CSI 2: Sickle Cell Disease Flashcards
Haemoglobin’s key structural features
- 4 globin chains [2 alpha, 2 beta]
- 4 haem groups [each a porphyrin ring bound to Fe2+, that can carry 1 oxygen molecule]
- Quaternary structure mainly alpha helices
- Hydrophobic core, hydrophilic surface
How many globin chains are there and what codes for each?
Eight
Alpha globin cluster - alpha1, alpha2, zeta
Beta globin cluster - epsilon, G gamma, G alpha, delta, beta
Types of Hb found in healthy adults
HbA - 2 alpha, 2 beta
HbA2 - 2 alpha, 2 delta
HbF [very small amount] - 2 alpha, 2 gamma
P_50
Value at which 50% of the haem groups are bound to oxygen
Describe why Hb’s affinity for oxygen helps it transport oxygen around the body appropriately
- In the lungs, partial pressure is 100 torr and Hb is quickly saturated, picking up oxygen due to its high affinity for oxygen
- Approaching the tissues, where partial pressure is 26 torr, Hb can release around 66% of the oxygen bound to it
What is positive cooperativity?
Once one oxygen molecule binds to a haem group within a Hb molecule, there is a conformational change, which makes it easier for another oxygen molecule to bind to the next haem group.
How is 2,3-BPG formed?
Via the Rapoport-Luebering Shuttle from some of the 1,3-BPG produced in glycolysis, using bisphosphoglycerate mutase [isomerisation]
What is 2,3-BPG’s effect on the oxygen dissociation curve?
- Allosteric effector that binds to a site in deoxyHb distant from haem groups stabilising the structure
- Decreases the affinity of Hb for oxygen, allowing more oxygen to be released at tissues
- Partially responsible for the oxygen dissociation curve (pure Hb looks like myoglobin)
What is the Bohr effect? Why is it helpful?
Rapidly respiring cells → decreasing pH, increasing CO2
H+ and CO2 - both allosteric effectors, decreasing affinity of Hb for oxygen, allowing for oxygen release at tissues
Foetal Hb and oxygen dissociation
- Gamma chain differs from beta chain by one single AA substitution
- Decreased affinity for 2,3-BPG
- Increased affinity for oxygen
- Allows for obtaining oxygen from maternal HbA
How is HbS different from HbA?
HbS → non-conservative missense mutation
Glutamate at position 6 on the betaglobin chain is replaced by valine
What kind of mutation causes the formation of HbS?
Autosomal recessive
- 2 copies of the mutation are required to manifest symptoms
How does sickling occur when Hb is deoxygenated?
- Conformational change means hydrophobic valine residue protrudes from surface
- Groove formed by phenylalanine-85, leucine-88 and alanine-70 on another Hb forms hydrophobic interactions with the valine-6
- HbS polymerises into long rods (7Hbs make a nucleus i.e. initiation point for fibre formation)
- Rigid fibres distort shape of RBC
What does the hydrophobic pocket refer to?
Hydrophobic interactions between the groove formed by phenylalanine-85, leucine-88 and alanine-70 on one Hb and valine-6 on another
What are the key indicators of SCD on blood films?
- Sickle cell
- Reticulocytes (compensation)
- Target cells (hyposplenism)
- Polychromasia
Also Howell-Jolly bodies (hyposplenism)
What are the key indicators of SCD on blood cell count?
- Low Hb count
- High reticulocyte count
- High mean cell volume
What are three ways you could diagnose SCA?
- Blood films & cell count
- Electrophoresis
- Sickle cell solubility [not very solid evidence]
Compare the band patterns of SS, AS and AA having done electrophoresis
AA - Band closer to anode (glutamate -ve)
AS - Bands at HbA and HbS
SS - Band further away from anode (valine neutral), stronger band for HbF
How does a sickle solubility test work?
- Reducing agent sodium dithionite added
- Oxygen released from haemoglobin and sickling occurs if positive
- Increase in optical density
*Can’t differentiate between HbSS or HbAS → electrophoresis, high performance liquid chromatagraphy
How does sickling lead to jaundice and gallstones?
- Repeated sickling weakens the red blood cell membrane causing intravascular haemolysis (SC shortened 20 day lifespan)
- Build up of unconjugated bilirubin (breakdown product of haem group of RBC) in blood
- Yellow colour → jaundice
- Hardening and solidifying of bile salts → gallstones
What are the main complications of SCD?
- Vision loss
- Jaundice
- Stroke
- Bone crises
- Acute crises (dactylytis, splenic sequestration, acute chest syndrome)
- Chronic conditions (kidney damage, retinopathy, gallstones)
- Splenomegaly
What is vaso-occlusion?
Blockage of microcirculation including capillaries, leading to ischaemia and tissue necrosis
How does vaso-occlusion occur?
- Rigid sickle cells get stuck by binding to p-selectin on the endothelial lining
- Sickle cells aggregate together, causing a blockage
How does SCA cause vision loss?
- Vasocclusion of microvessels in the retina, increasing pressure, damaging endothelial lining
- Sickle cell ischaemia
3a. Retinal cell detachment → cell death
3b. Release of cytokines → stimulates growth of new collateral blood vessels through angiogenesis
4b. Collaterals grow over areas of vision
5b. Collateral blood vessel rupture → haemorrhagic stroke
What are the two types of stroke?
Ischaemic - due to inadequate blood supply to an organ
Haemorrhagic - due to leakage of blood into tissues
How does SCA cause bone crises?
- Reduced blood flow to bone marrow due to microvascular occlusion
- Oxygen deprivation
- Prolonged ischaemia → infarction → avascular necrosis/osteonecrosis
- Ischaemia exacerbated sickling further
- Severe joint pain
What is an infarction?
Tissue death due to inadequate blood supply
What is dactylytis?
Pain in hand joints due to blood vessels being occluded by sickled cells
How is splenic sequestration caused by SCA?
Blood vessels leading out of spleen are occluded by sickle cells, causing a pooling of blood in spleen → sudden drop of Hb in bloodstream
How is hyposplenism caused and what are its implications?
Repeated vaso-occlusion in the abundant vasculature of the spleen impair its function
1. Increases vulnerability to encapsulated bacterial infections → sepsis
2. Splenomegaly - extra splenic tissue growth to compensate for hyposplenism
What is acute chest syndrome caused by?
Pulmonary vaso occlusion → fever, cough, chest pain, rapid breathing
What are the four main triggers of vaso-occlusive crises?
- Infection
- Exercise
- Cold temperature
- Dehydration
How does infection trigger a vaso-occusive crises?
Increased basal metabolic rate → more deoxyHb → more sickling
How does exercise trigger a vaso-occusive crises?
Increased demand for oxygen → more deoxyHb → more sickling
How does cold temperature trigger a vaso-occusive crises?
Peripheral vasoconstriction → narrowed blood vessels → increased susceptibility to blockage
How does dehydration trigger a vaso-occusive crises?
Increased blood viscosity → increased chance of aggregation and blockage
How can SCD be generally managed?
- Over the counter painkillers (also called analgesia)
- Limit exercise and prioritise resting
- High fluid intake
- Heat packs to stimulate vasodilation, increasing oxygen reaching local tissues
Treatments for SCD
- Blood transfusions to treat anaemia
- Painkillers to treat crises (standard ibuprofen, opioids for severe pain)
- Bone marrow transplants for severe cases in younger patients with a matched donor
- Gene therapy
- Prophylactic antibiotics
- VTE prophylaxis (preventing thrombosis)
- IV fluids (maintaining hydration with saline)
- Cognitive Behaviour Therapy (to take care of mental health issues arising from living with a chronic illness)
Interventions to reduce the frequency of future sickle cell crises
Hydroxycarbamide
Monoclonal antibody crizanlizumab
How does hydroxycarbamide work?
- Increases synthesis of HbF to compensate
- Reduces neutrophil and reticulocyte production (neutrophils promote vaso-occlusion)
How does crizanlizumab work?
It binds to p-selectin expressed on endothelial surfaces and block sickle cell adhesion to vessels
What is a potential downside to blood transfusions? How can this be mitigated?
Could cause hemosiderosis (iron overload in organs); iron chelation therapy can help remove excess iron
What is a potential downside to hydroxycarbamide?
Increased risk of infection because neutrophils numbers are reduced
How can CRISPR gene editing be used to tackle SCD?
1a. Cas9 enzyme can target the faulty betaglobin gene in sickle cells, using a guide RNA
1b. Incorrect T base is replaced by A, repairing gene
2a. Cas9 breaks gene that encodes repressor of fetal Hb production - BCL11A
2b. DNA damage and error prone repair to BCL11A gene → stop coding for repressor
2c. Gene expressed → fetal Hb produced
Applying ICE
Ideas → the patient’s perception of what they have, their reasoning
Concerns → what is the patient worried about?
Expectations → what do they think the doctor can do for them? what do they want?
