Schizophrenia (SZ) is a debilitating neurological disorder with a world-wide prevalence of 1 percent; there is a strong genetic component, with an estimated heritability of 80-85 percent. Though postmortem studies have revealed reduced brain volume, cell size, spine density, and abnormal neural distribution in the prefrontal cortex and hippocampus of SZ brain tissue and neuropharmacological studies have implicated dopaminergic, glutamatergic and GABAergic activity in SZ, the cell types affected in SZ and the molecular mechanisms underlying the disease state remain unclear. To elucidate the cellular and molecular defects of SZ, we directly reprogrammed fibroblasts from SZ patients into human induced pluripotent stem cells (hiPSCs) and subsequently differentiated these disorder-specific hiPSCs into neurons (Fig. 1). SZ hiPSC neurons showed diminished neuronal connectivity in conjunction (Fig. 2) with decreased neurite number, PSD95-protein levels, and glutamate receptor expression. Gene expression profiles of SZ hiPSC neurons identified altered expression of many components of the cAMP and WNT signaling pathways. Key cellular and molecular elements of the SZ phenotype were ameliorated following treatment of SZ hiPSC neurons with the antipsychotic Loxapine.

Fig. 1. Patient-specific hiPSCs, NPCs and neurons. Left. hiPSCs express NANOG (green) and TRA-1-60 (red). DAPI (blue). ×100, scale bar 100 μm. Centre. hiPSC neural progenitor cells (NPCs) express NESTIN (green) and SOX2 (red). DAPI (blue). ×600, scale bar 100 μm. Right. hiPSC neurons express βIII-tubulin (red) and the dendritic marker MAP2AB (green). DAPI (blue). ×200, scale bar 100 μm.

Though its characteristic symptoms generally appear late in adolescence, SZ is thought to be a neurodevelopmental condition wherein illness is the end stage of abnormal neurodevelopmental processes that began years before the onset of symptoms. Consistent with this, SZ is often predated by a prodromal period that can appear in early childhood. Childhood-onset schizophrenia (COS) is defined by an onset of psychosis prior to age twelve and is a rare and particularly severe form of the disorder. We have reprogrammed fibroblasts from patients with COS into human induced pluripotent stem cell (hiPSCs) and will subsequently differentiate these disorder-specific hiPSCs into neural cells. Our preliminary data demonstrates that hiPSC neural progenitor cells (NPCs) most resemble early human fetal brain tissue, specifically the embryonic ganglionic eminence and cortical forebrain regions. Therefore, we believe our hiPSC NPCs are an ideal platform with which to study the early embryonic development effects that contribute to disease initiation and progression in COS. Ongoing work in the laboratory is to identify aberrant mRNA and microRNA expression of COS neural cells and to begin mechanistic studies of candidate mRNAs and microRNAs we find to be altered in COS. We hope to use our novel hiPSC based platform of COS to identify molecular insights into COS which may be generalizable across SZ.

Fig. 2. Decreased neuronal connectivity in SZ hiPSC neurons. A. Representative images of control and SZ hiPSC neurons cotransduced with LV-SYNP-HTG and Rabies-ENVAΔG-RFP, 10 days post rabies transduction. All images were captured using identical laser power and gain settings. βIII-tubulin staining (purple) of the field is shown below each panel. ×400, scale bar 80 μm. B. Graph showing treatment of SZ hiPSC neurons with Loxapine resulted in a statistically significant improvement in neuronal connectivity. Error bars are s.e., *P < 0.05.
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Kristen Brennand, PhD
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