No abstract

Double-seronegative myasthenia gravis (dSN-MG, without detectable AChR and MuSK antibodies) presents a serious gap in MG diagnosis and understanding. Recently, autoantibodies against the low-density lipoprotein receptor-related protein 4 (LRP4) have been identified in several dSN-MG sera, but with dramatic frequency variation (∼2-50%). We have developed a cell based assay (CBA) based on human LRP4 expressing HEK293 cells, for the reliable and efficient detection of LRP4 antibodies. We have screened about 800 MG patient sera from 10 countries for LRP4 antibodies. The overall frequency of LRP4-MG in the dSN-MG group (635 patients) was 18.7% but with variations among different populations (range 7-32.7%). Interestingly, we also identified double positive sera: 8/107 anti-AChR positive and 10/67 anti-MuSK positive sera also had detectable LRP4 antibodies, predominantly originating from only two of the participating groups. No LRP4 antibodies were identified in sera from 56 healthy controls tested, while 4/110 from patients with other neuroimmune diseases were positive. The clinical data, when available, for the LRP4-MG patients were then studied. At disease onset symptoms were mild (81% had MGFA grade I or II), with some identified thymic changes (32% hyperplasia, none with thymoma). On the other hand, double positive patients (AChR/LRP4-MG and MuSK/LRP4-MG) had more severe symptoms at onset compared with any single positive MG subgroup. Contrary to MuSK-MG, 27% of ocular dSN-MG patients were LRP4 antibody positive. Similarly, contrary to MuSK antibodies, which are predominantly of the IgG4 subtype, LRP4 antibodies were predominantly of the IgG1 and IgG2 subtypes. The prevalence was higher in women than in men (female/male ratio 2.5/1), with an average disease onset at ages 33.4 for females and 41.9 for males. Overall, the response of LRP4-MG patients to treatment was similar to published responses of AChR-MG rather than to MuSK-MG patients.

Japanese encephalitis virus is a mosquito-borne flavivirus that causes approximately 10000 deaths annually in Asia. After a brief viraemia, the virus enters the central nervous system, but the means of crossing the blood-brain barrier is uncertain. We used routine histological staining, immunohistology and electron microscopy to examine brain material from four fatal human cases, and made comparisons with material from a mouse model. In human material there was oedema, perivascular inflammation, haemorrhage, microglial nodules and acellular necrotic foci, as has been described previously. In addition, there was new evidence suggestive of viral replication in the vascular endothelium, with endothelial cell damage; this included occasional viral antigen staining, uneven binding of the vascular endothelial cells to Ulex europaeus agglutinin I and ultrastructural changes. Viral antigen was also found in neurons. There was an active astrocytic response, as shown by glial fibrillary acidic protein staining, and activation of microglial cells was demonstrated by an increase in major histocompatibility complex class II expression. Similar inflammatory infiltrates and a microglial reaction were observed in mouse brain tissue. In addition, beta-amyloid precursor protein staining indicated impaired axonal transport. Whether these findings are caused by viral replication in the vascular endothelium or the immune response merits further investigation.

Objective: To determine whether the activation of innate immune responses, which can be elicited by pathogenic and endogenous triggers, is associated with the presence of Epstein-Barr virus (EBV) infection in the multiple sclerosis (MS) brain. Methods:White matter postmortem MS (n ϭ 10) and control tissue (n ϭ 11) was analyzed for the expression of the proinflammatory cytokine interferon ␣ (IFN␣) by immunohistochemistry and for EBV by using the highly sensitive method of EBV-encoded RNA (EBER) in situ hybridization. Results:We detected overexpression of IFN␣ in active areas of white matter MS lesions but not in inactive MS lesions, normal-appearing white matter, or normal brains. The presence of IFN␣ in macrophages and microglia (expressing human leukocyte antigen class II) is suggestive of local production as part of an acute inflammatory process. Interestingly, EBERs were also specifically detected in areas where IFN␣ was overexpressed in these preselected active MS lesions. EBERϩ cells were also found in CNS lymphoma and stroke cases, but were absent in other control brains. We next addressed a potential mechanism, e.g., the role of EBERs in eliciting IFN␣ production, and transfected EBERs into human embryonic kidney (HEK) cells. We used HEK cells that stably expressed Toll-like receptor-3, which recognizes double-stranded RNAs, associated with many viral infections. EBERs elicited IFN␣ production in vitro. Conclusion:These findings suggest that latent EBV infection may contribute to the inflammatory milieu in active MS lesions by activating innate immune responses, e.g., IFN␣ production. Unraveling the underlying mechanisms may help in uncovering causal pathways and developing better treatment strategies for MS and other neuroinflammatory diseases. Neurology ® 2012;78:15-23 GLOSSARY ds ϭ double-stranded; EBER ϭ EBV-encoded RNA; EBNA-1 ϭ EBV nuclear antigen 1; EBV ϭ Epstein-Barr virus; FCS ϭ fetal calf serum; H&E ϭ hematoxylin & eosin; HEK ϭ human embryonic kidney; HLA ϭ human leukocyte antigen; HSE ϭ herpes simplex encephalitis; IFN␣ ϭ interferon ␣; IgG ϭ immunoglobulin G; ISH ϭ in situ hybridization; LFB ϭ Luxol fast blue; mAb ϭ monoclonal antibody; MS ϭ multiple sclerosis; OHL ϭ oral hairy leukoplakia; PBS ϭ phosphate-buffered saline; pDC ϭ plasmacytoid dendritic cell; PLP ϭ protein lipid protein; SLE ϭ systemic lupus erythematosus; SSPE ϭ subacute sclerosing panencephalitis; TLR3 ϭ Toll-like receptor-3.The innate immune response is an essential part of the host response to many infections and type I interferon (IFN) is produced within hours of systemic infection.1 Our previous findings showed that Toll-like receptor-3 (TLR3) stimulation of human microglia with the viral-mimic polyIC led to IFN␣ production and downstream Th1 polarization of CD4ϩ helper T cells, 2 which may impact on CNS immunity.