Week 1 - D - Macrocytosis and Macrocytic Anaemia - Megaloblastic, Non megaloblastic, Spurious

Question 1

What is macrocytosis? What is microcytosis?

Answer 1

Macrocytosis - this is when there is the presence of large red blood cells Microcytosis - this is when there is the presence of small red blood cells

Question 2

What is microcytotic anaemia?

Answer 2

This is where there is anaemia in which the red cells have a larger volume than normal

Question 3

How is size of the red blood cell expressed? What is the units of measurment?

Answer 3

Size of the red blood cells is measured in mean corpuscle (cell) volume The unit of measurement is femtolitres (fl) = 1x10^-15L

Question 4

When the automated analyser analyses a blood sample, from it we can calculate the: Haemotocrit (Hct) Mean cell haemoglobin (MCH) Mean cell Haemoglobin concentration (MCC) What three things are measured by the automated analysers which allows for these to be calculated?

Answer 4

Automated analysers measures: * Haemoglobin concentration * Number of red blood cells (RBC concentration) * Size of red blood cells (mean cell volume) Hct- uses the number of red cells and the mean cell volume - then can use this to work a ratio to the total blood volume MCH - number of RBCs divided by the Hb concentration MCC - number of cells divided by the Hb concentration, divided by the size of the cell to get the MCC

Question 5

It is important to known the difference between macrocytosis and a macrocytic anaemia Does the image show a macrocytosis or a macrocytic anaemia?

Answer 5

The RBC concentration and Hb is low and therefore the person is anaemic (<130g/L in a male, below 115 g/L in a female) The mean cell volume is high - macrocytosis The person therefore has a macrocytic anaemia

Question 6

the difference between macrocytosis and a macrocytic anaemia Does the image show a macrocytosis or a macrocytic anaemia?

Answer 6

Hb and RBC is normal - patient is not anaemic MCV is moderately high The patient has a macrocytosis

Question 7

What is the size of a red blood cell often compared to? What is the normal mean cell volume of a red blood cell?

Answer 7

The size of a red blood cell is often compared to a mature lymphocyte nucleus - these lymphocytes are small and have a condensed nucleus with a pale rim of blue cytoplasm The normal MCV is 80-100 ft (femtolitres)

Question 8

RBC smaller than nucleus of a lymphocyte – microcytic RBC larger than nucleus of a lymphocyte - macrocytic What is the MCV for a microcytic anaemia? What is the MCV for a macrocytic? What is the difference between mature and activated (atypical) lymphocytes?

Answer 8

• MCV= <80fl for microcytic
• MCV = >100fl for macrocytic
• Mature lymphocyte - small with condensed nucleus and pale rim of blue cytoplasm
• Activated - large with open nucleus and plenty of blue cytoplasm surrounding neighbouring RBCs

Question 9

The causes of acrocytic anaemia are either: Genuine (true) or Spurious (false) What are the two different categories for genuine macrocytic anaemia?

Answer 9

Megaloblastic and non-megaloblastic

Question 10

What red blood cell have a nucleus?

Answer 10

Red cell precursors - before reticulocyte formation - have a nucleus and are usually located in the marrow

Question 11

Erythroblast/Normoblast: A normal red cell precursor with a nucleus Describe the difference between a reticulocyte and a erythrocyte?

Answer 11

Reticulocyte - this red cell circulates in the blood for a couple of days before becoming a mature red blood cell - it is slightly larger and contains a small amount of RNA - it is therefore usually polychromatic as has blue (RNA) and red (Hb) colours The reticulocyte can be biconcave

Question 12

Developing erythroid cells in the marrow Accumulate Hb Reduce in size Stop dividing and lose nucleus What regulates the developing erythroid cells to stop dividing and lose the nucleus?

Answer 12

The Hb content of the cell regulates the stopping of erythroid cell division and its enucleation

Question 13

Pronormoblast has absolutely no haemoglobin in it What does megaloblastic actually mean?

Answer 13

This means there is an abnormally large nucleated red cell precursor with an immature nucleus Megalobastic aenamia – means the bone marrow is actually full of megablasts

Question 14

Arrows - top to bottom Early normoblast Late normoblast Enucleation about to occur On the right can see megaloblasts have a much more open nucleus (immature) What are the predominant defects that cause megaloblastic anaemias?

Answer 14

Megaloblastic anaemias are characterised by predominant defects in DNA synthesis and nuclear maturation with relative preservation of RNA and haemoglobin synthesis

Question 15

Is the cytoplasm development affected in megaloblastic anaemias?

Answer 15

Cytoplasmic development is not affected (this is what is effected in microcytic anaemia)

Question 16

In the few erythroblasts that survive as ‘megaloblasts’, cytoplasm development occurs normally and triggers enucleation This leads to a ‘bigger-than-normal’ red cell But overall, there are fewer of these and hence the patient is anaemic What happens during development from primitive cells with megaloblasts? (ie cell division and apoptosis) How do the megaloblast become enucleated?

Answer 16

With the maturation in megaloblasts, there is increased primitive cell division but a greater increased apoptosis and therefore fewer red cells are present Because there is normal cytoplasmic development, the cell is enucleated and the red cells will enter the bloodstream however fewer will be present

Question 17

Important The megolblasts are not cells that increase in size, they are in fact cells that fail to decrease appropriately in size during erythroid division What happens to the colour of the RBC as it goes through development and why?

Answer 17

The colour of the RBC changes from blue to red This is because the RNA (blue) present in the cell which is required for making proteins produces more and more haemoglobin therefore making the cell turn more red until enough haemoglobin is present the the cell loses its nucleus

Question 18

Macrocyte on the right - shows there is enough Hb present for enucleation of the cell and the entry into the blood, it is just there a fewer of the cells present - therefore macroyctic anaemia

Answer 18

Just a pic for info

Question 19

Is the large cell size in megaloblastic anaemia due to the cell becoming bigger during erythroid development?

Answer 19

The larger cell size in megaloblastic anaemia is not due to an increase in the size of the developing cell, but A FAILURE TO BECOME SMALLER

Megaloblastic anaemias are characterised by predominant defects in DNA synthesis and nuclear maturation with relative preservation of RNA and haemoglobin synthesis resulting in an abnormally large nucleated red cell precursor with an immature nucleus - the megaloblast

Question 20

What are causes of megaloblastic anaemia?

Answer 20

B12 deficiency Folate deficiency Drugs related or rare inherited conditions

Question 21

Why does lack of B12 and folate cause megaloblastic anaemia?

Answer 21

This is because folate and vitamin B12 are essential cofactors for nuclear maturation and Enable chemical reactions that proide enough nucleosides for DNA synthesis

Question 22

State again why B12 and folate deficiency can cause megaloblastic anaemia? What are the two interlinked cycles that these two are involved n?

Answer 22

B12 and folate are essenital cofactors for nuclear maturation and They enable chemical reactions that provide nucleosides for DNA synthesis They are involved in the methionine and folate cycle which are interlinked

Question 23

Folate cycle is improtant for nucleoside synthesis - give an example of the nucleoside change? What does the methionine cycle produce?

Answer 23

Folate cycle is important for nucleoside synthesis - ie uridine to thmyidine Methionine cycle produces - s-adenosyl methionine - this is an important methyl donor (potential impact on DNA, RNA proteins lipids and folate intermediates)

Question 24

What conversion does the folate cycle catalyse? What does the methionine cycle provide?

Answer 24

Folate cycle catalyses the conversion of uridine to thymidine Methionine cycle helps to to prouce s-adenosyl methionine which provides a methyl donor - these are required for a number of different chemical reactions

Question 25

What change in the methionine cycle is dependent upon B12 and N5-metyl tetrahydrofolate?? If there is a deficiency in B12 (or folate), what condition screened for on day 5 guthrie test can this cause? (this condition is an autosomal recessive conditon - the B12/folate deficiencies can just cause the same thing later in life)

Answer 25

B12/N5 THF is important forthe conversion of homocysteine to methionine - is dependent on these vitamins If there is a problem with the homocysteine to methionine cycle - homocysteinuria can happen in which there is a build up of homocysteine - screened for Guthrie test

Question 26

When homocysteine recieves a methyl group from the folate cycle and from methylcobalamin, it becomes methionine which then creates s-adenonsyl methionine - this can give methyl group donors important for chemical reactions What can a build up of homocysteine result in?

Answer 26

This condition is homocystienuria

Question 27

Lets talk about vitamin B12 What is it also known as? Where is B12 found?

Answer 27

Cobalamin Vitamin B12 is mainly found in animal products It is released from these products into the acidic pH of the stomach

Question 28

How is B12 absorbed from the diet?

Answer 28

* When food enters, the gastric parietal cells secrete HCl (to break down the food) and intrinsic factor (cells found in fundus and body of stomach) - the B12 and intrinsic factor are unable to bind in the acidic pH and once reaching the duodenum - where pancreatic enzymes makes the pH more alkaline- the two can bind forming B12-intrinsic factor complex * This complex binds to cubilin receptors in the distal small bowel (terminal ileum) & is absorbed

Question 29

Important to know this to understand causes of B12 deficiency Name causes of this deficiency?

Answer 29

* Decreased dietary intake of the vitamin - ie vegans or vegetarians * Aatrophic gastiris, or pernicious anaemia, this can cause no production of intrinsic factor from the gastric parietal cells therefore no absorption * Chronic pancreatiits – cannot produce enzymes to make conditions alkalaine to allow fo the B12 and intrinsic factor to bind * PPIs or gastric bypass * Coeliac or chrons affecting absorption in the bowel

Question 30

What is pernicious anaemia?

Answer 30

This is the autoimmune destruction of the gastric parietal cells resulting in achlorhydria and no intrinsic factor therefore affecting vitamin B12 absoprtion Results in intrinsic factor deficiency with B12 malabsorption and deficiency

Question 31

Pernicious anaemia has some associations as many autoimmune conditions go together Name some common autoimmune conditions that go with pernicious anaemia

Answer 31

Hypothyroidism Vitiligo Addison’s disease Eg person with hypothyroidism begins to become anaemia, - suspect pernicious anaemia

Question 32

Which antibodies are raised in pernicious anaemia?

Answer 32

Anti gastric parietal antibodies - these are very sensitive but not specific Anti-intrinsic factor antibodies - these are very specific but not sensitive

Question 33

What is folate converted to for absorption and where is it absorbed?

Answer 33

Dietary folates are converted to monoglutamate and absorbed in the jejunum (both diffusion and active absorption)

Question 34

Folate - converted to monoglutamate and absorbed in the jejunum VitB12 - binds to intrinsic factor and attaches to cubulin receptors to be absorbed in the ileum What is the sources of both? How long do their body stores last? What is the daily requirements of both?

Answer 34

Folate - * Absorbed in jejunum (and duodenum) * Found in leafy vegetables and yeast * Body stores only last 4 months * Require 100 micrograms daily Vitamin B12 - * Absorbed in the ileum * Found in animal foods * Body stores last 2-4 years * Require 1-3 micrograms daily

Question 35

Need far more folate than B12 per day and folate stores do not last as long What are causes of folate deficiency? (diet, malabsorpton, utilisation reasons, drugs)

Answer 35

Inadequate intake - Dietary cause is more likely than B12 Malabsorption - Coeliac disease, Crohn’s disease Excess utilisation * Haemolysis * Exfoliating dermatitis * Pregnancy * Malignancy Drugs - anticonvulsants, methotrexate (folate antagonists)

Question 36

What are the clinical features of vitamin B12 and folate deficiencies?

Answer 36

Symptoms Anaemia symptoms - pallor (skin and conjuctival), fatigue Weight loss, diarrhoea, infertility Sore tongue, can have jaundice Developmental problems especially in children Neurological problems

Question 37

What is the difference between the tongue in iron deficiency and B12 deficiency anaemia?

Answer 37

In iron deficinecy anaemia- you get atrophic glossitis In B12/folate deficiency anemia - get a glossitis - but it is described aa a beefy tongue - big fat swollen red inflamed tongue

Question 38

The deficiencies may present exclusively with neurological symptoms and no haemtological findings therefor may be useful to check for megaloblastic problems or B12 problems when a person present with neurological signs What is the neruological condition caused by a vitB12 deficiency? What tract does it not affect?

Answer 38

This would be subacute combined degeneration of the spinal cord - it affects both the posterior (dorsal) column tract and the corticospinal tract It does not affect the spinothalamic tract and therefore pain, temperature and deep pressure sensations can still be intact

Question 39

B12 and folate deficiencies caused defects in the nuclear maturation which can affect multiple lineages Therefore what might you see on a FBC in a patient with these deficiencies?

Answer 39

Low haemoglobin Pancytopenia - erythropenia, leucopenia, thrombocytopenia (remember it is a thrombocyte so thrombocyt - to affect platelts , eg thrombophilia is love for clotting (not increased platelets - thrombocytosis is icnreased palatelets) MCV - increased

Question 40

Generally, the diagnosis should rely on the clinical situation and the vitamin B12 and folate levels. A blood film may be useful in certain cases if there is diagnostic uncertainty What would be seen on the blood count and film?

Answer 40

The blood count would show macrooavalocytes - the red blood cells are enlarged and oval shaped and Hypersegmeneted neutrophils - Normally 3-5 segments in neutrophils but in can increase in macrocytic anemia

Question 41

After carrying out a FBC, it eh MCV is high, the reticulocyte level can be measured to point in the cause of the macrocytic anaemia (dont need to measure of this if history, FBC and blood film point to B12/folate deficiency) If the reticulocyte level is high, what could this be saying is a cause of the anaemia? Why can patients with pernicious anaemia appear mildly jaundiced?

Answer 41

Anaemia due to haemolysis or haemorrhage Haemolytic anaemia - the RBCs are being broken down the and the bone marrow is trying to compensate - Patients with pernicious anaemia can appear mildly jaundiced due to intramedullary haemolysis Ineffective erythropoiesis: * Red cells die prematurely in the marrow * Haemoglobin and lactate dehydrogenase (LDH) are released from dead red cells * Haemoglobin converted to bilirubin

Question 42

If the reticulocyte level is low, what is carried out?

Answer 42

Measure Assay serumB12 and folate (beware cause may have false positives here may occur) If these are low then carry out antibody tests for pernicious anaemia

Question 43

What are the antibody tests to check for pernicious anaemia? Which is sensitive but not specific? Which is specific but not sensitive? If folate levels, are low, why is it still very important to check for low B12?

Answer 43

Anti-gastric parietal cell antibody - sensitive but not specific Anti-intrinsic factor antibody - specific but not sensitive Extremely important test to rule out vitamin B12 deficiency before initiating folic acid therapy, which may worsen neurological complications in undiagnosed vitamin B12 deficiency.

Question 44

(FLAWS: anti-GPC sensitive, not specific; anti-IF: more specific, not sensitive) a bone marrow examination is not usually required in patients In the treatment of megalblastic anaemia, it is improtant to treat the cause where possible WHat is the treatment of vitB12 and folate deficiencies? (vitB12 treatment if dietary may only require a low dosing Tx)

Answer 44

VitB12 deficiency - due to malabsorption - IM injections with hydroxycobalamin 1mg 6times in 2 weeks then 1mg every 3 months for life (Vitamin B12 (hydroxycobalamin in Europe) injections for life in pernicious anaemia) Folate deficiency - 5mg oral folicacid daily for 4 months

Question 45

What may be a different treatment to vitB12 deficiency? (not yet licensed in the UK)

Answer 45

IN some patients very high dose oral may be effective - works under the assumption that small amounts of vitB12 pass via diffusion red cell transfusions are the last resort

Question 46

The causes of non-megaloblastic macrocytic anaemia tends to affect the red cell membrane What are these causes? (usually screened for if the blood film didnt show hypersegmeneted neutropjils/macroovalocytes or if the vitB12/folate is normal)

Answer 46

Liver disease Hypothyroidism Alcohol Marrow failure - usually looked for if the other three are not the cause

Question 47

What conditions may have caused the marrow failure? How is this checked? Basically if test results dont show B12/folate deficiency, dont reveal liver or hypothyroid disease or alcohol then carry out marrow failure examaintion

Answer 47

Bone marrow biopsy * Aplastic anaemia * Myelodysplasia * Myeloma

Question 48

What is myelodysplasia? What is aplastic anaemia? Name a drug that can cause it?

Answer 48

A myelodysplastic syndrome is a type of cancer in which the bone marrow does not make enough healthy blood cells and there are abnormal (blast) cells in the blood and/or bone marrow. Aplastic anemia is a rare disease in which the bone marrow and the hematopoietic stem cells that reside there are damaged. - this causes pancytopneia - chloramphenicol used to treat eye infections can cause this

Question 49

What is spurious macrocytosis?

Answer 49

The size of the mature red cell is NORMAL, but the MCV is measured as being high! Usually a fault with the analyser

Question 50

What are the two causes of spurious macrocytosis? (important to carry out blood film in diagnoses of macrocytosis for this reason) Focus on reticulocyte cause here

Answer 50

When there is an increased reticulocyte production by the bone marrow in response to haemorrhage or haemolysis Reticulocytes are bigger than mature red cells and are analysed along with these for the MCV measurement. Blood film will show polychromasia

Question 51

What are the two causes of spurious macrocytosis? (important to carry out blood film in diagnoses of macrocytosis for this reason) Focus on other cause here

Answer 51

Cold agglutins disease – immune system produces a protein causing the red cell to agglutinate and the cells are recognized as a single giant cellThe actual size of the individual red cells are completely normal

Question 52

Why is it called cold agglutination disease?

Answer 52

at body temperature, the antibodies do not attach to the red blood cells. At lower temperatures, however, the antibodies react to Ii antigens, bringing the red blood cells together, a process known as agglutination

Question 53

It is not uncommon for people with profound megalblastic anaemia to present with jaundice What are the megaloblastic, non-megaloblastic and spurious causes of macrocytic anaemia?

Answer 53

• Megaloblastic -B12/folate deficiency or drugs
• Non-megaloblastic - alcohol, liver disease, hypothyroidism, marrow failure (aplastic anaemia, myelodysplasia, myleoma)
• Spurious - erythroid hyperplasia producing more reticulocytes, cold-agglutination disease

After FBC- do reticulocytes

Question 54

Pancytopenia can complicate severe megaloblastic anaemia Nuclear maturation defects can affect multiple lineages What is pancytopenia?

Answer 54

Low red blood cells - eryhrocytopenia Low white blood cells - leucopenia Low platelets - thrombocytopenia