatosis?
Hereditary haemochromatosis
Transfusion associated haemosiderosis
What happens in hereditary haemochromatosis?
Excessive absorption of dietary iron.
HFE protein binds to the transferrin receptor, reducing affinity for iron-bound transferrin. Therefore, defects in this protein result in greater cellular uptake of iron.
Mutations in the gene coding for which proteins could potentially cause hereditary spherocytosis?
spectrin band 3 erythrocyte membrane protein band 4.2 Ankyrin
These are components of the red cell membrane cytoskeleton network which play a critical role in determining cell shape and deformability. Therefore, mutations produce spherocytes and these deformed cells are destroyed by the spleen resulting in disease
Approximately when does the switch from fetal to adult haemoglobin occur?
3-6 months of age
Which breakdown product of haem is responsible for the yellow discolouration often seen around a bruise
Bilirubin is yellow in colour and is a product of the normal catabolic pathway that breaks down haem. Bilirubin is excreted in bile and urine, and elevated levels may indicate disease. Bilirubin is responsible for the yellow color of bruises and the yellow discoloration observed in jaundice. Subsequent breakdown products such as stercobilin, cause the brown color of faeces and another breakdown product, urobilin, contributes to the straw-yellow colour of urine.
Iron is mainly absorbed from which part of the GI tract?
Duodenum and upper jejunum
What facilitates uptake of ferrous iron from the intestinal lumen into enterocytes?
DMT1 is located on the apical membrane of enterocytes where it binds ferrous iron (Fe2+) in the intestinal lumen and facilitates its transport into the cell.
What inhibits the release of iron from enterocytes and macrophages?
Hepcidin
It binds to ferroportin located on the basolateral membrane of enterocytes. Inhibition of ferroportin by hepcidin prevents iron from being exported into the bloodstream thereby reducing dietary iron absorption. Iron release from macrophages is also reduced by hepcidin inhibition of ferroportin.
Which components of a meal can adversely affect the absorption of iron?
The tannins present in both tea and coffee can adversely affect iron availability.
Which form of non-heme dietary iron is absorbed by intestinal enterocytes?
Fe2+
Ferrous iron
Name a treatment option for a patient with severe hereditary haemochromatosis.
Therapeutic phlebotomy to remove excess iron
Define anaemia.
A haemoglobin concentration lower than the normal range.
Why might anaemia develop?
Bone marrow:
- abnormal erythropoiesis
- abnormal haemoglobin synthesis
Peripheral red blood cells
- abnormal function
- abnormal structure
- abnormal metabolism
Removal
- abnormal function of reticuloendothelial system
- excessive bleeding
Why do people with chronic kidney disease develop anaemia?
Abnormal erythropoiesis
Production of erythropoietin is insufficient to stimulate normal levels of erythropoiesis
Why might a patient undergoing chemotherapy develop anaemia?
Abnormal erythropoiesis
Exposure of bone marrow to this leads to ‘empty bone marrow’ as it is unable to respond to stimuli from erythropoietin
This leads to aplastic anaemia - inability of haematopoietic stem cells to generate mature blood cells
Why might someone who is infected with parvovirus develop anaemia?
Exposure of bone marrow to this virus leads to empty bone marrow as it is unable to respond to stimuli from erythropoietin
This leads to aplastic anaemia.
In what types of conditions is anaemia of chronic disease seen?
Chronic inflammatory conditions such as:
- rheumatoid arthritis
- chronic infections
- malignancy
What happens in anaemia of chronic disease?
- –>Increased activity of macrophages in these underlying conditions - iron is stored here and is not released for use in bone marrow
- –>Reduced lifespan of red blood cells
- –>Marrow shows a lack of response to erythropoietin
- –>Chronic release of cytokines such as IL-6 increase the production of hepcidin by the liver resulting in less iron absorption by the bloodstream
Is anaemia of chronic disease microcytic, microcytic or normocytic?
Any
What are myelodysplastic syndromes and how do they cause anaemia?
Production of abnormal clones of marrow stem cells.
Red cells are
- defective and large (macrocytic anaemia)
- prematurely destroyed by RES
How is anaemia caused by myelodysplastic syndromes treated?
Chronic transfusions of red cells
Anaemia can be caused due to deficiencies in the building blocks for DNA synthesis. What causes this?
Vitamin B12 deficiency
Vitamin B9 folate deficiency
What happens to red blood cells in a vitamin B12 or vitamin B9 deficiency?
- Inability of red blood cell precursor cells to synthesise DNA and therefore divide
- Nuclear maturation and cell division lag behind cytoplasm development so large partial replicated red blood cell precursors are released into the blood stream with inappropriately large nuclei and open chromatin
Is anaemia caused by deficiencies in the building blocks for DNA synthesis microcytic, normocytic or macrocytic?
Macrocytic
Where is vitamin B12 obtained from?
Vitamin B12 can only be obtained from food of animal origin so it is essential that people on a vegan diet eat foods fortified with vitamin B12 or take supplements
How is vitamin B12 absorbed?
B12 combines with intrinsic factor produced by parietal cells of the stomach.
IF-B12 complex binds in the ileum, leading to absorption of B12 and destruction of IF
In portal blood, B12 is bound to plasma protein transobalamin which delivers B12 to the bone marrow and other tissues.
What is pernicious anaemia?
An autoimmune disease affecting gastric parties also cells causing a lack of intrinsic factor
Therefore vitamin b12 cannot be absorbed
Why might someone have a B12 deficiency?
- vegan diet
- pernicious anaemia
- damage to iliac cells
- congenital
How is folate obtained?
Folate is present in a variety of animal and vegetable food sources. It is particularly abundant in green leafy vegetables. A typical Western diet usually contains sufficient folate to meet demands.
Why might someone have a folate deficiency?
- increased demands in pregnancy and lactation
- alcoholics may become deficient due to inadequate intake and damage to intestinal cells
- diseases that affect duodenum and jejunum
- certain drugs
However, we usually have high stores of folate in body
In a patient with vitamin B12 deficiency, apart from anaemia, what other symptoms may be present?
Neurological disease
- focal demyelination affecting the spinal cord, peripheral nerves and optic nerves
- depression
- dementia
Why are the symptoms of anaemia usually mild in people with sickle cell disease?
HbS gives up haemoglobin readily in comparison to normal haemoglobin
What are common triggers of sickle cell crisis?
Cold weather
Infection
Exertion
What types of anaemia are
Microcytic
And
Hypochromic
Iron deficiency
Thalassaemia
Why might someone with thalassaemia get iron overload?
Excessive absorption of iron due to ineffective haematopoiesis
Blood transfusions to treat
Why might a pyruvate kinase deficiency cause anaemia?
- Pyruvate kinase catalyses the final step in glycolysis transferring the phosphate from phosphenol pyruvate to ADP to form ATP
- Since red blood cells lack mitochondria, pyruvate kinase deficiency blocks their only metabolic pathway which can supply ATP for cellular processes
- The sodium-potassium ATPase pump stops working without ATP to provide the energy and the red cells lose K+ to plasma
- Water moves down its concentration gradient out of cells causing them to shrink and haemolytic anaemia
Why might G6PDH deficiency cause anaemia?
- Pyruvate kinase catalyses the final step in glycolysis transferring the phosphate from phosphenol pyruvate to ADP to form ATP
- Since red blood cells lack mitochondria, pyruvate kinase deficiency blocks their only metabolic pathway which can supply ATP for cellular processes
- The sodium-potassium ATPase pump stops working without ATP to provide the energy and the red cells lose K+ to plasma
- Water moves down its concentration gradient out of cells causing them to shrink and haemolytic anaemia
A patient has increased reticulocytes, raised bilirubin, and LDH in their blood. What type of anaemia is this?
Haemolytic
What are the common causes of microcytic anaemia?
Thalassaemia Anaemia of chronic disease (ACD) - can also be normocytic Iron deficiency anaemia Lead poisoning Sideroblastic anaemia
What are the common causes of normocytic anaemia?
Sickle cell anaemia G6PD deficiency B6 deficiency B2 deficiency Anaemia of chronic disease (can also be microcytic)
What are the common causes of macrocytic anaemia?
B12 or folate deficiency causing megoblastic anaemia
What are the common causes of a haemolytic anaemia?
Red blood cell membrane abnormalities
Eg. Hereditary spherocytosis
Defects in the genes of proteins in red blood cell membrane
Idiopathic autoimmune haemolytic anaemia
An immunoglobulin binds to proteins on the red blood cell membrane and destroys them
Red blood cell metabolism defects
Eg. Pyruvate kinase deficiency, G6PHD deficiency
How can anaemia of chronic disease be distinguished from iron deficiency?
They are both microcytic and hypochromic because in anaemia of chronic disease, iron is sequestered as stores in macrophages.
To differentiate, in anaemia of chronic disease:
- ferritin is raised (acute phase protein)
- C-reactive protein is raised
- Other symptoms of chronic inflammatory diseases are present
How would the concentration of 2,3-biphosphoglycerate change in response to anaemia?
It would increase in order to shift the haemoglobin oxygen dissociation curve to the right so haemoglobin would give oxygen more readily to tissues.
State some symptoms of anaemia.
Shortness of breath Palpitations Headaches Pain, discomfort or tiredness in the legs that occurs during walking and is relieved by rest (medical term = claudication) Sensation of chest pain, pressure, or squeezing (medical term = angina) Weakness Lethargy Confusion
State some signs of anaemia.
Pallor
Tachycardia
Systolic flow murmur
What is haemoglobinaemia a sign of?
Intravascular haemolysis
Haemoglobinaemia = excess haemoglobin in the blood. If the normal reticuloendothelial pathway for removal of red blood cells is overwhelmed or haemolysis is very severe (e.g. due to incompatible blood transfusion), a direct breakdown of red blood cells rusults in release of haemoglobin into the circulation. Normally free haemoglobin in the blood is picked up by the protein haptoglobin (produced by liver), but there is a limited amount of this protein and it can become saturated very quickly.
What is the underlying pathology of hereditary spherocytosis?
Increased red cell rigidity
Mutations resulting in loss of function of proetins (ankyrin, spectrin, band 3, Protein 4.2) involved in vertical interactions between the cytoskeleton and the lipid bilayer of the plasma membrane result in hereditary spherocytosis. Disruption of this interaction causes a local disconnection of the cytoskeleton and membrane resulting in vesiculation of unsupported membrane components and progressive reduction in membrane surface area. This causes the red cells to adopt a spherocyte shape which is more rigid compared to the normal biconcave shape and therefore less deformable. Spherocytes haemolyze as they pass through the small diameter blood vessels of the spleen. The poor deformability of spherocytes only appears to be a problem in the spleen since these cells have a nearly normal lifespan following splenectomy.
Pyruvate kinase deficiency is diagnosed in a 4 year old girl. She develops haemolytic anaemia due to complete absence of this enzyme in her red blood cells.
What would be the net gain of ATP from each molecule of glucose passing through glycolysis in the red blood cells of this patient?
Pyruvate kinase catalyses the final step in glycolysis, transferring the phosphate group from phosphenol pyruvate to ADP, resulting in the production of ATP and pyruvate. Normally in glycolysis 2 ATP are used in the “investment” phase and 4 ATP are produced in the “payback” phase. Without this final step catalysed by pyruvate kinase, the net gain is zero rather than the normal 2 ATP. Lack of ATP production in the red blood cells of patients with pyruvate kinase deficiency affects all processes requiring ATP. The sodium potassium ATPase pump stops working without ATP to provide energy and the red cells will loose potassium to the blood plasma. Water will move down its concentration gradient out of cells causing them to shrink resulting in cellular death and hemolytic anaemia. Red blood cells are particularly sensitive since they lack mitochondria and are reliant on glycolysis for ATP production.
What is measured by a direct Coombs test?
Antibodies bound directly to the surface of red blood cells.
The direct Coombs test (also known as a direct antiglobulin test) is used when immune-mediated hemolytic anemia is suspected. An immune response attacking the patient’s own RBCs could be by autoimmunity, alloimmunity or a drug-induced immune-mediated mechanism. The test determines if antibodies or complement system factors have bound to RBCs surface antigens in vivo.
Which base is folic acid required for the synthesis of?
Thymidine
What is the cause of pernicious anaemia?
Auto-antibodies interfering with the production or function intrinsic factor by the parietal cells of the stomach.
Intrinsic factor is essential for the absorption of vitamin B12 in the ileum. Without intrinsic factor, vitamin B12 deficiency will occur and production of red blood cells will be impaired casuing anaemia. “Pernicious” means “deadly”. Pernicious anemia was often fatal in the past before vitamin B12 treatments were available.
Which would be the most appropriate treatment for a patient with pernicious anaemia?
- orally administered B12 supplements
- intramuscular injections of a B12 supplement
Intramuscular injections
Pernicious anaemia results from intrinsic factor deficiency which prevents absorption. Orally administed B12 supplement would therefore also not be absorbed. In the past B12 injections were usually given to treat the disease. However nasal sprays containing B12 are now more frequently used.
Whhat is a schistocyte?
A fragmented part of a red blood cell seen in patients with haemolytic anaemia
What are NSAIDs used for?
Treatment of acute or chronic conditions with pain and inflammation
Eg.
Osteoarthritis Rheumatoid arthritis Back pain Headache Migraine Acute gout
What is microcytosis?
Condition in which red blood cells are unusually small as measured by their mean corpuscular volume
What is hypochromia?
Red blood cells paler than normal.
Red blood cells have an area of pallor in the centre due to a reduction of haemoglobin.
How does the body adapt to anaemia?
- increased BPG in cells
- increased release of erythropoietin
- decreased systemic vascular resistance to increase systemic venous return (increased preload)
- tachycardia
How does anaemia of chronic disease cause anaemia?
Inflammatory stimulus activated monocytes and T cells
- Inhibits erythropoietin release
- Inhibits erythroid proliferation
- Increased haemophagocytosis of macrophages
- Increased synthetic release of hepcidin
What test is used to diagnose hereditary spherocytosis?
EMA (eosin-5 maleimide) binding test
What is the EMA test?
Eosin-5-Maleimide binding test
Eosin-5-Makeimide is a fluorescent dye that binds to band 3 protein in the red cell membrane.
Therefore. It can be used in the diagnosis of hereditary spherocytosis
Why are spherocytes problematic?
Decreased surface area to volume ratio leads to loss of deformability
They are trapped and destroyed in the spleen.
This poor deformability is only a problem in the spleen because cells have a normal lifespan following a splenectomy
