Early upregulation of cytosolic phospholipase A2α in motor neurons is induced by misfolded SOD1 in a mouse model of amyotrophic lateral sclerosis

Abstract

Background: Amyotrophic lateral sclerosis (ALS) is a fatal multifactorial neurodegenerative disease characterized by the selective death of motor neurons. Cytosolic phospholipase A₂ alpha (cPLA₂α) upregulation and activation in the spinal cord of ALS patients has been reported. We have previously shown that cPLA₂α upregulation in the spinal cord of mutant SOD1 transgenic mice (SOD1^G93A) was detected long before the development of the disease, and inhibition of cPLA₂α upregulation delayed the disease's onset. The aim of the present study was to determine the mechanism for cPLA₂α upregulation.

Methods: Immunofluorescence analysis and western blot analysis of misfolded SOD1, cPLA₂α and inflammatory markers were performed in the spinal cord sections of SOD1^G93A transgenic mice and in primary motor neurons. Over expression of mutant SOD1 was performed by induction or transfection in primary motor neurons and in differentiated NSC34 motor neuron like cells.

Results: Misfolded SOD1 was detected in the spinal cord of 3 weeks old mutant SOD1^G93A mice before cPLA₂α upregulation. Elevated expression of both misfolded SOD1 and cPLA₂α was specifically detected in the motor neurons at 6 weeks with a high correlation between them. Elevated TNFα levels were detected in the spinal cord lysates of 6 weeks old mutant SOD1^G93A mice. Elevated TNFα was specifically detected in the motor neurons and its expression was highly correlated with cPLA₂α expression at 6 weeks. Induction of mutant SOD1 in primary motor neurons induced cPLA₂α and TNFα upregulation. Over expression of mutant SOD1 in NSC34 cells caused cPLA₂α upregulation which was prevented by antibodies against TNFα. The addition of TNFα to NSC34 cells caused cPLA₂α upregulation in a dose dependent manner.

Conclusions: Motor neurons expressing elevated cPLA₂α and TNFα are in an inflammatory state as early as at 6 weeks old mutant SOD1^G93A mice long before the development of the disease. Accumulated misfolded SOD1 in the motor neurons induced cPLA₂α upregulation via induction of TNFα.

Keywords: ALS; Cytosolic phospholipase A2α; Misfolded SOD1; Motor neurons; Mutant SOD1G93A; TNFα.