Haematology Flashcards
What is haptoglobulin and what do high levels of it suggest?
It is an alpha 2 globulin in the plasma and binds to haemoglobin release from dead erythrocytes to from haptoglobulin-haemoglobin complexes which are removed by the spleen
High levels of haptoglobulin suggest haemolytic anaemia - where RBCs are destroyed faster than they are made
What are the gamma globulins of the plasma and what do high levels of them suggest
C reactive protein and immunoglobulins
High levels suggest infection or myeloma (a blood cancer which develops from cells in bone marrow)
What is the main difference between plasma and interstitial fluid?
Plasma contains more protein than interstitial fluid
What is haemopoiesis
The formation and development of blood cells
What is haemopoiesis regulated by
Genes
Micro environment
Transcription factors
Growth factors (glycoprotein hormones which bind to cell surface receptors that regulate proliferation and differentiation of HSCs and regulated function of mature blood cells)
What is the erythropoiesis growth factor
Erythropoietin
What are the granulocyte and monocyte production growth factors
G-CSF
G-M CSF
cytokines eg interleukins
Megakaryocytopoiesis and platelet production growth factor
Thrombopoietin
What other cells/structures are distributed with HSCs in the bone marrow?
In the bone marrow HSCs interact with
- HSC progenitor cells
- mesenchymal stem cells
- endothelial cells
- vasculature
What can happen if the regulation of haemopoiesis is disrupted
The balance between proliferation and differentiation is disrupted and can get leukemia (due to too many cells) or bone marrow failure (due to not enough cells)
Life span of different blood cells
Erythrocytes - 120 days Platelets - 10 days Monocytes - several days Neutrophils - 7-10 hours Eosinophils- little less than neutrophils Lymphocytes - v variable
2 essential characteristics if HSCs
Can self renew - some daughter cells stay as HSCs to prevent HSC pool depletion
Can differentiate into mature progeny - other daughter cells can differentiate into other types of cells through myeloid or lymphoid progenitors
Changes in site of haemopoiesis between adult and foetus
1) in mesoderm of yolk sac
2) at 6-8 weeks of gestation, liver takes over
3) 10 weeks into gestation, bone marrow takes over. Adult haemopoiesis happens mainly in bone marrow esp in pelvis, femur, and sternum
What are polychromatic erythrocytes
Have a blue tinge due to high RNA content - lose ribosomes after a few days and become mature erythrocytes
What does it mean if we see nucleated RBCs in blood
There’s a high demand for bone marrow to produce RBCs so immature RBCs are being prematurely released into circulation
Overall progression of RBC development
In bone marrow: Common myeloid progenitor gives rise to proerythroblasts —> erythroblasts (have nucleus still)
In peripheral blood: erythroblasts —> reticulocytes (lost nucleus) —> erythrocytes
Where and how is erythropoietin (EPO) made and how does it work
Made in the kidneys in response to hypoxia and anaemia. Works by supply and demand feedback
Hypoxia or anaemia in kidneys makes kidneys produce EPO
EPO interacts with erythropoietin receptors on red cell progenitor membranes which increases bone marrow activity, causing bone marrow to produce more red cells
What is iron needed for in the body
1) Oxygen transport
2) mitochondrial proteins
- cytochrome a,b,c (for ATP synthesis)
- cytochrome P450 (for hydroxylation reactions)
Iron is important for healthy hair skin nails and mucous membranes
Where is iron absorbed
Duodenum
What is the best absorbed form of iron, and what does the other form of iron need for absorption
Haem (ferrous / Fe2+ ) is best absorbed
Non haem (ferric / Fe3+) needs reducing substances like vitamin C
(sources of non haem iron such as soya beans contain phytates which reduce absorption)
Why is iron haemostasis needed
Excess iron can be toxic to organs like liver and heart
There is no physiological mechanism that excretes iron so it’s absorption must be controlled to 1-2 mg absorbed per day from diet
What is transferrin
The transport protein in plasma that delivers iron to bone marrow fro erythropoiesis and for its use in enzymes and muscles
How does erythropoietic activity affect hepcidin synthesis and what does this do
It suppresses hepcidin synthesis which ensures iron supply by increasing ferroportin in the duodenum enterocyte - this increases iron absorption
What does ferroportin do
It’s a transmembrane protein that transports iron from inside to outside of cell
What is an enterocyte
Cell of intestinal lining
What happens when iron storage is high
When ferritin stores are full, hepcidin production increases
The hepcidin binds to and degrades ferroportin
This stops ferroportin transporting iron out of the enterocytes
So iron stays in enterocyte - this allows iron to be disposed of as soon as enterocyte dies and is shed into the gut lumen
How can chemicals released during inflammation or chronic disease cause anaemia
Pro inflammatory cytokines ( IL-6, IL-1 TNf alpha, IFN gamma)
- decrease EPO production
- increase hepcidin production
This causes reduced Fe transport availability and absorption, resulting in anaemia of inflammation/chronic disease
What is hepcidin
A hormone that regulates iron ABSORPTION in the GUT
What is another name for vit B12
Cobalamin
How can vit B12 or floats deficiency affect rapidly dividing cells
Vit B12 and folate are needed for dttp synthesis (an important precursor for making thymidine DNA synthesis)
So deficiency in them means DNA synthesis is inhibited
This can cause megaloblastic anaemia (production of megaloblasts - abnormally shaped large RBCs)
This affects rapidly dividing cells such as
- cells in bone marrow - can grow but not divide
- epithelial surfaces of mouth and gut
- gonads - nerves
How is vitamin B12 absorbed
B12 binds to intrinsic factors (IF) in gastric parietal cells
B12-IF binds to receptors in ileum
Causes of B12 deficiency
Malabsorption - coeliac disease
Low B12 intake - veganism
Low IF secretion - pernicious anaemia
Lack of stomach acid - achlorhydria
How do erythrocytes die
Get destroyed by macrophages of spleen after 120 days
The iron is returned to bone marrow by transferrin and recycled there
Death releases bilirubin - excreted in bile
Where is folic acid absorbed
Small intestine (duodenum and jejunum )
When do folic acid requirements increase
Pregnancy
Increased RBC production eg sickle cell anaemia
A patients results show she has folate deficiency. What changes in the RBCs would you expect to see and why
Presence of macrocytes
As a result of impaired DNA synthesis which causes the cells to lack the DNA to divide but to continue to grow - causes megaloblastic erythropoiesis
What is the most abundant cytoskeleton protein if erythrocytes
Spectrin
What is the advantage if the biconcave shape of RBCS
Increases manoeuvrability through small blood vessels
Incraeses ability to RELEASE O2
Where does haemoglobin originate
The haem component is synthesised in mitochondria
The glob in component is synthesised in RER
Found exclusively in RBCs
What is the structure of the haem group
Ferrous iron (Fe2+) bound by a porphyrin ring - each Fe2+ can bind to one O2 molecule
Where does the globin component of haemoglobin come from in haemoglobin
There are 8 functional globin chains which are encoded by several genes separated into 2 gene clusters - the alpha globin cluster and the beta globin cluster
Alpha cluster - zeta and alpha genes
Beta cluster - delta, epsilon, beta and gamma genes
What is the secondary and quaternary structure of Hb
Secondary: 75 percent of the alpha and beta chains are in alpha helix form
Quaternary: 4 globin chains and 4 haem groups
Forms a tetramer (4 subunit protein)
Hydrophobic core and hydrophilic outside
Forms an approximate sphere
What is the majority of Hb in healthy adults
HbA - 2 Alpha and 2 beta
What is HbA2 and HbF made of
HbA2 = 2 alpha 2 delta HbF = 2 alpha 2 gamma
What is the p50 value and how does it differ between myoglobin and haemoglobin
P50 = the partial pressure of oxygen at which 50 percent of the haem groups are saturated with oxygen
P50 values:
Myoglobin = 2 torr
- has greater affinity for o2
- saturated at lower pp of o2
- produces a hyperbolic curve
- reacted to its function as its found in muscle
Haemoglobin = 26 torr
- lower affinity for o2
- produces a sigmoidal curve
- Positive cooperativity
- saturated at higher pp of o2
What allosteric effectors affect haemoglobins affinity for o2 and how
2,3-BPG (aka 2,3-DPG)
- formed by phosphorylation of 1,3-BPG by diphosphoglycerate mutate (DPGM) via Rapoport Luebering shuttle
- binds to site on haemogloblin distant from haem groups and stabilises the structure, lowering the haemoglobins affinity for oxygen - thus causes it to have sigmoidal dissociation curve
H+ (released from formation of lactate which replenishes NAD+ stores) and CO2 (formed from decarboxylation of pyruvate)
- bind to sites away from haem groups and lower affinity for oxygen
- BOHR effect
- so a decrease in pH of blood will mean more O2 is released into tissues
What is Absorbance
A = log 10
= the fraction of light transmitted through a blank solution/test solution
(So If solution has absorbance of 1.0, 10 percent of light is transited and 90 percent is absorbed)
What is the beer lambert law
Absorbance = the extinction coefficient x the concentration of the absorbing substance x the path length
describe the role of myoglobin and how it’s properties allow it to carry out this role
It is an oxygen binding molecule found in muscle, with one name group
It’s high affinity for o2 (higher than haemoglobin) allows it to carry out this role
But it’s lack of cooperativity means that it is poor at releasing o2
Describe spectrophotometry spectrum for oxyhaemoglobin and deoxyhameoglobin
Also explain why they might overlap
The graph for deoxyhaemoglobin has one peak at 560 nm
The graph for oxyhameoglobin has two peaks at 540 and 580
The graphs would overlap if not enough sodium dithionite (reducing agent) was added when forming the deoxyhaemoglobin
What is a clinical use of spectrophotometry
Can be used to check the respiratory status of newborn infants (as can be used to follow changes in oxygen binding to haemoglobin)
What is pulse oximetry
A non invasive way of measuring oxygen saturation levels - relies on difference in absorbance of oxy and deoxyhaemoglobin
Oximetry is clipped onto finger and emits red light and infrared light
Oxyhameoglobin absorbs more infrared but less red light than deoxyhaemoglobin
The ratio of absorbances be converted into oxygen sat. via beer Lambert law
What is carboxyhameoglobin (CoHb)
generated by binding of carbon monoxide to ferrous iron
(Hb has a greater affinity for carbon monoxide than oxygen so carbon monoxide can outcompete o2 - makes carbon monoxide possibly fatal at even v low levels)
What is methaemoglobin (MetHb) and methaemaglobinaemia
MetHb is generated when Fe2+ is oxidised to Fe3+
- impairs oxygen binding
- left shift of dissociation curve
- results in tissue anoxia as o2 is less readily given to tissues
- blood is a blue ish / chocolate colour
Methemoglobin reducatse reduces MetHb back to Hb
Methaemaglobinaemia can be hereditary and due to
- deficiency in Methemoglobin reducatse
- production in mutant Hb: Hb M which is resistant to reduction
Or it can be due to exposure to chemicals (alanine dues) such as
- p-chloroalanine
- nitrates
-local anaesthetics eg benzocaine
Describe electrophoresis results of HbA /HbS/mixed and why
HbA ran further to positive electrode than HbS bc it’s more negatively charged than HbS
- HbA has glutamate which is hydrophilic and negative
- HbS has valine which is hydrophobic and uncharged
Mixed sample mimicked results of someone heterozygous for the condition as it contained both HbA and HbS bands
Causes of microcytosis
Defect in haem group:
- iron deficiency
- anaemia of chronic disease (as this reduces iron absorption and use)
Defect in globin chain
- defect in alpha chain —> alpha thalassemia
- defect in beta chain —> beta thalassemia
List the three types of macrocytes
Round macrocytes
Oval macrocytes
Polychromatic macrocytes
List the causes of macrocytosis
- vit B12/folate deficiency —> causes megaloblastic anaemia due to impaired DNA synthesis as dttp can’t be produced, so cells can grow but not divide and form megaloblasts
- pregnancy
- liver disease and alcohol toxicity
- haemolysis (polychromasia)
How much do a normal RBC is pale and why
1/3 of diameter is pale
Due to this flatter section in the centre of RBC having lower conc of haemoglobin
What is hypochromia and what other physical RBC feature is it usually associated with
- larger area of central pallor than normal
- due to lower Hb conc and flatter cell
- often associated with microcytosis (so is seen in thalassemia and iron deficiency)
What is polychromasia, it’s causes and what other physical feature of RBC is it squally associated with
- blue tinge to RBC
- seen in immature RBCs —> reticulocytes, so seen in reticulocytosis (increased conc of reticulocytes in blood)
- reticulocytosis occurs due to blood loss or destruction of RBC - haemolysis
- asssociated with macrocytosis (as reticulocytes are larger)
2 causes of reticulocytosis
Bleeding
RBC destruction - haemolysis
How to detect reticulocytes
- look for polychromatic macrocytic RBC on blood film
Or - due new methylene blue test which stains for higher RNA content of reticulocytes
What is anisocytosis
RBCs show more variation in size than normal
What is Poikilocytosis
RBCs show more variation in shape than normal
What are target cells? List some conditions where they may be seen
Accumulation of Hb in central pallor
- liver disease and alcohol toxicity
- haemoglobinopathies eg thalassemia or sickle cell
- hyposplenism
- obstructive jaundice
What is hyposplenism
Remove spleen or spleen doesn’t work properly
When can HbS occur
When One or two copies of an abnormal beta globin gene are inherited
Mutation is charged glutamate at position 6 on beta chain replaced by uncharged valine
When are red cell fragments aka schistocytes seen
Indicate that a red cell has fragmented
Result from shearing process caused by platelet rich platelets in small blood vessels —> disseminated intravascular coagulopathy
How are irregularly contracted cells produced and what do they look like
Caused by glucose 6 phosphate dehydrogenase deficiency
G6PD is in hexose monophosphate pathway which is linked to glutathione metabolism which protects RBC from oxidant damage
Deficiency causes oxidant damage to cell from infection or exogenous sources eg food/drugs
Causes episodes of severe intermittent intravascular haemolysis
Appearance :
Irregular outline
Microcytic
No central pallor
Advantage of G6DP deficiency
Resistance against malaria
How are Heinz bodies formed
Denaturation of Hb in G6PD deficiency
Forms round inclusions
Inheritance pattern for G6PD
X linked
Inheritance pattern for sickle cell anaemia (which is HbSS)
Autosomal recessive
What counts as sickle cell disease
HbSS - sickle cell anaemia
Some heterozygous states which cause sickling eg HbSC or HbS beta thalassemia
Effects of functional hyposplenism on blood film
Presence of:
Target cells
Howell Jolly bodies (DNA remnants)
- Would both have been removed by spleen if it worked
Diagnosis of SCD
Sickle solubility test - reducing agent reduces oxyHb to deoxyHb which precipitates out and turns solution turbid
But definitive diagnosis:
- electrophoresis
- high performance liquid chromatography HPLC
(Separate proteins based on charge)
Lab features of SCD
Lower Hb (60-80 g/L) Higher reticulocyte count Sickle cells Boat cells Howell jolly bodies Target cells
How can SCD cause need for a colicystectomy
Sickle cells have higher turnover of 20 days
So increased haemolysis
Which means increased bilirubin
Which can cause gall stones
Which needs removal of gall bladder which is a colicystecomy
How are children with SCD screened for stroke risk and what action is taken
Measure velocity of blood flow through cerebral veins
High risk children can be given blood exchange therapy
Why is diagnosis of acute chest syndrome usually delayed
Similar symptoms to infection so hard to differentiate
Symptoms of acute chest syndrome
Tachycardia Fever Cough Chest pain Pulmonary infiltrates on chest x ray
2 effects of sickling
1) increased RBC turnover and haemolysis
- causes anaemia, jaundice, gall stones
2) vaso occlusion
- can happen in bone, retina, spleen, brain, lung kidney etc
- stroke, hyposplenism, necrosis, etc
2 functions of spleen
Immunity
Removal of old or damaged RBC
Why is splenic sequestration only seen in children
Repeated vaso occlusion in spleen causes it to stop working by age 5 - functional hyposplenism
Why is there no symptoms of anaemia due to the low Hb in SCD
low Hb (60-80 g/L) but the HbS has a lower affinity to O2 than HbA so releases O2 to the tissues more easily (right shift on ox dissociation curve) This means than someone withSCD doesn’t get anaemia symptoms that so one with HbA at 60-80 g/L would get (They still have anaemia as anaemia is low Hb, but don’t get the symptoms)
Early presentations of SCD
Start at 4-6 Months when HbF changes to HbS
Dactylitis
Infection
Splenic sequestration (pooling of RBCs in spleen causing vaso occlusion)
How does SCD cause increased susceptibility to encapsulated bacteria and what actions are taken
Vaso occlusion/pooling of RBCs causes splenic sequestration
Repetition of this causes spleen to stop working (functional hyposplenism) and spleen may have to be removed
Spleen has role in immunity so this results in increased susceptibility to encapsulated bacteria eg that cause pneumonia or meningitis
- regular immunisations
-prophylactic antibiotics
On a blood film, what does the presence of clear vacuoles in the cytoplasm of neutrophils suggest?
Bacterial infection
Characteristic features in blood film of megaloblastic anaemia
Hypersegmented neutrophils and macrocytes
Esp oval macrocytes
Key functions of a monocyte
Phagocytosis of microbes
Production of cytokines
Phagocytosis of apoptotic cells
Key functions of lymphocytes
Production of cytokines
Production of antibodies
What do high leevls fo haptoglobin suggest
Haemolytic anaemia, where RBCs are being destroyed faster than they are made
What separates the red cells from the plasma after the addition of an anticoagluant and centrifugation
Buff coat
What is apheresis
When a blood donor is collected and passed through centrifuge to separate particular cellular components
What is buffy coat made of
Leaukocytes and platelets
How is interstitial fluid drained and what is it known as when it is drained
Through lymphatic vessles
Lymph
What do neautrophils release during inflammation
Neutrophil elastase
What is the effect of defective ir deficinet A1AT to the lungs
It can compromise the lung where degradation of lung tissue leads to loss of elasticty and respiratory problems
Main fucntions of albumin
Transportation of fatty acids
Maintaing osmotic pressure
Where are HSCs derived from in embryonic development
Yolk sac —> liver —> bone marrow
How long does it take for bone marrow to develop haematopoietic activity
10 weeks
Where does haemopoiesis occur in adults
Bone marrow of pelvis, sternum and femur
Where is erythropoietin synthesised
Kidneys
What gives rise to a pro erythroblast
Common myeloid progenitor
Cause of microcytic anaemia
Iron deficiency
Causes of macrocytic aneamia
Vitamin B12/folate deficiency
Major functions of iron
Transport O2 in Hb
Mitochondrial proteins cytochrome a,b,c for production of ATP
3 clinical signs of iron deficinecy
Koilonychia - spoon shaped nails
Glossitis - inflammation of tongue
Angular stomatitis - inflammation of corner of mouth
What does vitamin B12 bind to in the stomach during its absorption
Intrinsic factor
What suppresses hepcidin synthesis
Erythropoietic activity
Where is intrinsic factor made
Gastric parietal cells
What is pernicious anaemia
Inadequate secretion of intrinsic factor from the gastric parietal cells (so less vit B12 can be absorbed in digestion)
So u get macrocytic anaemia due to vit B12 deficiency
Where do red cells undergo phagocytosis
Spleen
Where does the absorption of folate take place
Duodenum
What does the catabolism of haem produce and where is this excreted
Billirubin
Excreted in bile
When do the requirments of folate increase
Pregnancy
Low RBC production - sickle cell anaemia
What causes spherocytes
Disruption of vertical linkages
Loss of cell membrane without loss of cell cytoplasm
RBCs becomes less flexible
What causes hereditary elliptocytosis
Disruption of horizontal linkages
Lack of iron
What is polychromasia associated with
Macrocytosis
What is hyperchromia associated with
Microcytosis
What is anisocytosis
Red cells showing more variation in size than normal
What is poikilocytosis
Red cells showing more variation in shape than normal
What is polychromasia associated with
Macrocytosis
