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Characterizing the role of secretory phospholipase A₂ group IIA in glial cell-mediated neurotoxicity

University of British Columbia

Microglia are a type of non-neuronal glial cell that represent the mononuclear phagocyte system and innate immunity in the central nervous system. Astrocytes are another glial cell type. Under pathological circumstances, both glia types are capable of sustaining chronic inflammation in the brain, which results in neuronal death. Immune response involves glial secretion of pro-inflammatory mediators. Phospholipases A₂ (PLA₂) convert cell membrane glycerophospholipids into arachidonic acid, which in turn is a precursor to several pro-inflammatory eicosanoids that are released by glial cells. It is well established that secretory PLA₂ group IIA (sPLA₂IIA) is a pro-inflammatory mediator, however little is known about the role this enzyme plays in neurodegeneration. This thesis focuses on sPLA₂IIA and its role in chronic inflammation underlying neurodegenerative disorders such as Alzheimer’s and Parkinson’s diseases. Activated astrocytes have been shown to secrete sPLA₂IIA. However, no information is available on sPLA₂IIA expression and regulation in microglia. This thesis hypothesizes that sPLA₂IIA is a toxin secreted by activated glial cells, which causes neuronal death. If sPLA₂IIA contributes to neurotoxicity, then agents that inhibit, modify, or remove sPLA₂IIA from extracellular space should reduce the cytotoxic effects of glial secretions. The following human cells were used to study sPLA₂IIA expression, secretion and functions: microglia-like promonocytic THP-1 cells, U-373 MG astrocytoma cells, primary human astrocytes and neuroblastoma SH-SY5Y cells. Reverse transcriptase polymerase chain reaction (RT-PCR) revealed that stimulation by pro-inflammatory mediators induced sPLA₂IIA mRNA expression by glial cells. Stimulated glial cell supernatants were toxic to neuronal SH-SY5Y cells. Stimulation caused increased sPLA₂IIA protein concentrations in supernatants from all three types of glial cells. Despite observing increased sPLA₂IIA-specific enzymatic activity in stimulated glial cell supernatants, neither specific nor non-specific PLA₂ inhibitors exhibited anti-cytotoxic effects. However, the removal of sPLA₂IIA from supernatants by immunosorbents resulted in significantly increased neuronal survival, suggesting that sPLA₂IIA neurotoxicity relies on a non-enzymatic mechanism. Based on in vitro experiments and a literature review, potential sPLA₂IIA neurotoxicity mechanisms are proposed. The data obtained provide valuable information toward a more detailed mechanistic understanding of neuroinflammation and may guide future research toward more effective therapeutic agents for neurodegenerative disorders.

Text

2011-04-08

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/33425

Biology

University of British Columbia

UBCO

http://creativecommons.org/licenses/by-nc-nd/4.0/