Background: In our preceding study, we assayed in a blind fashion the blood sera of young normal subjects and schizophrenic patients for levels of platelet autoantibodies (PAA). The recorded PAA titers of the schizophrenic patients were significantly higher than those of the normal subjects. This observation has lent support to this test being used as an objective evaluation of schizophrenia in young subjects in the future. In addition, this finding strongly suggested that the etiology of a distinct group of sufferers of this disorder could have originated from an autoimmune reaction against platelets which can, under certain conditions, cross-react with brain tissue. Aims: In the present study, PAA titers in the sera of adult schizophrenic patients and matched normal subjects were determined analogously to the preceding study. The effect of hospitalization and drug treatments on the apparent blood test scoring in adult subjects could thus be evaluated. Methods: A total of 46 schizophrenia patients (30 men and 16 women) aged 19–45 years (mean ± SD: 31.7 ± 8.0 years) with a minimum score of 60 on the Positive and Negative Symptom Scale and 43 healthy control subjects (22 men and 21 women) aged 21–44 years (mean ± SD: 31.9 ± 6.9 years) participated in the study. The blood titers of PAA were evaluated in a single-blind fashion using an optimized ELISA test scored by optical density (OD) units. A positive test was defined as a value above 1.3 OD units. Results: The titers of PAA in the group of schizophrenic patients (1.1 ± 0.55 OD units, range: 0.360–2.285 OD units) were significantly higher in comparison to those of the healthy control subjects (0.81 ± 0.37 OD units, range: 0.360–1.704 OD units; p = 0.004, two-tailed unpaired t-test). Significantly more schizophrenic patients showed a positive test (15 patients out of 46) than the control subjects (5 out of 43). However, significantly higher OD values of 1.55 ± 0.5 were recorded in the group of patients with less than 3 years of registered disease (n = 16, age 19–30 years), while in the group with 4–20 years of hospitalization (n = 30, age 24–45 years) the recorded OD values (0.85 ± 0.4 OD units) were practically indistinguishable from those of the control group. Conclusions: In the adult schizophrenic patients, the PAA blood test remains valid for patients who were hospitalized for less than 3 years. Drug treatment, length of disease and age can be assumed to reduce the PAA level considerably.

We previously found autoantibodies against platelets in schizophrenic patients. One of the platelet proteins that bind these antibodies is enolase. Here, we describe the isolation and sequencing of an immunoreactive peptide after enzymatic digestion of enolase. The 3-D structure of enolase indicates that, unexpectedly, this peptide is buried inside the protein. However, 3-D surface analysis leads to the identification of a conformational epitope that resembles the binding peptide and might constitute a specific binder of the autoantibodies. In a screening of antibody binding with the peptide LVVGLCK, we found in 50 serum samples of controls a mean of O.D.=0.46; s= +/- 0.21 relative enzyme immunoassay units, while in sera of 39 schizophrenic patients, we found a mean of O.D.=1.47; s= +/- 0.65; P<0.0001. Furthermore, an inverse correlation was observed between duration of schizophrenia and the level of the detected autoantibodies. A screening of autoantibodies in sera of various mental disorders with this peptide is currently in progress.

Peripheral blood lymphocytes (PBL) isolated from 17 healthy subjects were engrafted i.p. each to an individual SCID mouse. After 48 hours the mice were tested for skin response to the common recall antigen PPD at various sites on the body periphery. In 8 of them PPD elicited a distinct positive skin reaction only at the abdomen. This procedure could provide a basis for detection of specific cellular immune response such as in autoimmune diseases.

Various findings suggest auto-immune changes in schizophrenia. We have recently demonstrated that platelets from schizophrenic patients bear autoantibodies (PAA) which cross-react with brain antigens. Accordingly, treatment of schizophrenia with an immunosuppressant might be of potential benefit. In a recent case study, a chronic schizophrenic patient treated with azathioprine has demonstrated a clear psychiatric improvement preceded by a decrease in PAA level. A phase I study designed for assessing side-effects of short-term azathioprine treatment in a group of schizophrenic patients is described here. From a group of 40 chronic non-responsive patients, 14 patients demonstrating high PAA level have entered the study and 11 have complied all along. Two groups were tested in parallel. In the first (6 patients) 150 mg/day was given for 7 weeks while in the second (5 patients) the same regimen was given for two periods of 7 weeks with an interval of 6 weeks. Blood biochemistry and cell count, as well as determination of PAA were carried out weekly, starting 3 weeks before the trial and continuing up to 7 weeks after the treatment. Two out of 11 patients developed leucopenia in week 4. No other side-effects were recorded in any of the patients. A substantial reduction in PAA was observed in 3 out of 6 patients in group I and 4 out of 5 in group II. Two patients showed improvement of psychiatric symptomatology. Our results demonstrate that short-term azathioprine treatment induces transient leucopenia in 18% of the patients receiving the drug, much alike the percentage reported for other patient populations.

Platelets isolated from blood of demented and schizophrenic patients were found to bear surface antibodies at a considerably higher titer than those found on platelets from normal age-matched groups or patients with affective disorders. The platelet count in demented and schizophrenic patients correlated inversely with the level of the platelet associated antibodies (PAA) which suggested an autoimmune route of opsonization. In most individual cases of dementia or schizophrenia PAA and platelet count were found to oscillate with time between high PAA-low platelet number and low PAA-high platelet number in approximately inverse correlation. PAA isolated from demented patients were found to cross-react with platelets from normals and with brain tissue from rats. Furthermore, molecular weights of specific brain antigens were identified by binding to PAA. These observations support the possibility that PAA might be implicated in the etiology of some mental dysfunctions associated with dementia and schizophrenia.

Balb/c mice were immunized with rat striatal integral membrane proteins. After hybridization of splenocytes with myeloma cells, hybridoma lines secreting antibodies against serotonin, dopamine and opiate receptors were detected by inhibition of ligand binding to brain membrane preparations. Antibodies from two positive lines, Mab/a9 and Mab/a18, were able to inhibit ligand binding to the S2-serotonin (Kd range: 10-100 nM), the mu-opiate (Kd range: 0.4-3 microM) and the delta-opiate receptors (Kd range: 0.7-1.1 microM), while Mab/a9 was also found to inhibit ligand binding to the D2/D4-dopamine receptor (Kd approximately 50 nM). An apparent molecular mass of 60 kDa could be ascribed to the delta-opiate receptor and apparent molecular masses of 29 and 36 kDa to the mu-opiate receptor by ligand elution from immuno-precipitates.

The overt effect of pressure on biological membranes is mediated predominantly through lipid condensation and disintegration of cytoskeletal polymers. These may lead to selective shedding of integral proteins, which could then be isolated by conventional means. In this study we have used the well characterised human erythrocyte membrane in order to establish the technical requirements for future use of pressure, as an alternative to detergents, in isolation of membrane proteins. Pressure of varying magnitude (300-1640 bar) and duration (5-60 min) was applied on human erythrocyte ghost membranes in suspension at different temperatures (4, 24 and 37 degrees C) and in the presence of various solutes. After ultracentrifugation protein and lipids remaining in the supernatant were quantified and analysed. It is indicated that selective integral membrane proteins can be shed off under defined conditions and presumably remain in solution by the support of strongly associated phospholipids and specific solutes. On the basis of our findings a series of technical recommendations for the isolation of specific membrane proteins is outlined.