Differentiation of oligodendrocyte precursors is impaired in the prefrontal cortex in schizophrenia
Introduction
Schizophrenia can be conceptualized as the clinical manifestation of the dysfunction of information processing and integration of neural networks across brain regions, including the different areas of the cerebral cortex and subcortical domains, such as the thalamus and medial temporal structures. It is well established that neuronal circuits within these regions are functionally disturbed in schizophrenia. Furthermore, impaired myelin ensheathment of the axons that connect these neuronal circuits via the white matter can further exacerbate network dysfunction by disturbing the timing of the arrival of action potential, compromising spike-timing dependent potentiation and hence disrupting the precise temporal coherence of neuronal circuit output that is necessary for synchronized network activation (Fields, 2005, Fields et al., 2014, Pajevic et al., 2013) which, in turn, may underlie many of the cognitive and perceptual deficits that are characteristic of schizophrenia (Cho et al., 2006, Lisman and Buzsaki, 2008, Spencer et al., 2004, Uhlhaas et al., 2008, Uhlhaas and Singer, 2010).
Structural magnetic resonance imaging (MRI) investigations have converged upon the conclusion that the volume of the white matter is decreased in patients with schizophrenia (Bora et al., 2011, Breier et al., 1992, Buchanan et al., 1998, Sigmundsson et al., 2001), consistent with the notion that the integrity of myelin is compromised. Furthermore, diffusion tensor imaging (DTI) has revealed that fractional anisotropy, which is a measure of a combination of factors, including myelin sheath integrity, is also decreased in schizophrenia patients (Holleran et al., 2014). More direct evidence of myelin deficit in this illness has come from postmortem studies. Specifically, the density of oligodendrocytes has been found to be decreased by 25–31% in both the gray and white matter of the prefrontal cortex in schizophrenia subjects (Hof et al., 2003, Uranova et al., 2004, Uranova et al., 2007, Vostrikov and Uranova, 2011). Furthermore, ultrastructural examination of oligodendrocytes has revealed morphological changes associated with cell degeneration and death (Uranova et al., 2001, Uranova et al., 2007), in addition to disturbances in the integrity of myelinated axonal fibers (Uranova et al., 2011). Finally, gene expression profiling of RNA extracted from homogenized gray and white matter of the cerebral cortex have revealed that many oligodendrocyte- and myelin-associated genes appear to be differentially expressed in subjects with schizophrenia (Aston et al., 2004, Hakak et al., 2001, Haroutunian et al., 2007, Hof et al., 2002, Katsel et al., 2005, Mitkus et al., 2008, Sequeira et al., 2012, Sugai et al., 2004, Tkachev et al., 2003).
Myelin is actively maintained by a complex series of events that tightly regulate the generation, differentiation, survival and death of oligodendrocytes. In the adult rat brain, roughly 5–10% of cells are oligodendrocyte progenitors (OPCs) (Dawson, 2003, Polito and Reynolds, 2005), whereas approximately 20% of the cells in the white matter in humans may be OPCs (Lojewski et al., 2014). It is further estimated that up to 80% of OPCs are actively replicating or differentiating (Trotter et al., 2010, Young et al., 2013). Disturbances of any of the events that underlie the differentiation and mediate the integrity of oligodendrocytes could lead to myelin and myelination deficits.
In an attempt to gain insight into the possible molecular mechanisms specifically associated with the dysfunction of cells that belong to the oligodendrocyte lineage and thereby myelin and myelination deficits in the white matter, we profiled the expression of mRNA in these cells obtained by laser-capture microdissection (LCM). CNPase (2′,3′-cyclic nucleotide 3′-phosphodiesterase) was chosen as a marker, because it is expressed in OPCs in addition to differentiating and myelinating oligodendrocytes (Belachew et al., 2001, Dawson, 2003, Levine et al., 1993, Polito and Reynolds, 2005, Reynolds et al., 2002, Scherer et al., 1994, Sprinkle, 1989, Tomassy and Fossati, 2014). Together with immunohistochemical validation of findings of our gene expression profiling experiment using oligodendrocyte lineage markers including neural/glial antigen 2 (NG2), a marker for OPCs, and oligodendrocyte transcription factor 2 (OLIG2), which is present in maturing and mature, myelinating oligodendrocytes, the preponderance of evidence derived from the present study appears to favor the interpretation that impairment in the differentiation of OPCs may contribute to the pathophysiology of white matter deficits of schizophrenia.
Section snippets
Postmortem human brain tissue
Fresh-frozen tissue blocks containing the prefrontal cortex (Brodmann's area 9) from 9 schizophrenia and 9 normal control subjects, matched for age, sex and postmortem interval (PMI), were obtained from the Harvard Brain Tissue Resource Center (Table 1). A detailed methodology for tissue preparation, LCM and RNA processing has been described in detail elsewhere (Pietersen et al., 2009, Pietersen et al., 2011, Pietersen et al., 2014a, Pietersen et al., 2014b, Simunovic et al., 2009). Postmortem
Gene expression profile of CNPase-immunoreactive cells in schizophrenia
We identified 497 genes that were differentially expressed (i.e. fold-change > 1.5 and p < 0.05) in the CNPase-immunoreactive cells in subjects with schizophrenia (Supplementary Table S2), the majority of which (56%) were downregulated. Pathway analysis revealed that many of the signaling cascades related to oligodendrocyte proliferation and differentiation appeared to be differentially regulated in schizophrenia subjects (Table 2). Validation of microarray findings were performed by statistically
Discussion
Findings of the gene expression profiling experiment reported here suggest that dysregulation of the differentiation of OPCs may contribute to oligodendrocyte and myelin deficits of schizophrenia. To address this hypothesis, we immunohistochemically visualized cells that expressed NG2, a marker for OPCs, and OLIG2, an oligodendrocyte lineage marker for OPCs, maturing oligodendrocytes and some mature, myelinating oligodendrocytes. We found that the density of NG2-immunoreactive cells was
Role of the funding source
The NIH played no role in the design and execution of this study.
Authors' contributions
SAM and CYP carried out LCM and RNA processing for microarray experiments. SAM performed qRTPCR validation and immunohistochemistry, participated in the microarray data analysis and drafted the manuscript. KCS participated in data analysis and manuscript preparation. TUWW conceived of the study, participated in its design and data interpretation, and prepared the manuscript.
Conflicts of interest and financial disclosure
None.
Acknowledgments
This study was supported by grants P50MH080272 (Boston CIDAR: Vulnerability to Progression in Schizophrenia) and R01MH076060 from the National Institutes of Health.
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