Diagnostic testing for muscle disease Flashcards
What are the non-specific markers of muscle damage? What does their presence indicate? How are their kinetics different?
- Creatine Kinase (CK) and Aspartate Aminotransferase (AST)
- These enzymes are normally found intracellularly within muscle, their appearance in the systemic circulation represents rupture of cells -> damaged tissues
- Measured in terms of ACTIVITY not CONCENTRATION (same applies to hepatic enzymes)
- CK activities peak early (6 hours) and return to normal quickly (24 hours)
- AST activities increase more slowly (18-24 hours) and return to normal slowly (20 days)
- We can use this relationship to understand the chronicity of disease, and to identify ongoing muscle damage, especially with serial measurements at different timepoints.
What would be a significant finding in a dynamic CK testing? How is this undertaken?
Sample 2 ≥ 2 x Sample 1
(even if both are within the reference range!)
Systemic enzyme response to a bout of sub-maximal exercise (15-20 minutes of trotting on the lunge is sufficient for most horses)
How is urine a useful muscle marker?
Pigmenturia is another non-specific sign of muscle damage:
- Cell rupture -> myoglobin into circulation -> filtration by kidney -> PIGMENTURIA
The presence of dark red urine should alert the clinician to potential myopathy but needs to be differentiated from diseases leading to haematuria (blood contaminated urine)
- Usually by simply measuring muscle enzyme activities!
Myoglobin is highly nephrotoxic – indication for fluid therapy!
What are the indications and contraindications for muscle biopsy?
Indications for muscle biopsy:
* Repeated bouts of clinical disease, poorly managed
* Cases with doubling of baseline CK activity
* Suspicion of underlying storage myopathy
* If seeking a definitive diagnosis
* Poor performance work-up with suspicion of muscle disease
Contraindications for muscle biopsy:
* The diagnosis is clearly evident (atypical myopathy)
* Condition is successfully managed with husbandry changes and symptomatic treatment (equine rhabdomyolysis)
When investigating exertional myopathies, what muscle should be biopsied? What is the procedure?
Biopsy semimembranosus muscle
- Contact your laboratory (RVC) with at least 48 hours notice!!
- Sedate horse, restrain in stocks if possible
- Inject local anaesthetic subcutaneously – avoid injecting muscle!
- Aseptically prepare the skins with chlorhexidine and surgical spirit
- Make a linear incision (4cm) and retract skin with gelpi retractors
- Make two parallel incisions (3cm) into the muscle tissue 1cm apart and 1cm deep, following the direction of muscle fibres
- Join your two incisions at the proximal end and then slowly undermine the length of your incision, eventually transecting the distal end
- Place the sample on damp gauze while you close dead space and suture the skin
- Divide the sample in half – one part should be stretched onto a piece of card and preserved in formalin, the other half should be sent fresh.
What will the lab do with biopsy samples?
Periodic acid Schiff (PAS) straining for PSSM
* Amylase digestion first removes all normal accumulations of glycogen
* Dark staining cells from affected horse (right) denoting abnormal accumulations of resistant glycogen
Haematoxylin and eosin (H&E) staining suggestive of RER
* Internalised nuclei (should be peripheral), marked variation in fiber sizes, infiltration of macrophages (black arrow)
* Caffeine sensitivity is also a feature – performed on fresh samples
Desmin stain for myofibrillar myopathy
* Abnormal accumulations of desmin in an affected horse (left) compared to a normal desmin distribution (right)
What genetic testing is available for muscle diseases?
PSSM1 – validated genetic testing available
* mutation in glycogen synthase (GYS-1) gene
* Widely available, samples include blood and hair plucks (including follicles)
PSSM2 – many unvalidated tests available (group of undiagnoses diseases that look like PSSM1)
* There is no evidence for the use of genetic testing with other storage myopathies
What atypical myopathy toxin testing is available?
Hypoglycin-A (HGA) and its toxic metabolite methylenecyclopropylacetic acid (MCPA)
Principle metabolite causing AM
Submit whole blood or spun serum sample
What tests would you run to diagnose atypical myopathy, recurrent equine rhabdomyolysis, PSSM1 and PSSM2?
X - would not complete diagnostic test
What results would we expect when running diagnostic tests for atypical myopathy, recurrent equine rhabdomyolysis, PSSM1 and PSSM2?
