Karl Bechter

Cerebrospinal fluid analysis in affective and schizophrenic spectrum disorders: Identification of subgroups with immune responses and blood–CSF barrier dysfunction

Immune and inflammatory mechanisms are detected in a subgroup of treatment resistant hospitalized affective and schizophrenic spectrum disorder patients. We analysed albumin, IgG, IgA, IgM, oligoclonal IgG and specific antibodies in paired cerebrospinal fluid (CSF) and serum samples. Numerical and graphical interpretation of CSF protein data was performed by Reibergrams with a new CSF statistics tool for nonlinear group analysis with reference to a large control group (n=4100). In 41% of the psychiatric patients (n=63) we observed CSF pathologies: 14% displayed intrathecal humoral immune responses, 10% slightly increased CSF cell counts (5-8/microL) and 29% had moderate blood-CSF barrier dysfunctions, in 24% as the only pathological sign with normal IgG, IgA and IgM concentrations in CSF (p=0.9 testing the null hypothesis for intrathecal synthesis with reference to Qmean of the reference group). In the group of affective (n=24) spectrum disorders 20% displayed a systemic immune reaction as detected by oligoclonal IgG. CSF analysis and interdisciplinary clinical approach revealed 6% of psychiatric patients likely to represent a virusspecific, bacterial or autoimmune associated disorder with CNS involvement. Elevated CSF neopterin concentration in 34% of the patients was interpreted as an increased release from astrocytes or from other glia cells. The low level immune response and barrier dysfunctions are discussed on the base of a mild encephalitis pathomechanism in subgroups of psychiatric patients. CSF analysis is shown to be a useful diagnostic tool for differential diagnosis in psychiatric diseases.

Borna disease, a possible hazard for man?

SummaryEvidence is presented that Borna disease (BD) virus, which is known to cause encephalopathy in horses, sheep, and a broad range of experimental animals, or a related agent, can infect man and may induce mental disorders. BD virus-specific antibodies could be demonstrated in 4–7% of sera (depending on origin) from more than 5000 psychiatric or neurological patients from Germany, U.S.A. and Japan. Antibodies from seropositive patients reacted with a BD virus-specific protein translated by RNAs which were transcribed from a cDNA clone obtained from BD virus-infected tissues. When the cerebrospinal fluid from three seropositive patients was inoculated into rabbits or rabbit embryonic brain cell cultures, evidence was obtained that suggests the presence of BD virus or a related agent.

Immuno-inflammatory, oxidative and nitrosative stress, and neuroprogressive pathways in the etiology, course and treatment of schizophrenia

In recent decades, a significant role for altered immunoinflammatory, oxidative and nitrosative stress (IO&NS) pathways in schizophrenia has been recognized (Smith and Maes, 1995; Wood et al., 2009). Importantly, such processes have provided crucial clues to the etiology, course and management of this devastating disorder. This is the focus of this special edition. Epidemiological findings supporting a role for prenatal viral, bacterial and protozoan infections in the etiology of schizophrenia have provided a seminal contribution to the neurodevelopmental hypothesis of schizophrenia (Brown and Derkits, 2010). The early developmental etiology of schizophrenia to a lesser extent has been focused on decreased vitamin D in early development, including via vitamin D modulation of the immune response to infection (McGrath et al., 2003). Interactions between these factors is suggested by the fact that vitaminDhas a documented role in immunemodulation, especially during placental development and in early childhood (Battersby et al., 2012; Liu et al., 2011). The maximal risk period for maternal infection association with offspring schizophrenia is shown to be early pregnancy (Brown et al., 2004; Khandaker et al., 2012). Interestingly many schizophrenia susceptibility genes are regulated by hypoxia, suggesting close interactions among IO&NS genes and obstetric complications leading to enhanced risk of schizophrenia (Nicodemus et al., 2008; Schmidt-Kastner et al., 2006). Other conditions of pregnancy, including hypoxia associated preeclampsia (Kendell et al., 1996), also increase the risk of the offspring being classed as having schizophrenia, emphasizing the profound impact of prenatal events. The evidence for the role of prenatal infection, both epidemiological and experimental, is excellently reviewed byUrsMeyer (2013–this issue) who has published extensively in this area. Many of the developmental effects of infection are driven not only by O&NS and proinflammatory cytokine increases in the placenta and fetus, but also by associated hypoferremia and zinc deficiency (Ganz and Nemeth, 2009; Prasad, 2009). Such changes render the offspring prone to subsequent second hits over the course of post-natal development, contributing to both the emergence and progression of disease manifestations. This is an important area of experimental research given that the elimination of the effects of maternal infection is estimated to decrease the incidence of schizophrenia by as much as 46% (Brown and Derkits, 2010).

High-avidity human serum antibodies recognizing linear epitopes of borna disease virus proteins

BACKGROUND The recent observation that Borna disease virus (BDV)-reactive antibodies from psychiatric patients exhibit only low avidity for BDV antigen called into question their diagnostic value and raised the possibility that antigenically related microorganisms or self antigens caused the production of these antibodies. We further characterized the specificity of these antibodies. METHODS We established a peptide array-based screening test that allows the identification of antibodies directed against linear epitopes of the two major BDV proteins, the nucleoprotein (N) and the phosphoprotein (P). RESULTS Initial tests employing sera of BDV-infected mice and rats or horses with Borna disease revealed a high specificity and sensitivity of this test. All sera recognized epitopes of N, P, or both. Sera of noninfected rats, mice, and horses showed no signals on either peptide array. Several human sera that recognized BDV antigen by indirect immunofluorescence contained antibodies that recognized various linear epitopes of one or even both BDV proteins. Remarkably, antibodies purified from such human serum by matrix-immobilized peptides showed high-avidity binding to BDV antigens when assayed by IFA or Western blotting. CONCLUSIONS These data suggest that reactive antibodies found in psychiatric patients indeed indicate infection with BDV or a BDV-like agent. However, the poor affinity maturation of BDV-specific human antibodies remains unexplained.

Flow cytometric analysis of T cell subsets in paired samples of cerebrospinal fluid and peripheral blood from patients with neurological and psychiatric disorders

Recent studies suggest inflammatory mechanisms involved in the pathogenesis of major psychiatric disorders (MPD). T cells play a major role during inflammation, but little is known about T cell subpopulations in the cerebrospinal fluid (CSF). We investigated the frequency of cells positive for the surface markers CD4, CD8, CD25, CD45, CD69, and CD127 in 45 paired cerebrospinal fluid (CSF) and peripheral blood (PB) samples by multiparameter flow cytometry from patients with MPD of the schizophrenic and affective spectrum with normal CSF cell counts and compared them with those from patients with non-inflammatory (NIND), chronic inflammatory (CIND) neurological disorders, and meningitis (MEN). In MEN patients, CD4+ cell frequency in PB, but not in CSF, was significantly increased as compared to CIND and NIND. No difference between patient groups was observed for CD8+. CD4+CD45RO+ double positive cells in PB were significantly lower in CIND than in MEN or NIND. The frequency of CD4+CD25+ cells in PB was significantly higher in MEN than in MPD or CIND. For CSF, the percentage of CD4+CD127(dim) cells was significantly lower in MEN than in MPD. CD4+CD127(dim) in PB and CSF showed overlapping characteristic clusters between MPD and CIND and MEN patients. Overall, the hypothesis of low degree inflammation in a subgroup of MPD is supported. The analysis of lymphocyte subsets in PB and CSF constitutes a novel promising tool to understand underlying pathomechanisms in psychiatric and neurological disorders on an individual case level.

The role of transmitter diffusion and flow versus extracellular vesicles in volume transmission in the brain neural–glial networks

Two major types of intercellular communication are found in the central nervous system (CNS), namely wiring transmission (point-to-point communication, the prototype being synaptic transmission with axons and terminals) and volume transmission (VT; communication in the extracellular fluid and in the cerebrospinal fluid (CSF)) involving large numbers of cells in the CNS. Volume and synaptic transmission become integrated inter alia through the ability of their chemical signals to activate different types of receptor protomers in heteroreceptor complexes located synaptically or extrasynaptically in the plasma membrane. The demonstration of extracellular dopamine (DA) and serotonin (5-HT) fluorescence around the DA and 5-HT nerve cell bodies with the Falck–Hillarp formaldehyde fluorescence method after treatment with amphetamine and chlorimipramine, respectively, gave the first indications of the existence of VT in the brain, at least at the soma level. There exist different forms of VT. Early studies on VT only involved spread including diffusion and flow of soluble biological signals, especially transmitters and modulators, a communication called extrasynaptic (short distance) and long distance (paraaxonal and paravascular and CSF pathways) VT. Also, the extracellular vesicle type of VT was demonstrated. The exosomes (endosome-derived vesicles) appear to be the major vesicular carriers for VT but the larger microvesicles also participate. Both mainly originate at the soma–dendritic level. They can transfer lipids and proteins, including receptors, Rab GTPases, tetraspanins, cholesterol, sphingolipids and ceramide. Within them there are also subsets of mRNAs and non-coding regulatory microRNAs. At the soma–dendritic membrane, sets of dynamic postsynaptic heteroreceptor complexes (built up of different types of physically interacting receptors and proteins) involving inter alia G protein-coupled receptors including autoreceptors, ion channel receptors and receptor tyrosine kinases are hypothesized to be the molecular basis for learning and memory. At nerve terminals, the presynaptic heteroreceptor complexes are postulated to undergo plastic changes to maintain the pattern of multiple transmitter release reflecting the firing pattern to be learned by the heteroreceptor complexes in the postsynaptic membrane.

Low avidity of human serum antibodies for Borna disease virus antigens questions their diagnostic value

Borna disease virus (BDV) can induce neurological disease in animals.1,2 Since viral nucleic acid,3–5 infectious particles6–8 and antibodies recognizing BDV antigens9–11 were found at higher frequencies in psychiatric patients than in healthy controls, BDV is suspected to cause psychiatric disorders in humans. However, the human origin of these viruses has recently been questioned.12 To diagnose BDV infections, sera are usually analyzed for antiviral antibodies by indirect immunofluorescence (IFA) on virus-infected cells.9,10,13 This study reveals that the reactive antibodies in human sera mainly recognized the BDV phosphoprotein, whereas animal sera preferentially detected the viral nucleoprotein. Immunoglobulin (Ig) G in sera of experimentally or naturally infected animals bound to the viral antigen with high avidity, ie resisting 3 M urea, whereas reactive IgG in human sera did not. Longitudinal studies showed that reactive human antibodies persisted for many years without gaining high avidity for BDV antigens, indicating that they were probably not induced by BDV but rather by infection with an antigenically related microorganism of unknown identity or by exposure to other related immunogens.

Cerebrospinal fluid and serum cytokine profiling to detect immune control of infectious and inflammatory neurological and psychiatric diseases.

The present study aimed at profiling inflammatory cytokines for neurological and psychiatric diseases. A total of 86 patients with meningitis, multiple sclerosis, tension-type headache, idiopathic facial nerve palsy (IFNP), affective and schizophrenic disorders were tested for both, serum and cerebrospinal fluid (CSF) using a multiplexed cytokine ELISA for IFN-γ, TNF-α, IL-1β, IL-2, IL-4, IL-5, IL-8/CXCL8, IL-10, IL12p70, IL-13 and IL-17. Cases with viral and bacterial meningitis had unequivocally higher cytokine concentrations in the CSF when compared with serum. Bacterial meningitis was unique by extremely elevated IL-17, TNF-α and IL-1β, indicating a plethora of inflammatory pathways, selectively activated in the CSF. In relapsing multiple sclerosis, IFN-γ and IL-10 were elevated in both, serum and CSF, but IL-12p70, IL-5, IL-13, and TNF-α were more prominent in serum than in CSF. Qualitatively similar biomarker patterns were detected in patients with idiopathic facial nerve palsy and tension-type cephalgia. Affective and schizophrenic disorders clearly present with an inflammatory phenotype in the CSF and also serum, the cytokines determined were in general higher in schizophrenia. Except IFN-γ, schizophrenic patients had higher IL-12p70 and a trend of higher IL-10 and IL-13 in serum suggesting a more prominent TH2-type counter regulatory immune response than in affective disorders. These differences were also mirrored in the CSF. Elevated IL-8 appears to be the most sensitive marker for inflammation in the CSF of all diseases studied, whereas TNF-α was restricted to peripheral blood. With the exception of IL-8, all but viral and bacterial meningitis, studied, displayed higher means of elevated lymphokine concentrations in the serum than in the CSF. This observation supports the concept of immunological crosstalk between periphery and intrathecal immunity in neurological and psychiatric diseases.

Borna disease virus-related therapy-resistant depression improved after cerebrospinal fluid filtration.

K. Bechter *, S. Herzog , V. Schreiner , H. Brinkmeier , P. Aulkemeyer , F. Weber , K.H. Wollinsky , R. SchuE ttler a Department of Psychiatry II, University of Ulm, Ludwig-Heilmeyer-Strasse 2, 89312 Bezirkskrankenhaus GuEnzburg, Germany Institute of Virology, University of Giessen, Germany Department of General Physiology, University of Ulm, Germany Department of Anaesthesiology and Intense Care, University of Ulm, Germany

Updating the mild encephalitis hypothesis of schizophrenia

Emerging evidence indicates that low level neuroinflammation (LLNI) may not occur infrequently. Many infectious agents with low overall pathogenicity are risk factors for psychoses including schizophrenia and for autoimmune disorders. According to the mild encephalitis (ME) hypothesis, LLNI represents the core pathogenetic mechanism in a schizophrenia subgroup that has syndromal overlap with other psychiatric disorders. ME may be triggered by infections, autoimmunity, toxicity, or trauma. A ‘late hit’ and gene–environment interaction are required to explain major findings about schizophrenia, and both aspects would be consistent with the ME hypothesis. Preliminary criteria for subgrouping neurodevelopmental, genetic, ME, and other types of schizophrenias were provided. Considering recent investigations of CSF, the ME schizophrenia subgroup may constitute approximately 40% of cases. LLNI may involve dysfunction of the blood–brain barrier, the blood–CSF barrier of CNS-endogenous immunity in part mediated by the volume transmission mode involving CNS-extracellular-fluid and CSF signaling. Both together could represent a common pathogenetic link for the distributed brain dysfunction observed in schizophrenia. However, CSF signaling may even extend along nerves into peripheral tissues via the CSF outflow pathway and explain peripheral topologies of dysfunctions/lesions found (dysautonomia, muscle lesions). In general, CSF signaling including at the PCOP could play an underestimated role in neuroinflammatory disorders.