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Alterations in the activity of phospholipases A2 in postmortem white matter from patients with multiple sclerosis
S J Huterer1, W W Tourtellotte, J R Wherrett
1Division of Neurology, University of Toronto, Ontario, Canada.
Abstract:
Activities toward arachidonyl-labelled phospholipase A2 substrates were assayed in fractions of white matter and cerebral cortex from control subjects and in fractions of demyelinated plaque, normal-appearing white matter and cerebral cortex from subjects who died with multiple sclerosis. Membranous activity at pH 8.6 in the presence of Ca2+, characteristic of 14 kDa "secretory" phospholipase A2, in either multiple sclerosis white matter or cortex did not differ from controls, whereas membranous activity at pH 4.5 in the absence of added Ca2+, characteristic of lysosomal enzymes was increased over controls in both plaque and normal-appearing white matter but not cerebral cortex. Activity in the cytosol fraction, at pH 8.6 in the presence of Ca2+ and glycerol characteristic of the "cytosolic" 85 kDa enzyme was decreased by greater than 50% in both white matter and cortex samples from multiple sclerosis subjects. Immuno-precipitation and -blotting confirmed that the deficient activity was largely attributable to the 85 kDa enzyme although the enzyme protein was not similarly reduced.
Insights
Multiple sclerosis (MS) alters phospholipase A2 enzyme activity in the brain. Cytosolic phospholipase A2 (cPLA2) activity decreased significantly in MS white matter and cortex, while lysosomal enzyme activity increased in affected white matter.
Area of Science:
- Neuroscience
- Biochemistry
- Enzymology
Background:
- Phospholipase A2 (PLA2) enzymes play critical roles in cellular signaling and membrane phospholipid metabolism.
- Dysregulation of PLA2 activity has been implicated in various neurological disorders, including demyelinating diseases like multiple sclerosis (MS).
- Specific PLA2 isoforms, such as the 14 kDa secretory PLA2 and the 85 kDa cytosolic PLA2 (cPLA2), exhibit distinct biochemical properties and cellular localizations.
Purpose of the Study:
- To investigate the activity profiles of different phospholipase A2 enzyme isoforms in the central nervous system of multiple sclerosis patients.
- To compare enzyme activities in affected (demyelinated plaque) and unaffected (normal-appearing white matter, cerebral cortex) tissues from MS subjects with control tissues.
- To determine the contribution of specific PLA2 isoforms to observed enzymatic changes in MS.
Main Methods:
- Assay of arachidonyl-labelled phospholipase A2 substrates in subcellular fractions (membranous and cytosol) from human brain tissue.
- Enzyme activity measurements were performed under specific conditions (pH, presence of Ca2+, glycerol) to differentiate between PLA2 isoforms.
- Immuno-precipitation and immuno-blotting techniques were used to confirm the identity and quantify the protein levels of the affected enzyme.
Main Results:
- Membranous activity characteristic of 14 kDa secretory PLA2 did not differ between MS and control subjects.
- Membranous activity characteristic of lysosomal enzymes (pH 4.5, no added Ca2+) was elevated in demyelinated plaque and normal-appearing white matter from MS subjects compared to controls.
- Cytosolic activity, characteristic of 85 kDa cPLA2 (pH 8.6, Ca2+, glycerol), was significantly decreased (over 50%) in both white matter and cerebral cortex of MS subjects, despite comparable protein levels of the 85 kDa enzyme.
Conclusions:
- The study reveals distinct alterations in PLA2 enzyme activities in the multiple sclerosis brain.
- Lysosomal enzyme activity is upregulated in white matter affected by MS, potentially reflecting inflammatory or repair processes.
- A significant reduction in 85 kDa cPLA2 activity, not explained by reduced enzyme protein, suggests post-translational modifications or regulatory changes affecting enzyme function in MS.