1. Hepatic cases Kathleen Tennant BVetMed Cert SAM Cert VC FRCPath MRCVS
Clincial Lead
Diagnostic Laboratories
Langford Veterinary Services

3. Case 1 2 year old Norfolk terrier
Presented depressed, vomiting and lethargic after scavenging in dustbin including baby nappies, chocolate cake and sugar free gum
Clinical exam – slightly icteric

4. Haematology Haemoconcentrated (PCV 59 L/L)
Platelet count through the machine 100 (200 – 500 x 10^9/L)
Leucocytes unremarkable (numbers and morphology)

5. Test
Result
Range
Units
Total protein 65
g/L
Albumin 30
Globulins 35
Sodium
155
mmol/L
Potassium
4.0
4.1 – 5.3
Chloride
107
Calcium
2.3
2.13 – 2.7
Inorganic phosphorous
3.2
Urea
9.3
3 – 9.1
Creatinine
162
Umol/L
Cholesterol
9.0
3.3 – 8.9
Total bilirubin 16
0 – 2.4
AST
402
0 - 28
U/L
ALT 4
ALP
2900

6. How do we best interpret the ALT value?
Rules out hepatocellular damage 1.
Low due to lack of remaining hepatic parenchyma
Likely to be laboratory or technical error 3.
There was hepatocellular damage, but this is in the past, hence low ALT but AST still elevated

7. Test
Result
Range
Units
Total protein 65
g/L
Albumin 30
Globulins 35
Sodium
155
mmol/L
Potassium
4.0
4.1 – 5.3
Chloride
107
Calcium
2.3
2.13 – 2.7
Inorganic phosphorous
3.2
Urea
9.3
3 – 9.1
Creatinine
162
Umol/L
Cholesterol
9.0
3.3 – 8.9
Total bilirubin
8.2
0 – 2.4
AST
402
0 - 28
U/L
ALT 4
ALP
2900

8. Internal consistency
AST and ALT often both elevate with hepatocellular damage, with 2 x top of reference interval elevations mild, 4 x moderate and 10 x taken as marked
It would be unusual for ALT to be below reference if AST is this elevated in hepatocellular damage as AST generally elevates with more, not less, damage.
The half life of AST is actually shorter than that of ALT (< 1 day vs 2 -3 days), so severity and timing play a role in the final values
Low ALT due to absence of remaining hepatocytes extremely unusual, and not expected if AST still high
AST elevation without ALT may be due to muscle damage
No history or suspicion of muscle damage – could use creatine kinase to confirm its presence or absence
? Falsely low ALT

9. Trouble - shooting single dubious results
Check the gross appearance of the sample or machine flags for hemolysis, icterus or lipaemia if these interfere with the assay
This sample is icteric – does not affect ALT but:
Inorganic phosphorous 3.2 (ref mmol/L) – is affected by icterus – reject result

10. Trouble - shooting single dubious results (ALT)
Check the quality assurance/ quality control for that analyte
Usually by checking the machine’s log for low, medium and high control performance
Should indicate if there is a problem with reagents, and can be run again if needed to check for a machine glitch (e.g. bubble in the system)
Check the results of samples either side of this one
In this case – all fine, so re – run to rule out a ‘pipetting’ error
Also run in x 10 and x 100 dilution

11. ALT re-run results Neat sample – ALT 0 (13 – 88 U/L)
X 10 dilution - ALT 3502 (13 – 88 U/L)
X 100 dilution – ALT 3560 (13 – 88 U/L)
Some wet chemistry dynamic assays, confronted with extremely high activity in their target enzyme will ‘miss’ the activity because it’s all over too quickly to measure – default to 0 or a very low level

12. Falsely low readings with high value rate of change assays
Substrate depletion
Substrate depletion
Absorbence
Absorbence
delay
delay
measure
measure

13. Test
Result
Range
Units
Total protein 65
g/L
Albumin 30
Globulins 35
Sodium
155
mmol/L
Potassium
4.0
4.1 – 5.3
Chloride
107
Calcium
2.3
2.13 – 2.7
Inorganic phosphorous
N/A
Urea
9.3
3 – 9.1
Creatinine
162
Umol/L
Cholesterol
9.0
3.3 – 8.9
Total bilirubin 16
0 – 2.4
AST
402
0 - 28
U/L
ALT
3502
ALP
2900

14. What else SHOULD have been measured at the beginning, given the clinical history?
Presented depressed, vomiting and lethargic after scavenging in dustbin including baby nappies, chocolate cake and sugar free gum
Clinical exam – slightly icteric

15. What else SHOULD have been measured at the beginning, given the clinical history?
Bile acids
Glucose
PT/APTT
D – dimers
Ammonia

16. Xylitol toxicity Marked elevation in insulin following ingestion
Initial marked hypoglycaemia may lead to some of the central clinical signs
Hepatocellular necrosis, development of D.I.C./ bleeding disorders
Marked prolongations in PT and APTT and evidence of bleeding disorder, including petechiation, G.I. bleeding. D – dimers likely to elevate.
Although they are likely to be abnormal, bile acids and ammonia should not take precedence over glucose and assessing clotting ability.

17. Case 2 8 y.o. FN Irish Wolfhound
3 week history of vague lethargy, inappetance and intermittent diarrhoea
Owners noted a bloated abdomen that morning, followed by collapse – rushed in

18. Clinical findings Poor body condition (3/9)
Fluid thrill in abdomen – structures could not be palpated
Tachycardic, bounding pulses

19. Haematology – what describes the red cell picture?
Hb (12 – 18 g/dl)
HCT 27 (35 – 55 %)
RBC 5.0 (5.4 – 8.0 x 10^12/L)
MCV 64 (65 – 75 fl)
MCH 21 (22 – 25 pg)
MCHC 32.8 (34 – 37 g/dl)
This slide takes a while to look at.

20. What best describes the red cell picture?
Mild normochromic anaemia 1.
Moderate normochromic normocytic anaemia
Mild hypochromic normocytic anaemia 3.
Mild hypochromic microcytic anaemia 4.
Mild regenerative anaemia 5.

21. Haematology Hb 10.00 (12 – 18 g/dl) Mild anaemia HCT 27 (35 – 55 %)
RBC 5.0 (5.4 – 8.0 x 10^12/L)
MCV 64 (65 – 75 fl) Microcytic
MCH 21 (22 – 25 pg)
MCHC 32.8 (34 – 37 g/dl) Hypochromic

22. Smear examination – red cell morphology

23. What best describes the red cell morphology on the smear?
Unremarkable
Microcytic, hypochromic 2.
Echinocytes
Acanthocytes
Schistocytes

24. Erythrocytes in liver dysfunction
Poor iron handling compromises erythroid precursor haemoglobin manufacture – less haemoglobin per cell
Slowness in achieving a haemoglobin content threshold allows an extra precursor division, resulting in smaller cells
These will generally be picked up by the machine BEFORE they are obvious on the smear
No evidence of regeneration in this population – no obvious polychromatophils

25. Acanthocytes and schistocytes
Acanthocytes (black arrows) are red cells with large cytoplasmic projections. These are associated with liver diseases, lipid disorders and shear injury to the red cells
Schistocytes (red arrow) associated with shear injury to red cells

26. Test
Result
Range
Units
Total protein 65
g/L
Albumin 26
Globulins 20
Sodium
146
mmol/L
Potassium
4.5
4.1 – 5.3
Chloride
107
Calcium
2.2
2.13 – 2.7
Inorganic phosphorous
1.2
Urea
7.8
3 – 9.1
Creatinine
158
Umol/L
Cholesterol
7.2
3.3 – 8.9
Total bilirubin
3.3
0 – 2.4
AST
210
0 - 28
U/L
ALT
880
ALP
810

27. Is this more a cholestatic or a hepatocellular damage picture?
Both exactly the same 3.

28. Test
Result
Range
Units
Total protein 65
g/L
Albumin 26
Globulins 20
Sodium
146
mmol/L
Potassium
4.5
4.1 – 5.3
Chloride
107
Calcium
2.2
2.13 – 2.7
Inorganic phosphorous
1.2
Urea
7.8
3 – 9.1
Creatinine
158
Umol/L
Cholesterol
7.2
3.3 – 8.9
Total bilirubin
3.3
0 – 2.4
AST
x ref
0 - 28
U/L
ALT
x ref
ALP
810 3 x ref

29. Abdominal fluid – sampled through midline
PCV 18 %
Plt 5 x 10^9/L
Nucleated cell count 3 x 10^9/L
Total protein 30 g/L

30. Abdominal effusion – elsewhere on the slide

31. What best describes the nature of the fluid?
Transudate
Modified transudate 2.
Exudate
Iatrogenic haemorrhage superimposed on a transudate
Haemorrhagic

32. Abdominal fluid – sampled through midline
PCV 18 %
Plt 5 x 10^9/L
Nucleated cell count 3 x 10^9/L
Total protein 30 g/L
Transudates TP < 25 g/L, nucleated cell count < 1 x 10^9/L
Not expected here as serum albumin 26 g/L
Previous and current haemorrhage….

33. Imaging - appearance of liver

34. Coagulation testing
PT patient 9 secs , control 9 secs (expected < 11 s)
APTT patient 25 secs, control 18 secs (expected < 21 s)
D – dimers 0.5 (0 – 2 mg/dl)

35. Which of these fits best with the coagulation test results?
Coumarin toxicity
Liver dysfunction
Hemophilia A
Hemophilia B
Consumptive coagulopathy (prolongation due to factors being used up attempting to stop bleeding from another cause) 5.

36. Coagulopathies and liver disease
May be compromise in absorbing Vitamin K and other fat soluble vitamins
Many clotting factors made by liver
In some animals with liver disease, APTT may prolong ahead of PT
Hemophiliacs often have very prolonged APTT values
Can not rule out a consumptive coagulopathy, although often PT and APTT might be expected to prolong together
Has anyone noticed we haven’t looked at platelets yet?

37. Platelet count through machine = 60 x 10^9/L, smear estimate 40 – 60 x 10^9/L Likely consumption….

38. Hepatic cytology 100x magnification

39. Hepatic cytology
Normal
Patient

40. Hepatic cytology 500x

41. What is the cytology most compatible with?
Chronic hepatitis
Large cell lymphoma 2.
Haemangiosarcoma 3.
Hepatoma
Hepatic carcinoma

42. Final diagnosis: Hepatic carcinoma - ruptured

43. Cytology correlation with histopathology
Wang et al (2004): 94 cases, dogs and cats – overall agreement between histopathology and cytology in 30% canine and 51% of feline samples. Best agreement in vacuolar hepatopathy (7/11 dogs, 15/18 cats), poor correlation for inflammation (5/20 dogs, 3/11 cats)
Kristensen et al (1990): agreement in 66% between histo/ cyto in 44 cases

44. Case 3

45. Case 3 9 y.o. N.F. miniature Dachshund
Intermittent lethargy, abdominal pain, vomiting
Recent rapid deterioration – collapse, pyrexia, dehydration, icterus

46. Haematology WBC 35 (5.5 – 17 x 10^9/L)
Neutrophils 32 (3.0 – 11.5 x 10^9/L)
Other findings through machine unremarkable
Smear shows left shift and toxic change in neutrophils

47. Neutrophils

48. Test
Result
Range
Units
Total protein 84
g/L
Albumin 26
Globulins 58
Sodium
149
mmol/L
Potassium
5.1
4.1 – 5.3
Chloride
111
Calcium
2.3
2.13 – 2.7
Urea
9.3
3 – 9.1
Creatinine
162
Umol/L
Cholesterol
13.2
3.3 – 8.9
Total bilirubin 54
0 – 2.4
AST 45
0 - 28
U/L
ALT
210
ALP
1283
GGT 56
0 - 15

49. Which of these is true?
ALP is a more sensitive or equally sensitive marker compared to GGT for cholestasis in the dog
ALP is a more sensitive marker than GGT for cholestasis in the cat 2.
Bilirubin is a specific marker for cholestasis in the dog and cat 3.
Cholesterol decreases in cholestatic disease

50. Markers for cholestasis
ALP is a sensitive marker for cholestasis in the dog, but in the cat, GGT tends to increase more in hepatic/cholestatic conditions other than hepatic lipidosis
Due to potential elevations derived from increased haemolysis, bilirubin is not specific for cholestasis in the dog or cat
Cholesterol increases in cholestasis (hence the name!)

51. Further testing Canine Pancreatic Lipase within normal limits
PT/ APTT normal

52. Extracellular bile pigment

53. Gall bladder contents

54. Gall bladder contents

55. Final diagnosis
Pure growth of E coli cultured from gall bladder contents aspirate, and from biopsies of liver parenchyma
Ascending cholecystitis/ infection