engleski

Aggregation and co-aggregation of specific proteins in postmortem brain samples

Background: Schizophrenia and other chronic mental illnesses (CMIs), such as major depressive disorder, are complex conditions with hereditary and non-genetic components. Since depressive episodes are a common clinical characteristic in schizophrenia, schizophrenia patients diagnosed with depression have a worse outcome, compared to patients without depression. Hence, investigating their shared elements could provide insight into shared biological pathways. As a result of overlapping symptoms between neurodegenerative disorders and CMIs, protein aggregation could be a possible pathological mechanism. Several proteins have so far been implicated as aggregating in CMIs, including Collapsin Response Mediator Protein 1 (CRMP1), Disrupted in Schizophrenia 1 (DISC1), Neuronal PAS domain-containing protein (NPAS3), and TRIO and F-actin-binding protein (TRIOBP-1). Most of these have been detected in multiple CMIs, suggesting aggregation to cross diagnostic boundaries. So far, the aggregation of one of these proteins alone was investigated, but some data suggests the idea of one protein stimulating aggregation of the others or co- aggregation. Methods: We investigated the presence of aggregation of these proteins simultaneously in human brain samples, using suicide as an alternative, discrete non-subjective measure that spans diagnoses. Studies were performed on brains samples from 14 victims of suicide and 14 control individuals. 6 samples each from patients with depression and Alzheimer's disease were also included for context. Brains were donated as part of the Hungary-wide Lenhossék Program, after granting of family or a legal permission. Samples of the insular cortex were isolated, and flashfrozen after a post-mortem interval of 4.60±0.28 hours. Ethical approval for each stage of this work was obtained from the ethical committees and boards from the Ministry of Health of Hungary, the Semmelweis University, and/or the University of Rijeka, as appropriate. Brain samples were homogenized, and the insoluble (including aggregating) protein fraction was purified with a series of solubilization buffers and ultracentrifugation. Both the original homogenates and this purified insoluble fraction were analyzed by Western Blot. All procedures were performed under blinded conditions. Results: In our study, we have seen TRIOBP-1, previously only seen to aggregate in schizophrenia, as insoluble in suicide victims or depression patients. However, specific species of DISC1 (n=2) and CRMP1 (n=3) were found insoluble in suicide victims relative to control samples. Additionally, levels of CRMP1 were seen to vary in the whole sample between depression patients and control individuals or victims of suicide (in both cases p<0.05 by one-way ANOVA). Neither sex nor age showed any significant effect on the results. Notably, many of these samples showed multiple proteins to be aggregating simultaneously. We, therefore, performed follow-up in vitro studies in neuroblastoma cells, to determine if any of these proteins can co-exist in the same intracellular aggregates. In most instances, aggregation of each protein was independent of each other, although DISC1 co-aggregated with CRMP1 (confirming a previous result) and also with TRIOBP-1 (a novel interaction). Discussion: Protein aggregation could be a common factor in major mental illness, with most instances of simultaneous aggregation arising from common underlying factors, rather than direct co- aggregation of the proteins, with the notable exception of DISC1 that can induce aggregation of some other proteins. Replication studies, as well as confirmation in additional brain regions, are now required to examine further this phenomenon and understand its pathological relevance.

Schizophrenia ; Protein aggregation ; Suicide ; Post-mortem tissue

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