1. Anemia – What do you mean it’s not IMHA???
Jason M. Eberhardt DVM, MS, DACVIM

2. Overview One of the most common CBC abnormalities Back to basics
10-30% of patients
Why is it still so confusing?
Back to basics
Systematic approach to anemia
Avoiding common pitfalls

3. Some thoughts…
“You need to have the correct diagnosis before you can recommend the correct treatment.”
“If you always have the correct diagnosis then you’re not a really veterinarian…you’re probably a breeder.”
“You need to run a minimum of 5 diagnostic tests prior to starting steroids…”

4. Definitions Mean Corpuscular Volume (MCV) – Avg. RBC size
Macrocytosis
Microcytosis
Normocytic
Mean corpuscular Hgb concentration (MCHC) – [ ] of Hgb vol. RBC
Hypochromic
Normochromic
Macrochromic
Reticulocytes – Immature RBCs released from B.M. early
Normoblasts/metarubricytes – nucleated erythrocytes
Reduction in # of circulating RBCs, HCT and HB
Erythrocyte indices – remember they are MEAN calculations – a large # of cells have to be abnorm. To pull value out of ref. range – IMHA only 8.3 % are actually hypochromic on blood work
Macrochromic – almost all lab errors (due to heinz bodies or hemolysis or severe lipemia– people sickle cell anemia, hereditary spherocytosis
MCH – mean corpuscular hemoglobin Hgb x 10/PCV

5. Definitions continued…
Poikilocytosis – Variation of RBC shape
Rouleaux – Stacks of coins
Small amount is normal
Increased fibrinogen or acute phase proteins
Typically seen in inflammatory conditions
Autoagglutination – Aggregate in grapelike clusters
Must be differentiated from rouleaux
Rouleaux disperses when blood is mixed with saline
Spherocytes (RBCs lack central pallor near feather edge), Echinocytes, Acanthocytes (HAS, hepatic dz), Schistocytes (DIC, Fe def, CHF, myelofibrosis, HAS, malig. Histo)

6. Rouleaux or Autoagglutination

7. Before I go any further…
Where do I start…….
Back to basics!!!

9. The first step… Remember the Total Protein!!! DO NOT OVERLOOK!
It’s the other half of “blood”
It’s cheap!
It’s fast
DO NOT OVERLOOK!
Are just the RBCs being affected or the plasma as well?

10. The next steps… Morphologic classification Bone marrow response
RBC indices
Bone marrow response
Regenerative vs. Non-regenerative
Description of poikilocytosis?
Macrocytic, hypochromic, regenerative anemia with marked spherocytosis

11. Morphological classification
Usage of RBC indices (MCV/MCHC) to “describe” the RBCs.
Remember MCV/MCHC are MEAN calculations
Large # of RBCs affected prior to increases/decreases
Allows characterization of anemia into a category
Helps with ranking differential diagnoses
Are found on nearly all in-house CBC units
Normocytic, normochromic, Macrocytic hypochromic, Microcytic hypochromic

12. Normocytic normochromic
Most common
“Normal” RBCs
Most commonly denotes a non-regenerative anemia
Usually lacks RBC morphology changes
“Pre-regenerative”
First 1-3 days of acute loss/lysis
Total Protein can assist…

13. Macrocytic hypochromic
Usually indicates a regenerative anemia
Reticulocytes are relatively larger then mature RBCs
Hypochromic because Hgb synthesis is not complete
Only 8% of 6752 patients with reg. anemia had both increased MCV & decreased MCHC DiNicola et al.

14. Macrocytic normochromic
Usually misclassification due to insensitivity of MCV/MCHC
Autoagglutination?
Feline Leukemia
Poodles – Congenital dyserythropoiesis
Not anemic
Large problem in humans
B12 &/or folate deficiency
Role in veterinary medicine is questionable
FelV – usually no retics
Great Schnauzers

15. Microcytic hypochromic
Consistent with an iron deficiency anemia
Inadequate amount of Hgb is produced
Typically seen in chronic conditions
GI blood loss
Severe parasitism
PSS & Hepatic atrophy
Myelodysplastic syndromes
Congenital: Akitas, Shiba Inu, Chow breeds
Not typically hypochromic
Small RBCs (low MCV) Insufficient Hgb (low MCHC)
Young dogs – GI parasites; Old dogs – GI masses
Initially strongly reg. then non
Look at TP
Thrombocytosis can be marked

16. Bone marrow response Is there a regenerative response?
Evaluation of reticulocytosis
No reticulocytosis/polychromasia expected during first 1-3 days (maybe not at all if anemia stays mild)
Response peaks 4-5 days (with normal B.M.)
Erythrocyte indices start to change 7-14 days
Which species do not release produce reticulocytes (horses)

17. What is consider regenerative???
Normal patient should have <45,000-60,000 absolute retic count
Absolute counts
60, ,000 Early/mild response
150, ,000 Mild-moderate
>250, ,000 Moderate-Marked
Relative %
1-4 % - Mild
5-20 % - Moderate
> 20 % - Marked
Define relative

18. Regenerative anemia Loss vs. Lysis External blood loss
LOOK AT TOTAL PROTEIN!!!!
External blood loss
Low to low-normal T.P.
Hemolytic disease
High to high-normal T.P.

19. Acute external blood loss
PCV does not fully reflect severity first 1-3 days
Reticulocytosis should start by day 3
Peak reticulocytes day 4-7
PCV increases to low normal w/in 2 wks
May take up to 4-5 weeks to return to normal
Mild anemia does not stimulate strong erythropoietin release
Splenic contraction
Blood draws
OBVIOUS – Urinary, nasal, hemoabdomen, hemothorax

20. Chronic blood loss
Iron deficiency and negative protein balance develops after “several” weeks in adults
Occurs more rapidly in young animals (low iron stores)
Initially non/”pre” regenerative
Period of regenerative anemia depending on severity
Eventually returns to being poorly/non-regenerative
Often have thrombocytosis
Remember RBC indices do not change for 7-14 days
Getting blood transfusions???

21. Hemolytic anemia Hemolysis is a mechanism NOT a “disease”
Lots of “non” immune mediated causes
Low serum phosphorus
Normal to increased T.P.
Spherocytosis and/or autoagglutination
Over interpretation is common
Can be seen in diseases that are not “primary”
Positive Coomb’s Test?
Icterus occurs

22. Direct Coomb’s Test Identifies presence antibodies/compliment on RBCs
They may/may not actually be directed towards RBCs
This may/may not actually cause damage to RBCs
Neither highly specific or sensitive for IMHA
Positive in 60-70% of cases
Positive results – should have other evidence of IMHA
Effect of steroids?
**NOTE** – What is the end point of the test?????
FP seen in blood transfusions, drug reactions, RBC parasitic infections

23. Breaking it down…
Try to subclassify into intravascular vs. extravascular
Alters differential diagnosis
Intravascular – Rapid breakdown in vascular system
Pink urine, pink serum
Hemoglobinuria best indicator
Hyperbilirubinemia typically more profound then in extravascular
Extravascular – removal of RBCs by spleen, liver, B.M.
More common
Often has icterus, splenomegaly, hepatomegaly

24. Immune mediated “Immune-mediated” is a mechanism NOT a disease.
Can be 2nd to a number of possible causes
Infectious – Babesiosis, Ehrlichiosis, Leishmaniasis, Rickettsioses, Mycoplasma haemofelis, FeLV
Neoplasia
Drugs
Can be initially non-regenerative (esp. in cats)
Drugs: Sulfas, Carprofen, cephalosporins
Vax???
Look alikes – ie low phosphorus
Also snake envenomation

25. “Penny” 6 year FS Cocker
Presented for severe lethargy, “yellow skin” and “peeing blood”
Severe, macrocytic, normochromic strongly regenerative anemia with mild-moderate spherocytosis
Slide agglutination negative
High total protein
Abdominal ultrasound WNL
Infectious disease titers all negative

26. The “Penny” dilemma Needed multiple transfusion in a 5-6 day period
Continued to have hemolysis despite aggressive immunosuppressive therapy
Where do we go from here???
“Peeing” blood – hemoglobinuria
Intravascular hemolysis
Pred, cyclosporine, IVIG

27. Intravascular hemolysis
Immune mediated
Phosphofructokinase deficiency
Eng. Springers, Amer. Cockers
Babesia infection
Snake envenomation
Heavy metal to toxicity
Zinc
Copper
Zinc dogs can have spherocytes!!!!
PFK – Strenuous exercise with hyperventilation – leads to alkalosis, high pH inhibits PFK and RBC lysis

28. “Penny” 6 yr FS Cocker Spaniel
Presented for severe lethargy, yellow skin and “peeing blood”
Severe, macrocytic, normochromic strongly regenerative anemia with mild-moderate spherocytes
Abdominal ultrasound WNL
Infectious disease titers all negative

30. “Sheldon” 9 yr MC Jack Russell
Presented with clinical evidence of anemia
Severe leukocytosis (54,000), severe anemia (9%), high normal platelets, mild-moderate reticulocytosis
Total Protein – 4.9 g/dL
VF, Ehr. Neg.
Weakness, severe lethargy, decreased appetite
Pale mm, tachycardia, grade 2/6 systolic murmurs

31. IHMA??? Started on prednisone, cyclosporine, doxycycline
Needed 2nd transfusion 1 week later
Added azathioprine
PCV still low 2 weeks later
Chest rads and abd. u/s WNL
Increased prednisone, continued on cyclosporine and azathioprine
3rd transfusion in 4 weeks
Added leflunomide
Repeat abdominal ultrasound WNL

32. More anemia!!! Initial PCV/TP at EAC Reference lab work 12%/4.8
Hypoalbuminemia (2.6 g/dL), globulin WNL (1.7 g/dL), BUN increased (mild), Total bilirubin (mild)
Inflammatory leukogram
Severe reticulocytosis

33. What’s going on??? Horrible IMHA??? Another type of hemolytic anemia?
GI bleeding (from prednisone?, GI mass?)
Diagnostic plan???????????
Explain the decreased total protein

35. Non-regenerative anemia
Very common!!!
Usually normocytic normochromic
Microcytic, hypochromic anemias
Usually no poikilocytosis
Huge majority are mild-moderate in severity
2nd to systemic disease

36. Before going any further…
Is neutropenia and/or thrombocytopenia also present?
What is the duration of clinical signs?
How severe are the clinical signs?
Supports bone marrow disease

37. I need more RBCs… Mild-moderate NR anemia
Search for an underlying disease first
Anemia of chronic/inflammatory disease
Neoplasia, renal disease, hepatic disease, infectious, inflammatory, endocrine
Drugs
Phenobarb, sulfas, methimazole, chlorambucil, azathioprine

38. Severe non-regenerative anemia
Toxicity
Estrogen?
Drugs
Renal disease
More than just decreased erythropoietin
Chronic dz, decr. RBC lifespan, ineffective production, blood loss
Phenobarb, sulfas, methimazole

39. Why can’t it be easy??? Bone marrow exam Took a long time to develop
Can take even longer to resolve
Can still be very confusing
and frustrating

40. Bone Marrow disease Immune mediated
Maturation arrest vs. Pure Red Cell Aplasia
Myelophthisic syndromes - multiple cell lines often affected
Aplastic anemia – B.M. replaced by fat
Can be 2nd to chronic ehrlichiosis
Myelofibrosis – B.M. replaced by fibrous
Myelonecrosis – Drugs, toxins, viral
Neoplasia

41. “Howard” 9 yr MN DSH Progressive lethargy, wt. loss for several weeks
Marked (12%), macrocytic, normochromic anemia
Total protein 6.2 g/dL
Absolute reticulocyte count 40,000
Retic. total 2%
Corrected 0.65%
FelV/FIV negative
Chest radiographs, abdominal ultrasound WNL

42. Why cats are not small dogs…
50% of cats with immune mediated disease initially had a non-regenerative response Kohn et al. 2006
2/3 were <3 years (range was 1-9 yr)
Bone marrow disease – 53%
Infectious – 22%
Hemolysis – 11%
Immune Mediated – 6%
Severity of anemia associated with B.M. disease Korman et al. 2013

43. Bone marrow or bust Owner noticed gradual decline Transfusion
More consistent with non-regenerative disease
Transfusion
Recheck 2-3 days later vs. bone marrow now
Marked erythroid hypoplasia/aplasia
Immune mediated vs. FelV
Bone marrow IFA positive for FelV Stutzer et al. 2010

44. RBC shape descriptions
Many have little/no clinical significance
Anisocytosis, elliptocytes, codocytes, leptocytes, *echinocytes*
Spherocytes – Evidence of hemolysis
Acanthocytes - Hemangiosarcoma, hepatic dz
Schistocytes - DIC, Fe def, CHF, myelofibrosis, hemangiosarcoma, other neoplasia

45. Summary Anemia is a common abnormality
Cause can often be elusive
Vital to approach systematically
RBC indices, bone marrow response, poikilocytosis
DON’T FORGET THE TOTAL PROTEIN!!!

46. QUESTIONS???