Aberrant activity at rest of the associative striatum in schizophrenia: Meta-analyses of the amplitude of low frequency fluctuations.

Schizophrenia is a severe psychiatric disorder associated with brain alterations at rest. Amplitude of low-frequency fluctuations (ALFF) and its fractional version (fALFF) have been widely used to investigate alterations in spontaneous brain activity in schizophrenia. However, results are still inconsistent. Furthermore, while these measurements are similar, they showed some differences, and no meta-analysis has been yet performed to compare them in schizophrenia. Thus, we conducted systematic research in five databases and in the grey literature to find articles investigating fALFF and/or ALFF alterations in schizophrenia. Two separate meta-analyses were performed using the SDM-PSI software to identify fALFF and ALFF alterations separately. Then, a conjunction analysis was conducted to determine congruent results between the two approaches. We found that patients with schizophrenia showed altered fALFF activity in the left insula/putamen, the right paracentral lobule and the left middle occipital gyrus compared to healthy individuals. Patients with schizophrenia exhibited ALFF alterations in the bilateral putamen, the bilateral caudate nucleus, the bilateral inferior frontal gyrus, the right precuneus, the right precentral gyrus, the left postcentral gyrus, the right posterior cingulate gyrus, compared to healthy controls. ALFF increased activity in the left putamen was higher in drug-naïve patients and was correlated with positive symptoms. The conjunction analysis revealed a spatial convergence between fALFF and ALFF studies in the left putamen. This left putamen cluster is part of the associative striatum. Its alteration in schizophrenia provides additional support to the influential aberrant salience hypothesis of psychosis.

Deletion of IRE1α in podocytes exacerbates diabetic nephropathy in mice.

Protein misfolding in the endoplasmic reticulum (ER) of podocytes contributes to the pathogenesis of glomerular diseases. Protein misfolding activates the unfolded protein response (UPR), a compensatory signaling network. We address the role of the UPR and the UPR transducer, inositol-requiring enzyme 1α (IRE1α), in streptozotocin-induced diabetic nephropathy in mice. Diabetes caused progressive albuminuria in control mice that was exacerbated in podocyte-specific IRE1α knockout (KO) mice. Compared to diabetic controls, diabetic IRE1α KO mice showed reductions in podocyte number and synaptopodin. Glomerular ultrastructure was altered only in diabetic IRE1α KO mice; the major changes included widening of podocyte foot processes and glomerular basement membrane. Activation of the UPR and autophagy was evident in diabetic control, but not diabetic IRE1α KO mice. Analysis of human glomerular gene expression in the JuCKD-Glom database demonstrated induction of genes associated with the ER, UPR and autophagy in diabetic nephropathy. Thus, mice with podocyte-specific deletion of IRE1α demonstrate more severe diabetic nephropathy and attenuation of the glomerular UPR and autophagy, implying a protective effect of IRE1α. These results are consistent with data in human diabetic nephropathy and highlight the potential for therapeutically targeting these pathways.

Dysconnectivity of the Nucleus Accumbens and Amygdala in Youths with Thought Problems: A Dimensional Approach.

Background: Youths with thought problems (TP) are at risk to develop psychosis and obsessive-compulsive disorder (OCD). Yet, the pathophysiological mechanisms underpinning TP are still unclear. Functional magnetic resonance imaging (fMRI) studies have shown that striatal and limbic alterations are associated with psychosis-like and obsessive-like symptoms in individuals at clinical risk for psychosis, schizophrenia, and OCD. More specifically, nucleus accumbens (NAcc) and amygdala are mainly involved in these associations. The current study aims to investigate the neural correlates of TP in youth populations using a dimensional approach and explore potential cognitive functions and neurotransmitters associated with it. Methods: Seed-to-voxels functional connectivity analyses using NAcc and amygdala as regions-of-interest were conducted with resting-state fMRI data obtained from 1360 young individuals, and potential confounders related to TP such as anxiety and cognitive functions were included as covariates in multiple regression analyses. Replicability was tested in using an adult cohort. In addition, functional decoding and neurochemical correlation analyses were performed to identify the associated cognitive functions and neurotransmitters. Results: The altered functional connectivities between the right NAcc and posterior parahippocampal gyrus, between the right amygdala and lateral prefrontal cortex, and between the left amygdala and the secondary visual area were the best predictors of TP in multiple regression model. These functional connections are mainly involved in social cognition and reward processing. Conclusions: The results show that alterations in the functional connectivity of the NAcc and the amygdala in neural pathways involved in social cognition and reward processing are associated with severity of TP in youths.

Analysis of gene expression and use of connectivity mapping to identify drugs for treatment of human glomerulopathies.

Background: Human glomerulonephritis (GN)-membranous nephropathy (MN), focal segmental glomerulosclerosis (FSGS) and IgA nephropathy (IgAN), as well as diabetic nephropathy (DN) are leading causes of chronic kidney disease. In these glomerulopathies, distinct stimuli disrupt metabolic pathways in glomerular cells. Other pathways, including the endoplasmic reticulum (ER) unfolded protein response (UPR) and autophagy, are activated in parallel to attenuate cell injury or promote repair. Methods: We used publicly available datasets to examine gene transcriptional pathways in glomeruli of human GN and DN and to identify drugs. Results: We demonstrate that there are many common genes upregulated in MN, FSGS, IgAN, and DN. Furthermore, these glomerulopathies were associated with increased expression of ER/UPR and autophagy genes, a significant number of which were shared. Several candidate drugs for treatment of glomerulopathies were identified by relating gene expression signatures of distinct drugs in cell culture with the ER/UPR and autophagy genes upregulated in the glomerulopathies ("connectivity mapping"). Using a glomerular cell culture assay that correlates with glomerular damage in vivo, we showed that one candidate drug - neratinib (an epidermal growth factor receptor inhibitor) is cytoprotective. Conclusion: The UPR and autophagy are activated in multiple types of glomerular injury. Connectivity mapping identified candidate drugs that shared common signatures with ER/UPR and autophagy genes upregulated in glomerulopathies, and one of these drugs attenuated injury of glomerular cells. The present study opens the possibility for modulating the UPR or autophagy pharmacologically as therapy for GN.

The unfolded protein response transducer IRE1α promotes reticulophagy in podocytes.

Glomerular diseases involving podocyte/glomerular epithelial cell (GEC) injury feature protein misfolding and endoplasmic reticulum (ER) stress. Inositol-requiring enzyme 1α (IRE1α) mediates chaperone production and autophagy during ER stress. We examined the role of IRE1α in selective autophagy of the ER (reticulophagy). Control and IRE1α knockout (KO) GECs were incubated with tunicamycin to induce ER stress and subjected to proteomic analysis. This showed IRE1α-dependent upregulation of secretory pathway mediators, including the coat protein complex II component Sec23B. Tunicamycin enhanced expression of Sec23B and the reticulophagy adaptor reticulon-3-long (RTN3L) in control, but not IRE1α KO GECs. Knockdown of Sec23B reduced autophagosome formation in response to ER stress. Tunicamycin stimulated colocalization of autophagosomes with Sec23B and RTN3L in an IRE1α-dependent manner. Similarly, during ER stress, glomerular α5 collagen IV colocalized with RTN3L and autophagosomes. Degradation of RTN3L and collagen IV increased in response to tunicamycin, and the turnover was blocked by deletion of IRE1α; thus, the IRE1α pathway promotes RTN3L-mediated reticulophagy and collagen IV may be an IRE1α-dependent reticulophagy substrate. In experimental glomerulonephritis, expression of Sec23B, RTN3L, and LC3-II increased in glomeruli of control mice, but not in podocyte-specific IRE1α KO littermates. In conclusion, during ER stress, IRE1α redirects a subset of Sec23B-positive vesicles to deliver RTN3L-coated ER fragments to autophagosomes. Reticulophagy is a novel outcome of the IRE1α pathway in podocytes and may play a cytoprotective role in glomerular diseases.

Loss of Planar Cell Polarity Effector Fuzzy Causes Renal Hypoplasia by Disrupting Several Signaling Pathways.

In vertebrates, the planar cell polarity (PCP) pathway regulates tissue morphogenesis during organogenesis, including the kidney. Mutations in human PCP effector proteins have been associated with severe syndromic ciliopathies. Importantly, renal hypoplasia has been reported in some patients. However, the developmental disturbance that causes renal hypoplasia is unknown. Here, we describe the early onset of profound renal hypoplasia in mice homozygous for null mutation of the PCP effector gene, Fuzzy. We found that this phenotype is caused by defective branching morphogenesis of the ureteric bud (UB) in the absence of defects in nephron progenitor specification or in early steps of nephrogenesis. By using various experimental approaches, we show that the loss of Fuzzy affects multiple signaling pathways. Specifically, we found mild involvement of GDNF/c-Ret pathway that drives UB branching. We noted the deficient expression of molecules belonging to the Bmp, Fgf and Shh pathways. Analysis of the primary cilia in the UB structures revealed a significant decrease in ciliary length. We conclude that renal hypoplasia in the mouse Fuzzy mutants is caused by defective UB branching associated with dysregulation of ciliary and non-ciliary signaling pathways. Our work suggests a PCP effector-dependent pathogenetic mechanism that contributes to renal hypoplasia in mice and humans.

Role of IRE1α in podocyte proteostasis and mitochondrial health.

Glomerular epithelial cell (GEC)/podocyte proteostasis is dysregulated in glomerular diseases. The unfolded protein response (UPR) is an adaptive pathway in the endoplasmic reticulum (ER) that upregulates proteostasis resources. This study characterizes mechanisms by which inositol requiring enzyme-1α (IRE1α), a UPR transducer, regulates proteostasis in GECs. Mice with podocyte-specific deletion of IRE1α (IRE1α KO) were produced and nephrosis was induced with adriamycin. Compared with control, IRE1α KO mice had greater albuminuria. Adriamycin increased glomerular ER chaperones in control mice, but this upregulation was impaired in IRE1α KO mice. Likewise, autophagy was blunted in adriamycin-treated IRE1α KO animals, evidenced by reduced LC3-II and increased p62. Mitochondrial ultrastructure was markedly disrupted in podocytes of adriamycin-treated IRE1α KO mice. To pursue mechanistic studies, GECs were cultured from glomeruli of IRE1α flox/flox mice and IRE1α was deleted by Cre-lox recombination. In GECs incubated with tunicamycin, deletion of IRE1α attenuated upregulation of ER chaperones, LC3 lipidation, and LC3 transcription, compared with control GECs. Deletion of IRE1α decreased maximal and ATP-linked oxygen consumption, as well as mitochondrial membrane potential. In summary, stress-induced chaperone production, autophagy, and mitochondrial health are compromised by deletion of IRE1α. The IRE1α pathway is cytoprotective in glomerular disease associated with podocyte injury and ER stress.

Intrinsic tumor necrosis factor-α pathway is activated in a subset of patients with focal segmental glomerulosclerosis.

Focal segmental glomerulosclerosis (FSGS) is frequently found in biopsies of patients with steroid resistant nephrotic syndrome (SRNS). The pathogenesis of SRNS/FSGS is often unknown and the disease will recur in up to 50% of patients post-transplant, indicating the presence of circulating podocyte-toxic factor(s). Several studies have reported clinical improvement after anti-TNFα therapy. However, prediction of the clinical outcome in SRNS/FSGS is difficult, and novel predictive biomarkers are needed. An image-based assay, which measures disassembly of focal adhesion complexes in cultured podocytes, was used to ascertain the presence of podocyte toxic activity in SRNS/FSGS sera. Expression of TNFα pathway genes was analysed in the Nephroseq FSGS cohort and in cultured podocytes treated with SRNS/FSGS sera. Podocyte toxic activity was detected in 48/96 SRNS/FSGS patients. It did not correlate with serum TNFα levels, age, sex, ethnicity or glomerular filtration rate. In ~25% of the toxic samples, the toxicity was strongly inhibited by blockade of TNFα signaling. Transcriptional profiling of human FSGS biopsies and podocytes treated with FSGS sera revealed significant increases in expression of TNFα pathway genes. We identified patients with serum podocyte toxic activity who may be at risk for FSGS recurrence, and those patients in whom serum podocyte toxicity may be reversed by TNFα blockade. Activation of TNFα pathway genes occurs in podocytes of FSGS patients suggesting a causative effect of this pathway in response to circulating factor(s). In vitro analyses of patient sera may stratify patients according to prognostic outcomes and potential responses to specific clinical interventions.

Disease stage-dependent relationship between diffusion tensor imaging and electrophysiology of the visual system in a murine model of multiple sclerosis.

PURPOSE: Diffusion tensor imaging (DTI) is commonly used to evaluate white matter integrity in multiple sclerosis (MS), but the relationship between DTI measures and functional changes during disease remains ambiguous. Using a mouse model of MS, we tested the hypothesis that DTI measures would correlate to the visual evoked potential (VEPs) dynamically at different disease stages. METHODS: In vivo DTI, gadolinium-enhanced T1WI (Gd-T1WI) and VEPs were performed in 5 control and 25 mice after 2-12 weeks of experimental autoimmune encephalomyelitis (EAE). DTI indices, including fractional anisotropy (FA), axial and radial diffusivities (AD and RD), and Gd-T1WI enhancement, were measured in the optic nerve and tract (ON and OT), which were compared with measured VEPs. RESULTS: Gd-T1WI showed a 3- to 4-fold enhancement over controls beginning after 2 weeks of EAE. Across the time course, we found progressive reductions in FA and increases in RD with increases in VEP latency and reductions in amplitude. Significant correlations between DTI (FA and RD) and VEP evolved; in control/early asymptomatic EAE mice, both FA and RD were highly correlated with VEP latency (but not amplitude), while in late EAE, both DTI indices were highly correlated with VEP amplitude (but not latency). CONCLUSION: DTI measures FA and RD are associated to VEP latency in early stages of EAE but associated to VEP amplitude in later stages, suggesting that the patterns of DTI related to the functional decline may depend on the stage of disease progression.

Anterograde-propagation of axonal degeneration in the visual system of wlds mice characterized by diffusion tensor imaging.

PURPOSE: To evaluate the feasibility of using diffusion tensor imaging (DTI) to characterize the temporospatial profile of axonal degeneration and its relation to blood-brain barrier (BBB) permeability. MATERIALS AND METHODS: Longitudinal DTI was performed in Wallerian degeneration slow (WldS) mice following retinal ischemia. In parallel, gadolinium (Gd)-enhanced T1 -weighted imaging (Gd-T1 WI) was performed to evaluate BBB permeability in white matter during axonal degeneration. To confirm the in vivo findings, immunohistochemistry using SMI-31 and myelin basic protein (MBP) was performed to examine the axons and myelin, respectively, and Evans blue was used to evaluate the permeability of the BBB. RESULTS: Reduced axial diffusivity was found in the optic nerve (ON, -15%, P = 0.0063) 1 week and optic tact (OT, -18%, P = 0.0077) 2 weeks after retinal ischemia, which were respectively associated with an 11% (P = 0.0116) and 25% (P = 0.0001) axonal loss. Increased radial diffusivity was found 1-2 weeks after the colocated decrease of axial diffusivity (35% increase, P = 0.0388 in the ON at week 2 and an 80% increase, P = 0.0015 in the OT at week 4). No significant changes were observed using Gd-T1 WI (P = 0.13-0.75), although an approximately 1-fold increase in Evans blue staining intensity was found in the injured ON and OT starting 1 week after retinal ischemia. CONCLUSION: We demonstrated the utility of DTI to characterize anterograde-propagating axonal degeneration through the ON and OT following retinal damage. Evans blue staining revealed serum albumin accumulation at injured sites, although there was no BBB leakage detectable using Gd-T1 WI. LEVEL OF EVIDENCE: 2 J. Magn. Reson. Imaging 2017;45:482-491.

Novel unbiased assay for circulating podocyte-toxic factors associated with recurrent focal segmental glomerulosclerosis.

Focal segmental glomerular sclerosis (FSGS) is an irreversible renal pathology characterized by podocyte detachment from the glomerular basement membrane, hyalinosis, and sclerosis. Clinically, it manifests with proteinuria and progressive loss of glomerular filtration. Primary idiopathic FSGS can occur in isolation and frequently progresses to end-stage renal disease, requiring dialysis or kidney transplantation. In 30-50% of these patients, proteinuria and FSGS recur in the renal allograft, suggesting the presence of a podocyte-toxic factor(s) in the recipient's serum. Currently, there is no reliable way to quantify the serum activity or predict the subset of FSGS patients at risk for recurrence after transplantation. We describe a novel in vitro method that measures the podocyte-toxic activity of sera from FSGS patients using cultured human podocytes; we compare this with the effect of compounds such as adriamycin. Using immunofluorescence microscopy followed by computerized image-processing analysis, we show that incubation of human podocytes with adriamycin leads to a dose-dependent disassembly of focal adhesion complexes (FACs). We then demonstrate that sera from patients with posttransplant recurrent or idiopathic FSGS cause a similar FAC disturbance. In contrast, sera from nonrecurrent FSGS patients do not affect FACs. In some FSGS patients, toxic effects of serum can be prevented by blockade of the tumor necrosis factor-α pathway. We propose that this method may be useful as a diagnostic tool to identify FSGS patients with serum podocyte-toxic activity that presumably places them at increased risk for recurrence in the renal allograft.