Localisation of citrullinated proteins in normal appearing white matter and lesions in the central nervous system in MS
Neurobiology and immunology:
There is increasing evidence that citrullination may play an important role in MS pathogenesis. Excess citrullination has been reported in the CNS in postmortem MS brain tissue.
Using myelin basic protein (MBP) isolated from normal appearing white matter (NAWM) from MS patients and controls, and fractionation of the samples by column chromatography, previous studies found that 18% of MBP was citrullinated in control tissue compared to 45% of MBP in patients with MS. Since citrullination alters the charge of the protein, citrullinated MBP becomes partially unfolded and its interaction with phospholipids is weakened, resulting in myelin sheaths that are not as tightly packed as in normal myelin.
The study investigated the immunohistochemical localisation of citrullinated proteins in post mortem brain tissue from MS patients and normal control cases. Nineteen blocks of snap-frozen autopsy tissue from 12 clinically and neuropathologically confirmed secondary progressive multiple sclerosis (SPMS) cases, together with nine blocks from six normal control cases, were obtained.
All of the patients had a confirmed diagnosis of MS by both histological and clinical criteria. Control and MS blocks were matched for CNS location. The study confirmed the citrullinated proteins present by western blotting as well as expression of PAD2 mRNA in the CNS by quantitative real-time PCR. PAD2 mRNA expression was also examined in in vitro studies of primary human astrocytes, a human foetal microglial cell line and a human brain endothelial cell line. Using these techniques, the authors demonstrated that increased citrullinated GFAP was found in areas of both ongoing demyelination and myelin loss in active and chronic active MS lesions. Interestingly, where there was complete myelin loss, citrullinated proteins were absent.
Lower levels of citrullinated proteins were observed in the MS NAWM and control white matter. Western blot analysis of brain tissue from these patients confirmed that in addition to MBP, GFAP was the major citrullinated protein in the CNS, and the amount of citrullination was increased in active disease, suggestive of a role in the pathogenesis of MS. However, in this study they were unable to identify a difference in the levels of citrullination between control white matter and MS NAWM compared to MS lesional brain tissue.
The presented data supports the hypothesis that deimination is important in the pathogenesis of MS, both in terms of citrullination of MBP preceding demyelination and citrullination of GFAP and its possible effects on astrocyte functions. Future work will involve investigating factors which increase PAD2 activity in astrocytes in vitro, as this will provide a better understanding of the mechanisms underlying the increased citrullination of GFAP and MBP seen in MS brain.
: Bradford CM, Ramos I
: J Neuroimmunol. 2014 May 27. pii: S0165-5728(14)00149-0. doi: 10.1016/j.jneuroim.2014.05.007. [Epub ahead of print]
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