Downregulation of Ambra1 by altered DNA methylation exacerbates dopaminergic neuron damage in a fenpropathrin-induced Parkinson-like mouse model.

Fenpropathrin (Fen), a volatile pyrethroid insecticide, is used widely for agricultural applications and has been reported to increase the risk of Parkinson's disease (PD). However, the molecular basis, underlying mechanisms, and pathophysiology of Fen-exposed Parkinsonism remain unknown. Recent studies have revealed epigenetic mechanisms underlying PD-related pathway regulation, including DNA methylation. Epigenetic mechanisms are potential targets for therapeutic intervention in neurodegenerative diseases. After whole-genome bisulfite sequencing (WGBS) of midbrain tissues from a Fen-exposed PD-like mouse model, we performed an association analysis of DNA methylation and gene expression. Then we successfully screened for the DNA methylation differential gene Ambra1, which is closely related to PD. The hypermethylation-low expression Ambra1 gene aggravated DA neuron damage in vitro and in vivo through the Ambra1/Parkin/LC3B-mediated mitophagy pathway. We administered 5-aza-2'-deoxycytidine (5-Aza-dC) to upregulate Ambra1 expression, thereby reducing Ambra1-mediated mitophagy and protecting DA neurons against Fen-induced damage. In conclusion, these findings elucidate the potential function of Ambra1 under the regulation of DNA methylation, suggesting that the inhibition of DNA methylation may alleviate Fen-exposed neuron damage.

Small extracellular vesicles: a novel drug delivery system for neurodegenerative disorders.

Neurodegenerative diseases (NDs) have a slow onset and are usually detected late during disease. NDs are often difficult to cure due to the presence of the blood-brain barrier (BBB), which makes it difficult to find effective treatments and drugs, causing great stress and financial burden to families and society. Currently, small extracellular vesicles (sEVs) are the most promising drug delivery systems (DDSs) for targeted delivery of molecules to specific sites in the brain as a therapeutic vehicle due to their low toxicity, low immunogenicity, high stability, high delivery efficiency, high biocompatibility and trans-BBB functionality. Here, we review the therapeutic application of sEVs in several NDs, including Alzheimer's disease, Parkinson's disease, and Huntington's disease, discuss the current barriers associated with sEVs and brain-targeted DDS, and suggest future research directions.

Early antidepressant treatment response prediction in major depression using clinical and TPH2 DNA methylation features based on machine learning approaches.

OBJECTIVE: To identify DNA methylation and clinical features, and to construct machine learning classifiers to assign the patients with major depressive disorder (MDD) into responders and non-responders after a 2-week treatment into responders and non-responders. METHOD: Han Chinese patients (291 in total) with MDD comprised the study population. Datasets contained demographic information, environment stress factors, and the methylation levels of 38 methylated sites of tryptophan hydroxylase 2 (TPH2) genes in peripheral blood samples. Recursive Feature Elimination (RFE) was employed to select features. Five classification algorithms (logistic regression, classification and regression trees, support vector machine, logitboost and random forests) were used to establish the models. Performance metrics (AUC, F-Measure, G-Mean, accuracy, sensitivity, specificity, positive predictive value and negative predictive value) were computed with 5-fold-cross-validation. Variable importance was evaluated by random forest algorithm. RESULT: RF with RFE outperformed the other models in our samples based on the demographic information and clinical features (AUC = 61.2%, 95%CI: 60.1-62.4%) / TPH2 CpGs features (AUC = 66.6%, 95%CI: 65.4-67.8%) / both clinical and TPH2 CpGs features (AUC = 72.9%, 95%CI: 71.8-74.0%). CONCLUSION: The effects of TPH2 on the early-stage antidepressant response were explored by machine learning algorithms. On the basis of the baseline depression severity and TPH2 CpG sites, machine learning approaches can enhance our ability to predict the early-stage antidepressant response. Some potentially important predictors (e.g., TPH2-10-60 (rs2129575), TPH2-2-163 (rs11178998), age of first onset, age) in early-stage treatment response could be utilized in future fundamental research, drug development and clinical practice.

microRNAs profiling of small extracellular vesicles from midbrain tissue of Parkinson's disease.

Small extracellular vesicles (sEVs) are generated by all types of cells during physiological or pathological conditions. There is growing interest in tissue-derived small extracellular vesicles (tdsEVs) because they can be isolated from a single tissue source. Knowing the representation profile of microRNA (miRNA) in midbrain tissue-derived sEVs (bdsEVs) and their roles is imperative for understanding the pathological mechanism and improving the diagnosis and treatment of Parkinson's disease (PD). bdsEVs from a rat model of PD and a sham group were separated and purified using ultracentrifugation, size-exclusion chromatography (SEC), and ultrafiltration. Then, miRNA profiling of bdsEVs in both groups was performed using next-generation sequencing (NGS). The expression levels of 180 miRNAs exhibited significant differences between the two groups, including 114 upregulated and 66 downregulated genes in bdsEVs of PD rats compared with the sham group (p < 0.05). Targets of the differentially expressed miRNAs were predicted by miRanda and RNAhybrid, and their involvement in the signaling pathways and cellular function has been analyzed through the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO). Furthermore, we explored the expression levels of miR-103-3p, miR-107-3p, miR-219a-2-3p, and miR-379-5p in bdsEVs, sEVs derived from plasma, and plasma of both groups of rats. Interestingly, the expression levels of miR-103-3p, miR-107-3p, miR-219a-2-3p, and miR-379-5p were elevated in bdsEVs and sEVs from plasma; in contrast, their expression levels were decreased in plasma of the rat model of PD. In summary, miRNAs may play a significant role in the onset and development of PD, and miRNAs need to be selected carefully as a research subject for exploring the pathological mechanism and the potential therapeutic targets and diagnostic markers of PD.

Multiple roles of neuronal extracellular vesicles in neurological disorders.

Neuropathy is a growing public health problem in the aging, adolescent, and sport-playing populations, and the number of individuals at risk of neuropathy is growing; its risks include aging, violence, and conflicts between players. The signal pathways underlying neuronal aging and damage remain incompletely understood and evidence-based treatment for patients with neuropathy is insufficiently delivered; these are two of the reasons that explain why neuropathy is still not completely curable and why the progression of the disease cannot be inhibited. Extracellular vesicles (EVs) shuttling is an important pathway in disease progression. Previous studies have focused on the EVs of cells that support and protect neurons, such as astrocytes and microglia. This review aims to address the role of neuronal EVs by delineating updated mechanisms of neuronal damage and summarizing recent findings on the function of neuronal EVs. Challenges and obstacles in isolating and analyzing neuronal EVs are discussed, with an emphasis on neuron as research object and modification of EVs on translational medicine.

Comparison of the Variability of Small Extracellular Vesicles Derived from Human Liver Cancer Tissues and Cultured from the Tissue Explants Based on a Simple Enrichment Method.

A potential use of small extracellular vesicles (sEVs) for diagnostic and therapeutic purposes has recently generated a great interest. sEVs, when purified directly from various tissues with proper procedures, can reflect the physiological and pathological state of the organism. However, the quality of sEV is affected by many factors during isolation, including separation of sEV from cell and tissues debris, the use of enzymes for tissue digestion, and the storage state of tissues. In the present study, we established an assay for the isolation and purification of liver cancer tissues-derived sEVs (tdsEVs) and cultured explants-derived sEVs (cedsEVs) by comparing the quality of sEVs derived from different concentration of digestion enzyme and incubation time. The nano-flow cytometry (NanoFCM) showed that the isolated tdsEVs by our method are purer than those obtained from differential ultracentrifugation. Our study thus establishes a simple and effective approach for isolation of high-quality sEVs that can be used for analysis of their constituents.

Identifying patients with major depressive disorder based on tryptophan hydroxylase-2 methylation using machine learning algorithms.

OBJECTIVES: This study aimed to identify patients with major depressive disorder (MDD) by developing different machine learning (ML) models based on tryptophan hydroxylase-2 (TPH2) methylation and environmental stress. METHODS: The data were collected from 291 patients with MDD and 100 healthy control participants: individual basic information, the Negative Life Events Scale (NLES) scores, the Childhood Trauma Questionnaire (CTQ) scores and the methylation level at 38 CpG sites in TPH2. Information gain was used to select critical input variables. Support vector machine (SVM), back propagation neural network (BPNN) and random forest (RF) algorithms were used to build recognition models, which were evaluated by the 10-fold cross-validation. SHapley Additive exPlanations (SHAP) method was used to evaluate features importance. RESULTS: Gender, NLES scores, CTQ scores and 13 CpG sites in TPH2 gene were considered as predictors in the models. Three ML algorithms showed satisfactory performance in predicting MDD and the BPNN model indicated best prediction effects. CONCLUSION: ML models with TPH2 methylation and environmental stress were identified to possess great performance in identifying patients with MDD, which provided precious experience for artificial intelligence to assist traditional diagnostic methods in the future.

Short-term effect of fine particulate matter and ozone on non-accidental mortality and respiratory mortality in Lishui district, China.

BACKGROUND: In recent years, air pollution has become an imminent problem in China. Few studies have investigated the impact of air pollution on the mortality of the middle-aged and elderly people. Therefore, this study aims to evaluate the impact of PM2.5 (fine particulate matter) and O3 (ozone) on non-accidental mortality and respiratory mortality of the middle-aged and elderly people in Lishui District of Nanjing and provide the evidence for potential prevention and control measures of air pollution. METHOD: Using daily mortality and atmospheric monitoring data from 2015 to 2019, we applied a generalized additive model with time-series analysis to evaluate the association of PM2.5 and O3 exposure with daily non-accidental mortality and respiratory mortality in Lishui District. Using the population attributable fractions to estimate the death burden caused by short-term exposure to O3 and PM2.5。. RESULT: For every 10 µg/m3 increase in PM2.5, non-accidental mortality increased 0.94% with 95% confidence interval (CI) between 0.05 and 1.83%, and PM2.5 had a more profound impact on females than males. For every 10 µg/m3 increase in O3, respiratory mortality increased 1.35% (95% CI: 0.05, 2.66%) and O3 had a more profound impact on males than females. Compared with the single pollutant model, impact of the two-pollutant model on non-accidental mortality and respiratory mortality slightly decreased. In summer and winter as opposed to the other seasons, O3 had a more obvious impact on non-accidental mortality. The population attributable fractions of non-accidental mortality were 0.84% (95% CI:0.00, 1.63%) for PM2.5 and respiratory mortality were 0.14% (95% CI:0.01, 0.26%) for O3. For every 10 µg/m3 decrease in PM2.5, 122 (95% CI: 6, 237) non-accidental deaths could be avoided. For every 10 µg/m3 decrease in O3, 10 (95% CI: 1, 38) respiratory deaths could be avoided. CONCLUSION: PM2.5 and O3 could significantly increase the risk of non-accidental and respiratory mortality in the middle-aged and elderly people in Lishui District of Nanjing. Exposed to air pollutants, men were more susceptible to O3 damage, and women were more susceptible to PM2.5 damage. Reduction of PM2.5 and O3 concentration in the air may have the potential to avoid considerable loss of lives.

Role of autophagy and oxidative stress to astrocytes in fenpropathrin-induced Parkinson-like damage.

Fenpropathrin is an insecticide that is widely used in agriculture. It remains unknown whether fenpropathrin exposure increases the risk of Parkinson's disease. We found that fenpropathrin increased oxidative stress both in vitro and in vivo. Additionally, fenpropathrin increased production of ROS, NOS2, and HO-1, and decreased SOD and GSH in astrocytes. We further found that fenpropathrin-mediated oxidative stress might inhibit autophagic flow, including decreased expression of LC3A/B and enhanced expression of SQSTM1 via down-regulation of CDK-5, an upstream marker of autophagy. In mice, autophagy was slightly different from that found in astrocytes, as reflected in the increased expressions of LC3A/B and SQSTM1. Our findings elucidate the toxicological phenomena and pathogenic mechanisms of fenpropathrin and may provide guidance for improved pesticide control and environmental protection.

Fenpropathrin induces degeneration of dopaminergic neurons via disruption of the mitochondrial quality control system.

The synthetic pyrethroid derivative, fenpropathrin, is a widely used insecticide. However, a variety of toxic effects in mammals have been reported. In particular, fenpropathrin induces degeneration of dopaminergic neurons and parkinsonism. However, the mechanism of fenpropathrin-induced parkinsonism has remained unknown. In the present study, we investigated the toxic effects and underlying mechanisms of fenpropathrin on perturbing the dopaminergic system both in vivo and in vitro. We found that fenpropathrin induced cellular death of dopaminergic neurons in vivo. Furthermore, fenpropathrin increased the generation of reactive oxygen species, disrupted both mitochondrial function and dynamic networks, impaired synaptic communication, and promoted mitophagy in vitro. In mice, fenpropathrin was administered into the striatum via stereotaxic (ST) injections. ST-injected mice exhibited poor locomotor function at 24 weeks after the first ST injection and the number of tyrosine hydroxylase (TH)-positive cells and level of TH protein in the substantia nigra pars compacta were significantly decreased, as compared to these parameters in vehicle-treated mice. Taken together, our results demonstrate that exposure to fenpropathrin induces a loss of dopaminergic neurons and partially mimics the pathologic features of Parkinson's disease. These findings suggest that fenpropathrin may induce neuronal degeneration via dysregulation of mitochondrial function and the mitochondrial quality control system.

Generation of a Novel Mouse Model of Parkinson's Disease via Targeted Knockdown of Glutamate Transporter GLT-1 in the Substantia Nigra.

Parkinson's disease (PD) is a common neurodegenerative disease that is characterized by pathological dopaminergic (DA) neuronal death and α-synuclein aggregation. Glutamate excitotoxicity is a well-established pathogenesis of PD that involves dysfunctional expression of glutamate transporters. Glutamate transporter-1 (GLT-1) is mainly responsible for clearance of glutamate at synapses, including DA synapses. However, the role of GLT-1 in the aberrant synaptic transmission in PD remains elusive. In the present study, we generated small-interfering RNAs (siRNAs) to knockdown GLT-1 expression in primary astrocytes, and we report that siRNA knockdown of astrocytic GLT-1 decreased postsynaptic density-95 (PSD-95) expression in neuron-astrocyte cocultures in vitro. Using adeno-associated viruses (AAVs) targeting GLT-1 short-hairpin RNA (shRNA) sequences with a glial fibrillary acidic protein (GFAP) promoter, we abolished astrocytic GLT-1 expression in the substantia nigra pars compacta (SNpc) of mice. We found that GLT-1 deficiency in the SNpc induced parkinsonian phenotypes in terms of progressive motor deficits and nigral DA neuronal death in mice. We also found that there were reactive astrocytes and microglia in the SNpc upon GLT-1 knockdown. Furthermore, we used RNA sequencing to determine altered gene expression patterns upon GLT-1 knockdown in the SNpc, which revealed that disrupted calcium signaling pathways may be responsible for GLT-1 deficiency-mediated DA neuronal death in the SNpc. Taken together, our findings provide evidence for a novel role of GLT-1 in parkinsonian phenotypes in mice, which may contribute to further elucidation of the mechanisms of PD pathogenesis.

Regulatory Mechanism of miR-543-3p on GLT-1 in a Mouse Model of Parkinson's Disease.

Parkinson's disease (PD) features the degeneration and death of dopamine neurons in the substantia nigra pars compacta and the formation of Lewy bodies that contain α-synuclein. Among the numerous PD etiologies, glutamate excitotoxicity is a research hot spot, and glutamate transporters play key roles in this theory. It has been shown that the expression of the glutamate transporter is regulated by microRNAs. In this study, we found that the levels of expression and function of glutamate transporter type 1 (GLT-1) were significantly reduced and miR-543-3p was upregulated during the development of PD. Furthermore, our results indicated that GLT-1 plays an important role in the pathomechanism of PD. We found that miR-543-3p can suppress the expression and function of GLT-1 in MPP+-treated astrocytes and MPTP-treated mice. Inhibition of miR-543-3p can rescue the expression and function of GLT-1 and relieve dyskinesia in the PD model, which suggests that inhibition of miR-543-3p could serve as a potential therapeutic target for PD.

Gene Dysfunction Mediates Immune Response to Dopaminergic Degeneration in Parkinson's Disease.

Many publications reported that genetic dysfunction mediates abnormal immune responses in the brain, which is important for the development of neurodegenerative diseases, especially for Parkinson's disease (PD). This immune disorder results in subsequent inflammatory reaction, which stimulates microglia or other immune cells to secrete cytokines and chemokines and disturbs the proportion of peripheral blood lymphocyte subsets contributing to dopaminergic (DA) neuron apoptosis. Furthermore, the abnormal immune related signal pathways caused by genetic variants promote chronic inflammation destroying the blood-brain barrier, which allows infiltration of different molecules and blood cells into the central nervous system (CNS) exerting toxicity on DA neurons. As a result, the inflammatory reaction in the CNS accelerates the progression of Parkinson's disease and promotes α-synuclein aggregation and diffusion among DA neurons in the procession of Parkinson's disease. Thus, for disease evaluation, the genetic mediated abnormal immune response in PD may be assessed based on the multiple immune molecules and inflammatory factors, as well as the ratio of lymphocyte subsets from PD patient's peripheral blood as potential biomarkers.

ß N-O turns and helices induced by ß2-aminoxy peptides: synthesis and conformational studies.

Herein, we report an efficient route for the asymmetric synthesis of ß(2)-aminoxy acids as well as experimental and theoretical studies of conformations of peptides composed of ß(2)-aminoxy acids. The nine-membered-ring intramolecular hydrogen bonds, namely, ß N-O turns, are generated between adjacent residues in those peptides, in accordance with our computational results. The presence of two consecutive homochiral ß N-O turns leads to the formation of ß N-O helical structures in solution, although both helical (composed of two ß N-O turns of the same handedness) and reverse-turn (composed of two ß N-O turns with opposite handedness) structures are of similar stability, as suggested by theoretical studies. Nevertheless, two slightly different conformations, with the same handedness, of ß(2)-aminoxy monomers have been observed in the solid state and in solution according to our X-ray and 2D NOESY studies.

Disulfide bond creates a small connecting loop in aminoxy peptide backbone.

Disulfide-bond formation between the side chains of cysteine-cysteine pairs is often critical to the folding behavior, stability, and functionality of proteins. In this paper, we report that sulfur atoms can be introduced into the amide groups of aminoxy peptides to form a novel type of disulfide bridge, which creates a connecting loop in the peptide backbone.

[Features of heart rate, blood pressure, heart rate variability and cerebral blood flow during sequential head-up tilt test].

OBJECTIVE: To investigate the features of BP, HR, cerebral blood flow (CBF) and heart rate variation (HRV) during sequential head-up tilt table test (HUT) and provide information for methodology of testing tendency to syncope in pilots. METHOD: Sixteen healthy male were exposed to HUT +45 degrees 20 min, +60 degrees 20 min, and +75 degrees 20 min respectively with 5 min rests in between. CBFV, HR, HRV, BP were measured with transcranial doppler image (TCI), 12-Lead in phase ECG. RESULT: 1) As the tilt angle became larger, HR and DBP increased gradually (P<0.05), with SBP, PP and CBF Velocity (Vs,Vm,Vd) descended. There were significant differences among the various conditions. CV became smaller before +75 degrees, and then it began to increase. 2) Correlation between physiological indices and techno-indices were analyzed. At the point of 1 min, tilt angle was significantly positively correlated with SBP, DBP, MAP, Vd, and sdRR (P<0.01); At the point of 20 min, tilt angle was significantly positively correlated with HR and sdRR (P<0.01), but negatively correlated with SBP, PP, Vs and mRR (P<0.01), time of exposure was positively correlated with HP (P< 0.01), but negatively correlated with Vm, Vd, and mRR (P<0.01). 3) Correlation among physiological indices were also analyzed. HR were negatively correlated with SBP, Vs, Vm and Vd (P<0.01), SBP were significantly positively correlated with Vd, and mRR (P<0.01). Vs, Vm, Vd were significantly positively correlated with mRR and sdRR (P< 0.01). CONCLUSION: By investigating HR, BP, HRV and CBFV during of different angles, some advises for evaluating the tendency to syncope in pilots were put forward.