Recovery from severe
spinal injury in adults is limited, compared to immature animals who demonstrate some capacity for repair. Using laboratory opossums (Monodelphis domestica), the aim was to compare proteomic responses to injury at two ages: one when there is axonal growth across the lesion and substantial behavioural recovery and one when no axonal growth occurs. Anaesthetized pups at postnatal day (P) 7 or P28 were subjected to complete transection of the spinal cord at thoracic level T10. Cords were collected 1 or 7 days after injury and from age-matched controls.
Proteins were separated based on isoelectric point and subunit molecular weight; those whose expression levels changed following injury were identified by densitometry and analysed by mass spectrometry. Fifty-six unique
proteins were identified as differentially regulated in response to spinal transection at both ages combined. More than 50% were cytoplasmic and 70% belonged to families of
proteins with characteristic binding properties.
Proteins were assigned to groups by biological function including regulation (40%), metabolism (26%),
inflammation (19%) and structure (15%). More changes were detected at one than seven days after injury at both ages. Seven identified
proteins: 14-3-3 epsilon, 14-3-3 gamma,
cofilin,
alpha enolase,
heart fatty acid binding protein (FABP3), brain
fatty acid binding protein (FABP7) and
ubiquitin demonstrated age-related differential expression and were analysed by qRT-PCR. Changes in
mRNA levels for FABP3 at P7+1day and
ubiquitin at P28+1day were statistically significant. Immunocytochemical staining showed differences in
ubiquitin localization in younger compared to older cords and an increase in oligodendrocyte and neuroglia immunostaining following injury at P28. Western blot analysis supported proteomic results for
ubiquitin and
14-3-3 proteins. Data obtained at the two ages demonstrated changes in response to injury, compared to controls, that were different for different functional
protein classes. Some may provide targets for novel drug or gene
therapies.