Dual-monomer solvatochromic probe system (DSPS) for effectively differentiating lipid raft cholesterol and active membrane cholesterol in the inner-leaflet plasma membrane.

Monitoring active membrane cholesterol and lipid raft cholesterol in the inner leaflet of the plasma membrane is significant for understanding the membrane function and cellular physiopathological processes. Limited by existing methods, it is difficult to differentiate active membrane cholesterol and lipid raft cholesterol. A novel dual-monomer solvatochromic probe system (DSPS) that targets two types of cholesterol was developed. Acrylodan-BG/SNAP-D4 composed of SNAP-D4 cholesterol-recognizing monomers and solvatochromic acrylodan-BG-sensing monomers exhibits excellent cholesterol detecting properties in terms of selectivity, accuracy, convenience and economic benefits. Cell imaging revealed that lipid raft cholesterol emitted blue fluorescence, whereas active membrane cholesterol (which partially bobbed in aqueous cytosol) displayed green fluorescence; both the fluorescence emissions increased or decreased in a cholesterol-dependent manner. This system provides a new technology for the determination of two types of cholesterol, which is beneficial for the further study of membrane function, intracellular cholesterol trafficking, and cell signaling.

Pigment epithelium-derived factor may induce antidepressant phenotypes in mice by the prefrontal cortex.

Pigment epithelium-derived factor (PEDF) is a multifunctional glycoprotein encoded by SERPINF1 and our previous study reported that PEDF may have antidepressant effects. As a key brain region regulating cognition, memory and emotion, the prefrontal cortex (PFC) has been studied extensively in major depressive disorder (MDD), but there are few reports on the relationship between PEDF and the PFC. In this study, enzyme-linked immunosorbent assay showed that the PEDF level was decreased in the plasma of MDD patients compared with that of healthy controls. Western blotting validated that the PEDF expression in the PFC was downregulated in the mouse chronic social defeat stress and rat chronic unpredictable mild stress models of depression. Correspondingly, normal mice overexpressing PEDF in the PFC showed depression-resistant phenotypes. We detected PFC metabolite levels by liquid chromatography-tandem mass spectrometry and found significant upregulation of 5-hydroxyindoleacetic acid, kynurenine, 5-hydroxytryptamine, ornithine and glutamine, and downregulation of 5-hydroxytryptophan, glutamic acid and aspartic acid in PEDF-overexpressing mice compared with control mice, in which no such changes were detected. Combined with the above findings, this provides an insight into a potential mechanism of the antidepressant effects of PEDF via the PFC, which may help to improve understanding of depression pathophysiology.

Ferritin disorder in the plasma and hippocampus associated with major depressive disorder.

Major depressive disorder (MDD) is a debilitating mental illness that can cause significant emotional disturbances and severe socioeconomic burdens. Rodent and nonhuman primate-based depression models have been studied, such as brain-derived neurotrophic factor (BDNF) and monoamine acid disorder hypotheses, as well as peripheral microbiota disturbances causing MDD; however, the pathogenesis is still largely unknown. This study aims to explore the relationship between ferritin and MDD. First, alterations in ferritin, including ferritin light chain (FTL) and ferritin heavy chain (FTH), in MDD patient plasma compared with healthy control (HC) plasma were detected using ELISA. Then, serum ferritin expression in cLPS-depressed mice was measured by ELISA. The existence of FTH in the hippocampus was validated by immunofluorescence, and the change in FTH levels in the hippocampus of mice injected with cLPS was detected by western blotting. FTL levels in MDD patients were decreased compared with those in HCs. In cLPS-depressed mice, serum ferritin was not different from that in the control group, while the expression of FTH in the hippocampus was significantly reduced in depressed mice. Our findings demonstrate the alteration of ferritin expression in MDD and provide new insight into the pathogenesis of MDD.

Non-targeted Metabolomics Profiling of Plasma Samples From Patients With Major Depressive Disorder.

Background: Major depressive disorder (MDD) is a neuropsychiatric disorder caused by multiple factors. Although there are clear guidelines for the diagnosis of MDD, the direct and objective diagnostic methods remain inadequate thus far. Methods: This study aims to discover peripheral biomarkers in patients with MDD and promote the diagnosis of MDD. Plasma samples of healthy controls (HCs, n = 52) and patients with MDD (n = 38) were collected, and then, metabolism analysis was performed using ultrahigh-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). Heatmap analysis was performed to identify the different metabolites. Meanwhile, receiver operating characteristic (ROC) curves of these differential metabolites were generated. Results: Six differential metabolites were found by LC-MS/MS analysis. Three of these were increased, including L-aspartic acid (Asp), diethanolamine, and alanine. Three were decreased, including O-acetyl-L-carnitine (LAC), cystine, and fumarate. In addition, LAC, Asp, fumarate, and alanine showed large areas under the curve (AUCs) by ROC analysis. Conclusion: The study explored differences in peripheral blood between depressed patients and HCs. These results indicated that differential metabolites with large AUCs may have the potential to be promising biomarkers for the diagnosis of MDD.

Alterations of neurotransmitters and related metabolites in the habenula from CUMS-susceptible and -resilient rats.

Although major depressive disorder (MDD) has caused severe mental harm to overwhelming amounts of patients, the pathogenesis of MDD remains to be studied. Due to the in-depth discussion of the mechanism of new antidepressants like ketamine, the habenula (Hb) was reported to be significant in the onset of MDD and the antidepressant mechanism. In the Hb of depressive-like rodents, various molecular mechanisms and neuronal electrical activities have been reported, but neurotransmitters disorder in response to stress are still unclear. Thus, we divided stress-susceptible and stress-resilient rats after exposure to chronic unpredictable mild stress (CUMS). Multiple metabolites in the Hb were determined by liquid chromatography-tandem mass spectrometry. Based on this approach, we found that glutamate was significantly increased in susceptible group and resilient group, while dopamine was significant decreased in two groups. Gamma-aminobutyric acid was significantly upregulated in susceptible group but downregulated in resilient group. Our study firstly provides quantitative evidence regarding alterations of main neurotransmitters in the Hb of CUMS rats, showing the different role of neurotransmitters in stress susceptibility and stress resilience.

iTRAQ-based proteomics implies inflammasome pathway activation in the prefrontal cortex of CSDS mice may influence resilience and susceptibility.

AIMS: Major depressive disorder, as a destructive mental health disorder, is a major contributor to disability and death. Numerous studies have illustrated that activation of inflammation and fluctuating immune reactions play a crucial role in the physiopathology of depression. The effectiveness of antidepressants is affected by the intensity of the inflammatory response. Thus, we aim to reveal the correlation of inflammatory factors and depression. MAIN METHODS: Isobaric tags for relative and absolute quantitation (iTRAQ™)-based proteomics was applied to verify the quantitation of target proteins in the PFC of chronic social defeat stress (CSDS) model mice. Ingenuity pathway analysis (IPA) was performed to explore related pathways, and the involvement of molecules was validated by western blotting and real time-quantitative polymerase chain reaction (RT-qPCR). KEY FINDINGS: According to the IPA results, CSDS-susceptible mice and CSDS-resilient mice both exhibited alterations of the inflammasome pathway in the PFC. Compared with control mice, susceptible mice subjected to CSDS showed an increased ATP-activated purinergic receptor P2X7 (also known as P2RX7) protein level. Nevertheless, the expression levels of cysteinyl aspartate-specific protease 1 (Caspase 1) and apoptosis-associated speck-like protein containing a CARD (ASC) were reduced in CSDS mice, and downregulation of interleukin-1ß (IL-1ß) was found in susceptible mice. Moreover, no significant difference was found in nuclear factor-κB levels among the three groups. SIGNIFICANCE: CSDS administration leads to dysfunctions of key molecules in the inflammasome pathway, promoting depressive-like behaviors in mice.

Major depression accompanied with inflammation and multiple cytokines alterations: Evidences from clinical patients to macaca fascicularis and LPS-induced depressive mice model.

BACKGROUND: Inflammation progress has been consistently implicated in the pathophysiology of major depressive disorder (MDD). However, the underlying mechanism of inflammation and depressive symptoms still far from being fully elucidated. In addition, studies on emotional disorders could also benefit from model of the non-human primates. To explore the difference of serum multi-cytokines levels among the MDD patients and depressed macaca fascicularis as well as LPS-treated mice, thus may find the reliable potential biomarkers for MDD. METHODS: Serum multi-cytokines levels among MDD patients (n = 44) and depressed macaca fascicularis (n = 6) together with controls (n = 22 for human, n = 6 for macaques) were detected by the Bio-Plex cytokines panel. Then five of these serum cytokines in LPS-treated mice were measured via ELISA. Furthermore, these cytokines protein expressions were validated by western blotting in three depression-related regions of LPS-treated mice. RESULTS: Here, we found that MDD patients displayed increased concentration of 13 proinflammatory and anti-inflammatory cytokines accompanied with one decreased cytokine in peripheral serum. Meanwhile, the naturally occurring depression (NOD) macaca fascicularis merely exhibited elevated concentration of 4 peripheral cytokines (IL-6, IL-8, MCP-1, VEGF), which were in accordance with the outcomes of MDD patients. Importantly, the serum and brain tissues of LPS-treated mice also emerged similar cytokines alterations. CONCLUSION: In summary, our findings strengthen the evidence that cytokines were associated with the depression, and the IL-6 and VEGF may as predictive biomarkers for novel diagnostic as well as therapeutic of depression. The hypothalamus may as a key brain region involve in the inflammatory related depressive-like behaviors.

Metabolomic abnormalities of purine and lipids implicated olfactory bulb dysfunction of CUMS depressive rats.

Major depressive disorder (MDD) is a serious mood disorder and leads to a high suicide rate as well as financial burden. The volume and function (the sensitivity and neurogenesis) of the olfactory bulb (OB) were reported to be altered among the MDD patients and rodent models of depression. In addition, the olfactory epithelium was newly reported to decrease its volume and function under chronic unpredictable mild stress (CUMS) treatment. However, the underlying molecular mechanism still remains unclear. Herein, we conducted the non-targeted metabolomics method based on gas chromatography-mass spectrometry (GC-MS) coupled with multivariate statistical analysis to characterize the differential metabolites in OB of CUMS rats. Our results showed that 19 metabolites were categorized into two perturbed pathways: purine metabolism and lipid metabolism, which were regarded as the vital pathways concerned with dysfunction of OB. These findings indicated that the turbulence of metabolic pathways may be partly responsible for the dysfunction of OB in MDD.

Entorhinal cortex-based metabolic profiling of chronic restraint stress mice model of depression.

Despite that millions of people suffer from major depressive disorder (MDD), the mechanism underlying MDD remains elusive. Recently, it has been reported that entorhinal cortex (EC) functions on the regulation of depressive-like phenotype relying on the stimulation of glutamatergic afferent from EC to hippocampus. Based on this, we used liquid chromatography-tandem mass spectrometry method to explore metabolic alterations in the EC of mice after exposed to chronic restraint stress (CRS). Molecular validation was conducted via the application of western blot and RT-qPCR. Through this study, we found significant upregulation of glutamate, ornithine aspartic acid, 5-hydroxytryptophan, L-tyrosine and norepinephrine in CRS group, accompanied with downregulation of homovanillic acid. Focusing on these altered metabolic pathways in EC, we found that gene levels of GAD1, GLUL and SNAT1 were increased. Upregulation of SERT and EAAT2 in protein expression level were also validated, while no significant changes were found in TH, AADC, MAOA, VMAT2, GAD1, GLUL and SNAT1. Our findings firstly provide evidence about the alteration of metabolites and related molecules in the EC of mice model of depression, implying the potential mechanism in MDD pathology.

Different Serotypes of Adeno-Associated Virus Vector- and Lentivirus-Mediated Tropism in Choroid Plexus by Intracerebroventricular Delivery.

Regulation of gene expression by viral vectors is an effective method for researchers to explore the function of gene products in a target tissue. The choroid plexus (CP) is an important target for gene therapy of neuropsychiatric diseases such as Alzheimer's disease and major depressive disorder. However, viral tropism in CP has not been well studied as a result of limited viral vector applications. To identify CP-specific viral vectors, we intracerebroventricularly administered six different serotypes of adeno-associated virus (AAV) vectors (AAV2/1, AAV2/5, AAV2/8, AAV2/9, AAV2-BR1, and AAV2-PHP.eB) and lentivirus in adult mice. Tropism in CP was compared among these viruses. We found that AAV2/5 and AAV2/8 displayed remarkable infections in CP, while AAV2/1 infected both ependymal cells and cells in the CP. Except for the low infection intensity of AAV2/9 and lentivirus in the CP, no infection intensity was found for CP tissues injected with AAV2-BR1 or AAV2-PHP.eB. Green fluorescence protein expression in the CP after AAV2/5 infection was confirmed by Western blotting. AAV2/5-mediated tropism in epithelial cells of the CP was verified by immunostaining with transthyretin. In this study, we identified for the first time that serotype-specific AAVs 5 and 8 may be robust research tools for intracerebroventricular gene delivery.

The 25(OH)D/VDR signaling may play a role in major depression.

Although the current evidences may suggest that the 25(OH)D associated with depression, still there exists conflicting results. In addition, little known is concerning the relationship between the 25(OH)D and the chronic stress-induced depressive-like behaviors. We detected the 25(OH)D levels in serum and the VDR protein expression in different brain regions aiming to explore the relationship between 25(OH)D/VDR signaling and major depression. The chemiluminescent microparticle immunoassay (CMIA) was used to detect the serum concentration of 25(OH)D in patients, the enzyme-linked immunosorbent assay (ELISA) was applied to measure the serum 25(OH)D levels in both CRS-treated and CSDS-treated mice models of MDD. Meanwhile, the VDR protein expression levels were validated among three MDD related brain regions from CRS-treated mice by western blotting. In this study, we mainly observed that the concentration of the 25(OH)D was decreased in the serum of MDD patients comparing to healthy controls. Consistent with the clinical findings, the CRS-treated mice also displayed down-regulated 25(OH)D level comparing with control mice. While in the CSDS model, the serum 25(OH)D status of depressive mice remained unchanged. Moreover, we found the protein level of VDR was significantly decreased in the hippocampus while increased in the hypothalamus of CRS-treated mice. Nevertheless, the prefrontal cortex exhibited no change regarding VDR protein expression compared with control mice. Taken together, these findings further confirmed that the 25(OH)D together with VDR may involve in the pathophysiological mechanism of depression-like behaviors induced by chronic stress.

Validation of the targeted metabolomic pathway in the hippocampus and comparative analysis with the prefrontal cortex of social defeat model mice.

The proportion of major depressive disorder (MDD) patients around the world has increased remarkably. Although many studies of MDD have been conducted based on classic hypotheses, like alteration of the hypothalamic-pituitary-adrenal axis or monoamine neurotransmitters, the mechanisms underlying MDD remain unclear. Aiming to further investigate the mechanisms of MDD, liquid chromatography-tandem mass spectrometry was employed to measure target metabolites in the hippocampus (HIPPO) of chronic social defeat stress model mice. Compared with control mice, stress-susceptible mice showed a reduction of 5-hydroxyindoleacetic acid and kynurenic acid in the tryptophan pathway, and an increased level of dopamine in the catecholamine pathway, while stress-resilient mice displayed a reduction of 5-hydroxytryptamine in the tryptophan pathway. The altered levels of key molecules related to the tryptophan or dopamine metabolic pathways were validated by real-time quantitative polymerase chain reaction or western blotting. Comparative analysis with previous targeted metabolomics results in the prefrontal cortex (PFC) of chronic social defeat stress mice revealed that the altered metabolites manifested in specific brain areas, and only the dopamine metabolic pathway was perturbed in both the HIPPO and PFC after stress. Additionally, correlation analysis validated that levels of kynurenic acid in the HIPPO, along with glutamic acid, L-3, 4-dihydroxyphenylalanine, and vanillylmandelic acid in the PFC, were correlated with depression-like behaviors. This study provides a unique perspective on the potential molecular mechanisms of stress susceptibility and stress resilience.