Identification of autoantibodies to glial fibrillary acidic protein in spinal cord injury patients

Abstract

Traumatic spinal cord injury (SCI) is a devastating injury causing significant morbidity and mortality. Experimental studies have demonstrated that SCI induced cellular damage and disruption of the blood-spinal cord barrier can initiate an autoimmune response. This response is thought to be pathogenic and contribute to poor outcome. The objective of this research was to investigate whether human SCI mounts an autoimmune response to self-antigens. Plasma samples were collected longitudinally from SCI patients (n=18) at acute (T1, <48 h) and subacute (T2, 2-4 weeks) time points to probe western blots of human brain homogenates in order to screen patients for the presence of putative autoantibodies. To identify the corresponding antigens, two-dimensional gel electrophoresis, western blot and liquid chromatography coupled with mass spectrometry (LC-MS/MS) analyses were performed. We found that four of 18 patients (22%) had novel immunoreactive bands ranging in size from 36 to 42 kDa present in subacute, but not in acute, plasma samples suggesting postinjury production. To identify the cross-reacting antigens, we separated brain proteins by two-dimensional gel electrophoresis and identified nine immunoreactive spots. Amino acid sequence analysis of these spots identified peptides that mapped to glial fibrillary acidic protein. Our results suggest that ∼ 22% of SCI patients generated autoantibodies to glial fibrillary acidic protein. Future studies will be required to determine whether these autoantibodies contribute to the pathogenic sequelae of SCI.

Figures

Figure 1

Western blots of human SCI plasma Time 1 and Time 2. Pictures of representative western blots probed with T1 (

Figure 2

Western blots and Coomassie stained gel showing increased immunoreactivity after SCI. Picture of a representative western blot from membranes probed with A. T1 plasma, and B. T2 plasma from a patient positive for autoantibodies. C. Coomassie-stained 2D gel from the same area as shown in the membranes. Spots excised for LC-MS/MS are indicated by numbers; * indicates location of β-actin. (T1, (