Functional characterization of the dural sinuses as a neuroimmune interface.

Despite the established dogma of central nervous system (CNS) immune privilege, neuroimmune interactions play an active role in diverse neurological disorders. However, the precise mechanisms underlying CNS immune surveillance remain elusive; particularly, the anatomical sites where peripheral adaptive immunity can sample CNS-derived antigens and the cellular and molecular mediators orchestrating this surveillance. Here, we demonstrate that CNS-derived antigens in the cerebrospinal fluid (CSF) accumulate around the dural sinuses, are captured by local antigen-presenting cells, and are presented to patrolling T cells. This surveillance is enabled by endothelial and mural cells forming the sinus stromal niche. T cell recognition of CSF-derived antigens at this site promoted tissue resident phenotypes and effector functions within the dural meninges. These findings highlight the critical role of dural sinuses as a neuroimmune interface, where brain antigens are surveyed under steady-state conditions, and shed light on age-related dysfunction and neuroinflammatory attack in animal models of multiple sclerosis.

Prozone phenomenon observed in indirect immunofluorescence assay by antibodies against neuronal antigens.

A marked prozone effect was observed in indirect immunofluorescence with human sera and human cerebrospinal fluid in two clinical cases involving breast carcinoma with paraneoplastic neuronal antibodies, and anti- N-methyl-D-aspartic acid (NMDA) receptor antibodies. Anti-Yo antibodies and anti-NMDA antibodies were not detectable under high concentrations (1:10 serum dilution and neat CSF respectively) but showed a true effect when sufficiently diluted at 1:80 and 1:5 respectively. This paper demonstrates that prozone effects have their occurrences in indirect immunofluorescence, and clinicians and laboratory technicians should be wary of its implications during screening of autoantibody markers in neurological diseases.

Antigen-antibody dissociation in Alzheimer disease: a novel approach to diagnosis.

With the ever-increasing population of aged individuals at risk of developing Alzheimer's disease (AD), there is an urgent need for a sensitive, specific, non-invasive, and diagnostic standard. The majority of efforts have focused on auto-antibodies against amyloid-beta (Abeta) protein, both as a potential treatment, and a reliable biomarker of AD pathology. Naturally occurring antibodies against Abeta are found in the CSF and plasma of patients with AD as well as healthy control subjects. To date, differences between diseased and control subjects have been highly variable. However, some of the antibody will be in preformed antigen-antibody complexes and the extent and nature of such complexes may provide a potential explanation for the variable results reported in human studies. Thus, measuring total amounts of antigen or antibody following unmasking is critical. Here, using a technique for dissociating antibody-antigen complexes, we found significant differences in serum antibodies to Abeta between AD and aged-matched control subjects. While the current study demonstrates the relevance of measuring total antibody, bound and unbound, against Abeta in AD, this technique may be applicable to diseases such as acquired immune deficiency syndrome and hepatitis B where determination of antigen and antibody levels are important for disease diagnosis and assessing disease progression.

Intrathecal anti-alphaB-crystallin IgG antibody responses: potential inflammatory markers in Guillain-Barré syndrome.

OBJECTIVE: alphaB-crystallin (alphaBC), a small stress protein with cytoprotective and anti-apoptotic functions, is a potent antigen in autoimmune demyelinating diseases. To address the role of alphaBC in Guillain-Barré syndrome (GBS) we analyzed humoral responses against alphaBC in relation to clinical, electrophysiological and CSF features in GBS. METHODS: Anti-alphaBC-IgG antibodies were measured in serum and cerebrospinal fluid (CSF) of patients with GBS (n = 41), infectious inflammatory neurological diseases (n = 21), multiple sclerosis (n = 42), and other, non-inflammatory neurological disorders (n = 40) by ELISA using human recombinant alphaBC. Expression of alphaBC was immunohistochemically analyzed in postmortem peripheral nerve tissue of GBS and controls without neuropathy. RESULTS: Serum alphaBC-IgG antibody levels did not differ between disease groups, whereas alphaBC-IgG antibodies in CSF were increased in GBS and infectious inflammatory neurological diseases. Calculation of an antigen specific alphaBC-IgG index (alphaBC-Ig-G(CSF) x total IgG(CSF))/(alphaBC-IgG(Serum) x total IgG(Serum)) revealed significantly elevated values in patients with GBS compared to other disease groups (p < 0.001). alphaBC-IgG indices exceeding a cut off value > 0.8 had an 85 % specificity and a 76 % sensitivity for GBS. alphaBC was overexpressed in dorsal root ganglia and spinal roots of autopsy cases with GBS. CONCLUSIONS: We demonstrate increased alphaBC-IgG indices in a high proportion of our GBS patients, which reflect enhanced antigen-specific intrathecal antibody responses against abnormally expressed alphaBC in inflamed peripheral nerve tissue. Elevated alphaBC-IgG indices might therefore serve as markers of PNS inflammation and supplement currently used laboratory tests in the diagnosis of GBS.

Increased ALZ-50 immunoreactivity in CSF of pseudotumor cerebri patients.

Pseudotumor cerebri (PTC) is characterized by increased intracranial pressure and papilledema without a mass lesion. PTC predominantly affects obese women. Currently, the pathogenesis of PTC is obscure. Since cytoskeletal abnormalities are found in many neurodegenerative diseases, we hypothesized that some cytoskeletal protein might be involved in the pathophysiology of PTC. Western blotting with specific antibody probes was employed to evaluate ALZ-50 immunoreactive protein, cytoskeletal microtubule-associated protein (MAP), and glial fibrillary acidic protein (GFAP) in cerebrospinal fluid (CSF) samples from 8 PTC patients and 6 controls. Immunoblotting of ALZ-50 in CSF revealed intense staining of 50 kDa protein bands in 7 of 8 PTC patients, while weak staining was found in 4 of 6 controls. Moderate staining of ALZ-50 was seen in 1 of 8 PTC patients and in 2 of 6 controls. CSF blots with anti-ALZ-50 antibody also showed intense staining of a 65 kDa protein band in 3 of the 8 patients but in none of the controls. In anti-MAP CSF blots of the PTC patients and controls, weak staining of the MAP 60 kDa and 50 kDa protein bands was observed. Weak staining of 60 kDa bands was also observed in anti-GFAP CSF blots of all PTC patients and controls. In CSF blots reacted with anti-GFAP antibody, 65 kDa and 32 kDa bands were evident in some PTC patients, but in none of the controls. This study indicates that ALZ-50 immunoreactivity is elevated in CSF of PTC patients. The ALZ-50 immunoreactive protein, either normal tau protein or its phosphorylated variant, may be useful as a biomarker for the diagnosis of PTC. Since the ALZ-50 monoclonal antibody was generated against brain homogenate from Alzheimer's disease (AD) patients, this study suggests a possible link between PTC and AD.

Consensus recommendations of the Italian Association for Neuroimmunology for immunochemical cerebrospinal fluid examination.

In 2002-2003, the Italian Association for Neuroimmunology (AINI) ran a program for procedure and method standardization in neuroimmunology. The main purposes of the program were: a) to improve the overall quality of analytical performance and, simultaneously, to reduce costs by resource optimization; b) to establish the bases for clinical guidelines in neurology; c) to promote the formation of laboratory networks and of joint research projects; d) to facilitate the procedures for certification required by governmental/non-governmental agencies. This report summarizes the consensus recommendations of a panel of AINI neuroimmunologists/biochemists involved in the field of cerebrospinal fluid examination. The collection process for said recommendations was guided by "impact-factored" literature and the knowledge of the experts involved. Communication was by email and face-to-face at two dedicated AINI workshops.

Use of immunoassays in neurological diagnosis and research.

OBJECTIVES: To review the use of immunoassays in the diagnosis and research of disorders affecting the nervous system. METHODS: Systematic review of the English literature. RESULTS: Immunoassays have demonstrated utility for: (1) the detection of antigen (molecules, genes, gene products, peptides, hormones and drug metabolites) and (2) the detection of an immune response (antigen-antibody complexes and specific and non-specific populations of antibodies) in serum, cerebrospinal fluid, and central nervous system tissue. DISCUSSION: The specificity of the antibody-antigen interaction makes immunoassays an ideal diagnostic and research tool for the investigation of neurological disease. A number of immunoassays are available for this purpose, and the choice of a particular methodology generally depends upon whether one is detecting antigen, antibody or antigen-antibody complexes, and the nature of the biologic sample that is being tested. Ease of testing, sensitivity, specificity and cost are other important considerations.

Search for a multiple sclerosis-specific brain antigen.

Specific reactivity of multiple sclerosis (MS) cerebrospinal fluid (CSF) IgG against central nervous system (CNS) tissue has been sought. Brain proteins were subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), reacted with CSF followed by 125I-Staph protein A, and developed for autoradiography. Pooled CSF from MS patients, CSF from individual MS patients, and pooled CSF from patients with other neurologic diseases and from normal patients, were used. Proteins were extracted from the white matter of MS and normal brains and from MS plaque and peri-plaque tissue. No reactivity specific for MS was observed.

Neurosyphilis in the antibiotic era.

Among 30 patients with neurosyphilis diagnosed between 1970 and 1981, 43% had symptoms attributable to neurosyphilis, 43% had unrelated symptoms, and 14% were asymptomatic. Serum VDRL was positive in 86%, and the CSF VDRL was positive in 53%. Meningovascular and vascular syphilis were relatively more common than in the prepenicillin era; tabes dorsalis and general paresis were unchanged in relative frequency.

Increased expression of B7-1 costimulatory molecule on cerebrospinal fluid cells of patients with multiple sclerosis and infectious central nervous system disease.

The expression of the costimulatory molecule B7-1 (BB-1; CD80) and its ligand CD28 was investigated on peripheral blood (PB) and cerebrospinal fluid (CSF) T and B lymphocytes and monocytes in 11 patients with relapsing-remitting multiple sclerosis (MS) 21 age-matched healthy controls and 10 patients with central nervous system (CNS) infectious disease (CID). Three channel flow cytometry was used with a novel gating technique in order to unambiguously identify the low numbers of B lymphocytes present in normal CSF. There was a significantly higher fraction of B7-1+ B lymphocytes in the CSF of patients with MS (72%) and CID (69%) when compared with healthy individuals (53%; p < 0.0001 and p < 0.002, respectively). Furthermore, two patients with a clinical picture of encephalitis showed a profoundly increased B7-1 expression on CSF monocytes. Comparison of absolute numbers of B7-1+ B lymphocytes/mL CSF between MS patients and healthy controls revealed a highly increased frequency of these cells among MS patients (235 cells/mL in MS patients versus 3.9 cells/mL in controls; p < 0.0001) with no overlap between the groups, which was otherwise seen for all other analyzed cell populations. We therefore hypothesize that activated B lymphocytes expressing high levels of B7-1 may be of pathogenetic importance in the development and maintenance of the MS disease.

Factor VIII-related antigen in cerebrospinal fluid.

A hundred and one samples of cerebrospinal fluid (CSF) were obtained from patients with bacterial meningitis (18), viral meningitis (9), lymphoproliferative disorders (33), 15 with meningeal infiltrations, multiple sclerosis (8), stroke (8) and 25 subjects with normal CSF. All samples were studied for VIIIR:Ag with specific and sensitive immunoradiometric assay (IRMA) and Laurell's technique. Prothrombin and factor IX antigenic activities were investigated by Laurell's technique. Simultaneously, plasma specimens from ten patients with bacterial meningitis were evaluated. Only a selective increase of VIIIR:Ag was demonstrated in CSF from bacterial meningitis whereas prothrombin and factor IX were not detected. VIIIR:Ag plasma and CSF levels were uncorrelated. Similarly, no relationship could be established between the degree of elevation of VIIR:Ag in the CSF and their protein concentration. These findings suggest that VIIIR:Ag elevation in CSF has diagnostic value for bacterial meningitis and that disruption of the blood-brain barrier is not responsible for their elevated levels. Accordingly, the presence of VIIIR:Ag in CSF may be an indication of endothelial damage in the choroid plexi.

Detection of antigens in Alzheimer cerebrospinal fluid by monoclonal antibodies.

Monoclonal antibodies raised against cerebrospinal fluid (CSF) and human ventral forebrain from patients with Alzheimer's disease were tested against CSF pools derived from patients with Alzheimer's disease and normal controls. Antibodies that appeared to distinguish the two pools were subsequently tested against 58 CSF samples from individual Alzheimer patients, normal controls, and individuals with other neurologic diseases. The mean CSF content of two antigens was decreased in patients with Alzheimer's disease compared with controls; the mean CSF content of one of these antigens was also decreased in patients with other neurologic diseases. Although group differences could be detected, the degree of overlap did not follow for the separation of individual patients within these three groups.

Immunoreactive S-antigen in cerebrospinal fluid: a marker of pineal parenchymal tumors?

This investigation evaluated the possibility that the occurrence of S-antigen in cerebrospinal fluid (CSF) might be used as a preoperative marker of pineal parenchymal tumors (pineoblastoma and pineocytoma). Such a marker could provide a means of preoperatively differentiating these neoplasms from pineal region tumors of other origin. The S-antigen, also known as the 48-kD protein or arrestin, is a highly antigenic protein originally found in the retina and pineal gland. In the retinal photoreceptors and submammalian pineal photoreceptors the protein is thought to be involved in phototransduction; its function in the mammalian pinealocyte is unknown. S-Antigen immunoreactivity also occurs in certain neoplastic cells of retinoblastomas, pineocytomas, pineoblastomas, and cerebellar medulloblastomas. This study included a group of 13 patients with tumors of the pineal region. Samples of CSF were obtained preoperatively and analyzed for the S-antigen using western blot technology. Tumor biopsy material was classified according to conventional neurohistological criteria and was also examined by immunocytochemical techniques for the presence of the S-antigen. S-Antigen immunoreactivity was found in the preoperative CSF of the one patient found to have pineocytoma; tumor tissue removed from this patient was the only neoplastic tissue examined in this study which contained S-Antigen immunoreactive tumor cells. Furthermore, hydroxyindole-O-methyltransferase activity was detectable in the pineocytoma but not in three other pineal tumors, and melatonin levels in the CSF of the pineocytoma patient were the highest in the patient group examined. These preliminary results suggest that testing for S-antigen in CSF might be useful in characterizing and treating tumors of the pineal region and, when identified in conjunction with other markers, it might also help to better define pineal parenchymal tumors. This study needs confirmation with a larger number of patients. If this approach is eventually found to be a reliable predictor of pineal cell tumors, it may supplant the need for surgical biopsies before initiating appropriate adjunctive therapy.

Low levels of soluble NG2 in cerebrospinal fluid from patients with dementia with Lewy bodies.

The proteoglycan NG2 plays a major role in proliferation, migration, and differentiation of pericytes and NG2 cells in the brain. We have previously reported decreased soluble NG2 (sNG2) levels in cerebrospinal fluid (CSF) from patients with Alzheimer's disease (AD) and a relationship between sNG2 and AD biomarkers in these patients. To further investigate whether alterations in sNG2 is specific to AD pathology, we measured levels of sNG2 in CSF from a patient cohort consisting of non-demented controls (n = 51), patients with Parkinson's disease (PD) (n = 61), and patients with dementia with Lewy bodies (DLB) (n = 37), two synucleinopathies whereof the latter disorder frequently coincides with amyloid-ß pathology similar to AD. We found decreased sNG2 concentrations in DLB patients, but not in PD patients, compared to controls. Levels of sNG2 in controls and PD patients correlated to T-tau, P-tau, α-synuclein, and neurosin. Only one correlation, between sNG2 and neurosin, was found in DLB patients. Analysis of a second cohort consisting of controls (n = 23) and DLB patients (n = 31) showed that the result was reproducible, as lower levels of sNG2 again were found in DLB patients compared to controls. We conclude that lower levels of sNG2 levels indicate a DLB-related impact on NG2 expressing cells foremost associated with neuropathology linked to accumulation of amyloid-ß and not α-synuclein.

[Detecting infectious agents in central nervous system inflammation by cerebrospinal fluid analysis]. / Liquoranalytik in der Diagnostik erregerbedingter Erkrankungen des Zentralnervensystems.

Cerebrospinal fluid analysis is the method of choice in CNS infection and provides the basis for appropriate treatment. Due to the proximity of CSF and CNS, the infectious agent may be detected directly by microscopy or antigen or nucleic acid detection--the latter by polymerase chain reaction--in native CSF or after culture. Furthermore, intrathecal antibody synthesis against the infectious agent may identify the cause of infection. This indirect antigen detection method requires correction for a systemic antibody response and a blood-CSF barrier disturbance. The following text gives an overview of appropriate detection methods and their relevance to the most important CNS infections.