Red Cell parameters

what are the three parts of a haemotology report

erythron, leukon, thrombon,

what are we evaluating in a red cell assessment

1. mass

2. evidence for effective and appropriate eryhtropoeisis

3. size and variation

4. haemoglobinisation

5. shapes and inclusions

how is red cell mass assessed

PCV/Hct (Packed Cell Volume / Hematocrit): proportion of blood occupied by RBCs

RBCC (Red Blood Cell Count)

Hemoglobin (Hgb)

how is evidence for effective and appropriate erythropoeisis assessed?

§size and colour (MCV, MCHC)

§reticulocyte count

how is red cell size and variation assessed?

MCV (Mean Corpuscular Volume): average size of RBCs

RDW (Red cell Distribution Width): variation in RBC size (anisocytosis)

how is red cell haemoglobinisation (colour) assessed?

MCHC (Mean Corpuscular Hemoglobin Concentration): average hemoglobin concentration in a red cell

Hypochromic → pale cells (iron deficiency)

Normochromic → normal color

how is red cell shape and inclusions assessed?

smear

questions to ask when evaluating the erythron

• Is there inadequate, adequate or excessive red cell mass to deliver oxygen to tissues?

• Is there evidence of anaemia?

  • Is there evidence of regeneration?

  • What is the cellular character of the anaemia?

    • Normocytic/macrocytic/microcytic; normochromic/hypochromic

• Is there evidence of polycythaemia

  • Relative or absolute?

polycythemia

too many rbc

• can be relative or absolute

what is relative polycythaemia?

a reduced plasma volume makes it appear as though there are more red blood cells (higher concentration) despite normal RBC levels

what is absolute polycythaemia?

True increase in RBC mass due to increased RBC production/release

what are PCV, RBCC and Hgb used to measure?

three are measures of red cell mass and oxygen carrying capacity

Usually interpret them as a block

what are pcv rbcc and hgb all equally effected by?

haemoconcentration

• Will usually increase and decrease in line with one another

why would a PCV be wrong?

1. RBC's miscounted

• Mistaken for platelets

• Aggregated into pairs and triplets

2. MCV misleading

• Cell shrinkage or swelling

  • Transport, tube filling

  • Osmotic effects in machine

how to calculate PCV

MCV x RBCC

why is a high MCHC misleading?

§Not physiologically possible to cram more Hgb into red cells than they will take

what are causes of a high MCHC

§Haemolysis (sample handling or intravascular)

§Lipaemia

why may a MCV reading be misleading?

§Swelling from transport

§Mis-identification - pairs and triplets, cross over with large platelets

§Cell shrinkage or expansion in sample e.g. hyperosmolar

§Will impact on calculated PCV/HCT

how can haemolysis of a blood sample occur

• sample handled poorly or theres a problem

• cells release the haemoglobin

how can lipaemic samples interfere with MCHC

will interfere with light transmission through the sample and so will interfere with how we measure Hb

how can a rule of three error be picked up?

by looking at MCHC

Hct should roughly equal 3 × Hgb. If it doesn't, check for errors in the sample, the machine, or unusual red cell size/color.

Hct (%) approx. = Hgb (g/dL) x 3 (+/- 3%).

What is the classification of anaemia based on?

MCV and MCHC

What is a limitation of using MCV and MCHC for anaemia classification?

They are blunt measures.

We may see changes with the microscope that may not be sufficient to push parameter out of reference range

What can be more sensitive than MCV and MCHC in detecting changes in anaemia?

Machine dot-plots and histograms.

What type of anaemia is often classified as normocytic normochromic?

Anaemia of illness or pre-regenerative or occasionally non-regenerative.

What type of anaemia is classified as macrocytic hypochromic?

Classic highly regenerative anaemia.

• lots of young red cells coming in which are bigger but theyve got the right amt of Hb but bc its a bigger space the Hb is conc is less

What can sometimes cause macrocytic hypochromic anaemia aside from regeneration?

Cell swelling of transport.

What type of anaemia is classic for iron deficiency?

Microcytic hypochromic anaemia.

What condition can occur without anaemia related to microcytic hypochromic classification?

Portosystemic shunts.

what is polycythaemia

Increase in PCV, Hgb concentration and RBC count

What is relative polycythaemia?

PCV is increased but there is no increase in RBC production.

• Apparent increase in RBC due to a decrease in fluid in circulation (often ­ total protein and albumin)

• theres just less fluid

common causes of relative polycythaemia?

1. Dehydration (loss of plasma/water)

• Causes: Vomiting, diarrhoea, Polyuria, Extensive burns, Water deprivation or adipsia

2. Exercise, fear, excitement, severe pain - stress:

• Adrenaline secretion, splenic contraction and transient redistribution of RBC from the spleen to the circulation

how does relative polycythaemia resolve?

§Resolves after rehydration or removal of cause of splenic contraction

what is absolute polycythaemia

True increase in RBC mass due to increased RBC production/release (usu polychromasia, anisocytosis and reticulocytes)

• more rbcs produced

what does polycythaemia imply

increased number of several haemopoetic cell lines (human), however dogs & cats with polycythaemia vera usually have normal neutrophil & platelet counts!

what is primary polycythaemia? (polycytheamia vera)

• rare myeloproliferative disorder

• abnormal response of RBC precursors

• Normal EPO levels

what is secondary polycythaemia?

• Chronic tissue hypoxia of renal tissues (low arterial pO2) due to:

  • heart/lung diseases, high altitude, thrombosis, constriction of renal vessels

• Renal tumor or cysts [↑intra-capsular pressure]

• increased epo

how can the example of a kidney tumour cause secondary polycythaemia

• kidney has tight capsule

• turmour → pressure in capsule increases

• less blood gets in there

• sensors dont get the blood flow and o2 levels to think things are normal

• so release epo to ‘correct’ this

• polycythaemia

what are reticulocytes?

Young (immature/non-nucleated) erythrocytes prematurely released to blood from the bone marrow in regenerative anaemias.

how to visualise reticulocytes?

New methylene blue (NMB) precipitation demonstrates RNA-protein complexes (ribosmal RNA & mitochondria).

whats this

reticulocytes

• stained with new methylene blue → shows rna protein complexes

young red cells including reticulocytes have a what kind of appareance on romanowsky routine stain?

polychromatophil

clinical applications of reticulocytes

Evaluation of erythropoiesis in bone marrow.

Differentiation of regenerative and non-regenerative anaemia.

how to conduct a reticulocyte count?

Manual

Automated (some haematological analysers)

how to calculate absolute reticulocyte count (ARC)?

observed % reticulocytes x RBC (x10^12/l) x 10

Why ARC is important

Independent of variation of RBC numbers

More accurate than just the % reticulocytes:

A patient with anemia may have a high % reticulocytes, but the absolute number may be normal if RBC count is low

whats the difference between the 2 types of reticulocytes in cats,

difference is to do with a different pattern of dots in the red cell.

• aggregate: blue stained coarse clumping

• punctate: small, blue stained dots

What is the typical reticulocyte response in dogs?

Low number of reticulocytes (<1%)

What reticulocyte count is expected in regenerative anaemias in dogs?

At least (>60x10^9/L)

What is the typical reticulocyte response in cats?

Low number of reticulocytes (0.2-1.6%)

What are the two morphological types of reticulocytes in cats?

'Aggregate' and 'punctate'

What is the percentage of 'aggregate' reticulocytes in cats?

0.5% of erythrocytes

What is the percentage of 'punctate' reticulocytes in cats?

1-10% of erythrocytes

Which type of reticulocyte is considered in the assessment of regeneration in cats?

'Aggregate' reticulocytes

What reticulocyte count is expected in regenerative anaemia in cats?

At least (>50x10^9/L)

What is the typical reticulocyte response in ruminants and horses?

Virtually no reticulocytes in normal blood

Can reticulocytes appear in severe anaemias in horses?

No, reticulocytes may not appear even in very severe anaemias

When does peak reticulocyte production occur in cattle after acute blood loss?

7-14 days post acute blood loss

what does this show

Of the species we commonly deal with, dogs have probably

got the biggest red cells, whereas goats have got really,

really tiny ones.

But because there needs to be a certain amount of

haemoglobin to transport the right amount of oxygen across all

of these species, what we can recognise is that animals

that have got smaller volumes

will generally have larger numbers, so they all end up

with quite a similar pack cell volume.

describe the blood morphology in a dog

Larger erythrocytes

Uniform size

Central pallor

describe the blood morphology in a cat

Smaller erythrocytes

Mild anisocytosis (variation in size)

Scarce central pallor (less concave)

describe the blood morphology in a horse

Rouleaux

(sedimentationtendency)

describe the blood morphology in a ruminant

Anisocytosis and crenation (spikes)

what are the Variations within dog breeds

Macrocytosis in some poodles

Akitas have unusually small erythrocytes & particularly high potassium content

Greyhounds have high PCVs (0.55-0.6 L/L)

what is poikilocytosis

abnormally shaped RBCs

what can alterations in red cell shape indicate?

abnormal erythropoeisis

specific organ dysfunction

examples of common abnormal rbc shapes

• Codocytes (Target cells, Fe defic)

• Spherocytes (IMHA)

• Acanthocytes → irregularly spiked cells

• Schistocytes→ broken cells

• Echinocytes (Artefacts) crenation - 'burr cells' → regularly spiked cells

what are red cell inclusions?

abnormal material inside RBCs

what are examples of inclusions?

• Howell Jolly bodies → nuclear remnants

• Basophilic stippling → in cytoplasm

• Nucleated RBC's

• Infectious agents

  • Mycoplasma

  • Babesia

  • Viral inclusions

• Heinz bodies

describe the rbc

schistocytes

• RBC fragments

describe the rbc

acanthocytes

• irregular elongations of rbc border with sounded ends

describe the rbc

crenation/ echinocytes

• numerous pin point projections, even and regular

what are the types of red blood cell distribution

§Rouleaux formation

agglunination

explain the roleaux formation

§Clustering, sticky, piling of RBCs (look like stacks of coins)

§Normal finding in horses

what does the roleaux formation occur?

Relates to increased "stickiness" of plasma with increased globulin content

indicates inflam in SA

When does agglutination occur?

§Immune-mediated haemolytic anaemia

§Mismatched blood transfusion

how do you confirm if its agglutination

saline agglutination test:

Mix 1 drop of blood with 1 drop of saline

Æ Agglutination will persist, rouleaux formation will disperse