Resp Lecture 1

Anaemia is defined as decreased erythrocyte mass

Free haemoglobin in the plasma (so when you spin down the blood, you may notice a red tint to the plasma)

EPO is the principle growth factor promoting viability, proliferation, and differentiation

Adults-from the kidney and to a lesser degree the liver

Foetus- from the liver

hormones and other growth factors, low blood volume

TNF-alpha, IL-1, IFN, TGF-beta

These are all inflammatory mediators, they inhibit EPO release and its activity on RBC, hence why animals with inflammation will often become anaemic

Start with a pluripotent stem cell-->myeloid stem cel (committed cell) --> erythroid CFU --> primitive/mature --> rubriblast--> prorubricyte--> rubricyte--> metarubricyte-->chromatophil-->erythrocyte

All of this occurs within the marrow.

Pluripotent stem cell--> myeloid stem cell (committed cell)--> megakaryoblast-->megakaryocyte--> platelets

undifferentiated mother cell for all the RBCs. Rubriblasts, prorubricyte, and rubricytes can all continue to divide to make more cells, particularly under the influence of EPO. When the Hb reaches a desirable threshold, it signals to the rubricyte to stop dividing and mature.

Metarubricytes, polychromatophils, and mature erythrocytes can no longer divide. The nucleus shrinks down and the Hb builds up in the cell. The cytoplasm turns from blue to pink. Finally, when the nucleus is shrunk down, the macrophages take out the nucleus and donates an iron to make the Hb. This process takes 3-5D

Decreased haematocrict/PCV

Decreased RBC/L

Decreased Hb level

if any of the following are low, then the pt is said to be anaemic, resulting in decreased O2 carrying capacity

RBC loss (haemorrhage, haemolysis)

or decreased RBC production

There's a marrow response

Evidence: reticulocytosis, polychromasia, hypochromic macrocytic

No marrow response

Pre-regenerative (marrow has not had sufficient time to respond)

or non-regenerative-> no reticulocytosis, minimal polychromasia

Normocytic normochromic or hypochromic microcyctic

Blood smear: look for polychromasia

Reticulocytosis: done with a analyzer, most accurate means of assessment, lets us known the levels of immature RBCs in blood.

We'll also see evidence of macrocytosis and hypochromasia
Normocytic normochromic anaemia does NOT mean non-regenerative, may be pre-regenerative

Bone marrow evaluation: Erythroid hyperplasia

So we'll perform serial monitoring of PCV/CBC to assess improvement

Polychromasia (large blue cells)

Macrocytosis

anisocytosis

Hypochromasia

Increased howell-jolly bodies

Increased nucleated RBCs

Increased levels of basophilic stippling

Manually using a new methylene blue stain--> equal parts blood and stain are mixed and then incubated 10-15 minutes before smear is made. Count the number of reticulocytes per 1000 RBCs

Analyzer

Aggregate and punctate (more mature reticulocytes). In all other species besides cats, the maturation is fast and won't tell us anything.

Polychromatophils released in low numbers in dogs (1%) and cats (.4%). However, in active regenerative anaemia you will see these numbers increase. Aggregates tell us what the marrow did recently.

More mature form of reticulocyte. Reticulocytes take up to 2 weeks to mature in cats, so up to 10% punctate reticulocytes are seen in health (not very useful diagnostically)

Corrected reticulocyte %

Absolute count

Reticulocyte % x RBC

If we determine the % of reticulocytes then we need to determine if this is normal. Perform corrected reticulocyte %

Retic % x (pt hct/ avg species Hct)

Average species Hct in dog: .45, cat:.37

CRP 0-1% in health

We want higher counts with more severe anaemia otherwise considered non-regenerative

healthy birds will have 4-5%

Mean Corpuscular Volume (how much space do the cells take up)

It's the average you can take, you can have quiet a few immature cells in circulation and it still won't take the average size above normal. So sometimes animals with regenerative anaemia will have normal average cell volume; they may not show up as macrocyte.

Mean corpuscular Hb concentration

Amount of Hb per average cell.

Increase is not possible, hyperchromic indicates either free Hb or lipaemia.

Red cell distribution width

Measures the variation in cell size

An increase in RDW indicates anistocytosis

Per-acute

Acute stage I

Acute stage II

No change in the Hct or the TP (just started to Haemorrhage, so we lose a lot of plasma, so we won't notice the change in the density of the blood).

Stage I within hours, decrease Hct and TP as the fluid shifts from extravascular to the intervascular space, activation of RAAS.

Pre-regenerative anaemia

Within 3-5d.

EPO is produced (stimulated by hypoxia). Evidence of regeneration in blood (polychromesia, macrocytes, reticulocytes)

Increase in Hct and protein IF haemorrhage is controlled.

If we lose bood we lose plasma. So after an animal has haemorrhaged, the first thing we check is the protein level. If this has dropped, then we need to be concerned about haemhorrage. If not, then look for alternatives to RBC loss.

Intravascular (rupture of RBC in circulation) and extravascular (phagocytosis of RBC by macrophages in spleen marrow and liver)

Within hours, decreased Hct with normal protein, no evidence of regeneration (pre-regenerative), +/- haemoglobinuria and haemoglobinuria (if filtered by the kidneys).

within 3-5D, EPO is produced, evidence of regeneration in the blood. If under control, Hct increases.

If ongoing, bone marrow may not meet destruction, same can occur with haemhorrage.

Often see hyperbilirubinaemia/jaundice (as the RBC is broken down, bilirubin production increases bc increased Hb recycling)

Spherocytosis (extravascular)

Schistocytes (irregular shapes)

Keratocytes (tags hanging off) if microvascular fragmentation.

Normal protein levels.

IMHA, neonatal isoerythrolysis, transfusion reaction (disease)

Oxidative injury
Infections

Severe hypophosphatemia (phosphorous is essential in RBC metabolism, decreased levels may make RBC easy target for macrophages)
Zinc and copper toxicosis

Genetic disorders

Within the bone marrow, spleen, or induced via macrophages.

Immune mediated disease (primary or secondary , e.g. drugs)

Infections

Oxidative damage

neoplasia

Fragmentation (e.g. DIC, haemangiosarcoma, heart worm)

Genetic disease

Primary is idiopathic autoimmune, common in dogs.

Secondary is induced by drugs and vaccinations, infections, neoplasia; more common in cats.

Antibody against foreign RBC, antigen of same species (neonatal isoerythrolysis-more common in horses), blood transfusions.

Antibody or drug or parasite can bind to the RBC. Macrophages will identify and either destroy the cell (more common) or take a small chunk out, RBC becomes spherocyte. Clue of extravascular anaemia.

Large antibodies may bind more then one RBC causing agglutionation. Need a positive saline test to differentiate between that and rouleax.

Antibody or drug or parasite may bind, activating classical complement pathway, MAC formation and cell lysis, resulting in ghost cells. Indicative of intravascular haemolysis. RBC ruptures and we get membrane and free Hb. Won't see free Hb in extravascular haemolysis domination, Hb will go to macrophages.

presence of spherocytosis or ghost cells
Autoagglutination
Coombs test allows us to detect agglutination by detecting auto-antibodies.
Exclusion of other primary diseases.

D.

Unconjugated bilirubin is a byproduct of erythrocyte breakdown.

D. Reticulocytosis