Immunization with Recombinantly Expressed LRP4 Induces Experimental Autoimmune Myasthenia Gravis in C57BL/6 Mice.

BACKGROUND: Myasthenia gravis (MG) is an autoimmune disease of the neuromuscular junction (NMJ), characterized with muscle weakness. While MG develops due to acetylcholine receptor (AChR) antibodies in most patients, antibodies to muscle-specific receptor tyrosine kinase (MuSK) or low-density lipoprotein receptor-related protein 4 (LRP4) may also be identified. Experimental autoimmune myasthenia gravis (EAMG) has been previously induced by both LRP4 immunization and passive transfer of LRP4 antibodies. OBJECTIVE: Our aim was to confirm previous results and to test the pathogenic effects of LRP4 immunization in a commonly used mouse strain C57BL/6 (B6) using a recombinantly expressed human LRP4 protein. METHODS: B6 mice were immunized with human LRP4 in CFA, Torpedo Californica AChR in CFA or only CFA. Clinical and pathogenic aspects of EAMG were compared among groups. RESULTS: LRP4- and AChR-immunized mice showed comparable EAMG clinical severity. LRP4-immunized mice displayed serum antibodies to LRP4 and NMJ IgG and complement factor C3 deposits. IgG2 was the dominant anti-LRP4 isotype. Cultured lymph node cells of LRP4- and AChR-immunized mice gave identical pro-inflammatory cytokine (IL-6, IFN-γ and IL-17) responses to LRP4 and AChR stimulation, respectively. CONCLUSION: Our results confirm the EAMG-inducing action of LRP4 immunization and identify B6 as a LRP4-EAMG-susceptible mouse strain. Demonstration of complement fixing anti-LRP4 antibodies in sera and complement/IgG deposits at the NMJ of LRP4-immunized mice indicates complement activation as a putative pathogenic mechanism. We have thus developed a practical LRP4-induced EAMG model using a non-conformational protein and a widely available mouse strain for future investigation of LRP4-related MG.

Sorcin antibody as a possible predictive factor in conversion from radiologically isolated syndrome to multiple sclerosis: a preliminary study.

OBJECTIVE: To identify an antibody biomarker for prediction of conversion from radiologically isolated syndrome (RIS) to relapsing remitting multiple sclerosis (RRMS). METHODS: Sera of 13 RIS patients were screened by a protein macroarray derived from human fetal brain cDNA library. RESULTS: Sequencing of a clone with the highest signal intensity revealed sorcin as a potential target autoantigen in RIS patients. ELISA studies showed high-titer sorcin-antibodies in 3 of 4 RIS patients who converted to RRMS in a 5-year follow-up period and 13 of 23 control RRMS patients. CONCLUSION: The value of sorcin antibody as a predictor of conversion from RIS to RRMS requires to be tested in larger prospective studies.

Switch-associated protein 70 antibodies in multiple sclerosis: possible association with disease progression.

OBJECTIVE: This study was conducted to identify a biomarker for multiple sclerosis (MS) that can be used as a predictor of relapse and disability. MATERIALS AND METHODS: Sera of 26 consecutive relapsing-remitting MS (RRMS) patients were screened for switch-associated protein 70 (SWAP-70) antibody, which was previously identified by protein macroarray. The serum levels of several cytokines, chemokines and soluble adhesion molecules related to MS attacks were measured by enzyme-linked immunosorbent assay (ELISA). A possible correlation was sought among levels of SWAP-70 antibody, measured humoral factors and disability scores. RESULTS: ELISA studies showed high-titre SWAP-70 antibodies in 16 (61.5%) RRMS sera obtained during the attack period and 9 (34.6%) sera obtained during remission. There was a significant inverse correlation between SWAP-70 antibody levels and expanded disability status scale scores, CXCL10, soluble VCAM-1, CXCL13 and soluble VLA-4 levels. CONCLUSION: Our results showed that SWAP-70 antibodies could potentially be utilized as relapse and prognostic biomarkers in MS. Whether or not SWAP-70 antibodies have any effect on disease mechanisms requires further investigation.

CACNA1H antibodies associated with headache with neurological deficits and cerebrospinal fluid lymphocytosis (HaNDL).

BACKGROUND: Patients with the syndrome of headache with neurological deficits and lymphocytosis (HaNDL) typically present with recurrent and temporary attacks of neurological symptoms and cerebrospinal fluid lymphocytosis. AIM AND METHODS: To identify potential HaNDL-associated antibodies directed against neuronal surface and/or synapse antigens, sera of four HaNDL patients and controls were screened with indirect immunohistochemistry, immunofluorescence, cell-based assay, radioimmunoassay, protein macroarray and enzyme-linked immunosorbent assay (ELISA). RESULTS: Although HaNDL sera did not yield antibodies to any of the well-characterized neuronal surface or synapse antigens, protein macroarray and ELISA studies showed high-titer antibodies to a subunit of the T-type voltage-gated calcium channel (VGCC), CACNA1H, in sera of two HaNDL patients. CONCLUSION: Our results support the notion that ion channel autoimmunity might at least partially contribute to HaNDL pathogenesis and occurrence of neurological symptoms.

Progressive encephalomyelitis with rigidity and myoclonus: a syndrome with diverse clinical features and antibody responses.

BACKGROUND/AIMS: To better characterize progressive encephalomyelitis with rigidity and myoclonus (PERM) syndrome and identify novel PERM phenotypes. METHODS: The clinical features and antibody status of PERM patients were investigated using immunoblots, cell-based assays, RIA, protein macroarray and ELISA. RESULTS: Two patients with supratentorial involvement showed abnormal PET or EEG findings. One patient was discovered to have renal cell carcinoma, and protein macroarray revealed Ma3-antibodies. Another patient with leucine-rich, glioma-inactivated 1 (LGI1) and glutamic acid decarboxylase (GAD) antibodies showed a good response to immunotherapy. CONCLUSION: The heterogeneity of the immunological features suggests that PERM is caused by diverse pathogenic mechanisms. Seropositivity to well-characterized neuronal cell surface antigens might indicate a good treatment response.

Switch-associated protein 70 antibodies in multiple sclerosis: relationship between increased serum levels and clinical relapse.

OBJECTIVE: To identify an antibody biomarker for multiple sclerosis (MS) that can be used as a predictor of MS relapses. METHODS: MS patients' sera were screened by a protein macroarray derived from human fetal brain cDNA library (hEX1). Sera of 90 consecutive relapsing remitting MS (RRMS) patients and age-matched 145 Behçet's disease (BD) patients, 40 infectious meningoencephalitis patients, and 70 healthy controls were screened by ELISA for serum antibodies against the selected clone. RESULTS: Sequencing of the clone with the highest signal intensity revealed switch-associated protein 70 (SWAP70) as a potential target autoantigen in RRMS. ELISA studies showed high-titer SWAP70-antibodies in 21 (23.3 %) RRMS and 7 (4.8 %) BD patients. SWAP70 antibodies were more likely to be found positive in sera obtained during or shortly after a relapse. CONCLUSION: Detection of SWAP70 antibodies during the attack period might suggest that SWAP70 is involved in MS relapse pathogenesis. Whether serum SWAP70 antibody detection may be utilized as an MS relapse predictor should be tested in prospective studies.

Mitochondrial carrier homolog 1 (Mtch1) antibodies in neuro-Behçet's disease.

Efforts for the identification of diagnostic autoantibodies for neuro-Behcet's disease (NBD) have failed. Screening of NBD patients' sera with protein macroarray identified mitochondrial carrier homolog 1 (Mtch1), an apoptosis-related protein, as a potential autoantigen. ELISA studies showed serum Mtch1 antibodies in 68 of 144 BD patients with or without neurological involvement and in 4 of 168 controls corresponding to a sensitivity of 47.2% and specificity of 97.6%. Mtch1 antibody positive NBD patients had more attacks, increased disability and lower serum nucleosome levels. Mtch1 antibody might be involved in pathogenic mechanisms of NBD rather than being a coincidental byproduct of autoinflammation.

Anti-neuronal and stress-induced-phosphoprotein 1 antibodies in neuro-Behçet's disease.

No disease-specific neuronal antibodies have so far been defined in neuro-Behçet's disease (NBD). Immunohistochemistry and immunocytochemistry studies showed antibodies to hippocampal and cerebellar molecular layers and the surface antigens of cultured hippocampal neurons in sera and/or cerebrospinal fluids (CSF) of 13 of 20 NBD and 6 of 20 BD patients but not in multiple sclerosis or headache controls. Screening with a protein macroarray led to identification of stress-induced-phosphoprotein-1 (STIP-1) as an antigenic target. High-titer STIP-1-antibodies were detected in 6 NBD patients' sera but not in controls. These results suggest that neuronal antibodies could be useful as diagnostic biomarkers in NBD.