The effects of methylphenidate and atomoxetine on Drosophila brain at single-cell resolution and potential drug repurposing for ADHD treatment.

The stimulant methylphenidate (MPH) and the non-stimulant atomoxetine (ATX) are frequently used for the treatment of attention-deficit/hyperactivity disorder (ADHD); however, the function of these drugs in different types of brain cells and their effects on related genes remain largely unknown. To address these questions, we built a pipeline for the simultaneous examination of the activity behavior and transcriptional responses of Drosophila melanogaster at single-cell resolution following drug treatment. We selected the Drosophila with significantly increased locomotor activities (hyperactivity-like behavior) following the administration of each drug in comparison with the control (same food as the drug-treated groups with 5% sucrose, yeast, and blue food dye solution) using EasyFlyTracker. Subsequently, single cell RNA sequencing (scRNASEQ) was used to capture the transcriptome of 82,917 cells, unsupervised clustering analysis of which yielded 28 primary cell clusters representing the major cell types in adult Drosophila brain. Indeed, both neuronal and glial cells responded to MPH and ATX. Further analysis of differentially expressed genes (DEGs) revealed distinct transcriptional changes associated with these two drugs, such as two well-studied dopamine receptor genes (Dop2R and DopEcR) were responsive to MPH but not to ATX at their optimal doses, in addition to genes involved in dopamine metabolism pathways such as Syt1, Sytalpha, Syt7, and Ih in different cell types. More importantly, MPH also suppressed the expression of genes encoding other neurotransmitter receptors and synaptic signaling molecules in many cell types, especially those for Glu and GABA, while the responsive effects of ATX were much weaker. In addition to monoaminergic neuronal transmitters, other neurotransmitters have also shown a similar pattern with respect to a stronger effect associated with MPH than with ATX. Moreover, we identified four distinct glial cell subtypes responsive to the two drugs and detected a greater number of differentially expressed genes associated with ensheathing and astrocyte-like glia. Furthermore, our study provides a rich resource of candidate target genes, supported by drug set enrichment analysis (P = 2.10E-4; hypergeometric test), for the further exploration of drug repurposing. The whole list of candidates can be found at ADHDrug ( http://adhdrug.cibr.ac.cn/ ). In conclusion, we propose a fast and cost-efficient pipeline to explore the underlying molecular mechanisms of ADHD drug treatment in Drosophila brain at single-cell resolution, which may further facilitate drug repurposing applications.

rVarBase: an updated database for regulatory features of human variants.

We present here the rVarBase database (http://rv.psych.ac.cn), an updated version of the rSNPBase database, to provide reliable and detailed regulatory annotations for known and novel human variants. This update expands the database to include additional types of human variants, such as copy number variations (CNVs) and novel variants, and include additional types of regulatory features. Now rVarBase annotates variants in three dimensions: chromatin states of the surrounding regions, overlapped regulatory elements and variants' potential target genes. Two new types of regulatory elements (lncRNAs and miRNA target sites) have been introduced to provide additional annotation. Detailed information about variants' overlapping transcription factor binding sites (TFBSs) (often less than 15 bp) within experimentally supported TF-binding regions (∼ 150 bp) is provided, along with the binding motifs of matched TF families. Additional types of extended variants and variant-associated phenotypes were also added. In addition to the enrichment in data content, an element-centric search module was added, and the web interface was refined. In summary, rVarBase hosts more types of human variants and includes more types of up-to-date regulatory information to facilitate in-depth functional research and to provide practical clues for experimental design.

Fibroblast growth factor (FGF21) protects mouse liver against D-galactose-induced oxidative stress and apoptosis via activating Nrf2 and PI3K/Akt pathways.

FGF21 is recently discovered with pleiotropic effects on glucose and lipid metabolism. However, the potential protective effect of FGF21 against D-gal-induced injury in the liver has not been demonstrated. The aim of this study is to investigate the pathophysiological role of FGF21 on hepatic oxidative injury and apoptosis in mice induced by D-gal. The 3-month-old Kunming mice were subcutaneously injected with D-gal (180 mg kg(-1) d(-1)) for 8 weeks and administered simultaneously with FGF21 (5 or 1 mg kg(-1) d(-1)). Our results showed that the administration of FGF21 significantly alleviated histological lesion including structure damage, degeneration, and necrosis of hepatocytes induced by D-gal, and attenuated the elevation of liver injury markers, serum AST, and ALP in a dose-dependent manner. FGF21 treatment also suppressed D-gal-induced profound elevation of ROS production and oxidative stress, as evidenced by an increase of the MDA level and depletion of the intracellular GSH level in the liver, and restored the activities of antioxidant enzymes SOD, CAT, GSH-Px, and T-AOC. Moreover, FGF21 treatment increased the nuclear abundance of Nrf2 and subsequent up regulation of several antioxidant genes. Furthermore, a TUNEL assay showed that D-gal-induced apoptosis in the mouse liver was significantly inhibited by FGF21. The expression of caspase-3 was markedly inhibited by the treatment of FGF21 in the liver of D-gal-treated mice. The levels of PI3K and PBK/Akt were also largely enhanced, which in turn inactivated pro-apoptotic signaling events, restoring the balance between pro- and anti-apoptotic Bcl-2 and Bax proteins in the liver of D-gal-treated mice. In conclusion, these results suggest that FGF21 protects the mouse liver against D-gal-induced hepatocyte oxidative stress via enhancing Nrf2-mediated antioxidant capacity and apoptosis via activating PI3K/Akt pathway.

[Effect of FGF-21 on learning and memory ability and antioxidant capacity in brain tissue of D-galactose-induced aging mice].

This study aims to investigate the effects of fibroblast growth factor 21 (FGF-21) on learning and memory abilities and antioxidant capacity of D-galactose-induced aging mice. Kunming mice (37.1 +/- 0.62) g were randomly divided into normal control group, model group and FGF-21 high, medium and low dose groups (n = 8). Each group was injected in cervical part subcutaneously with D-galactose 180 mg x kg(-1) x d(-1) once a day for 8 weeks. At the same time, FGF-21-treated mice were administered with FGF-21 by giving subcutaneous injection in cervical part at the daily doses of 5, 2 and 1 mg x kg(-1) x d(-1). The normal control group was given with normal saline by subcutaneous injection in cervical part. At seventh week of the experiment, the learning and memory abilities of mice were determined by water maze and jumping stand tests. At the end of the experiment, the mice were sacrificed and the cells damage of hippocampus was observed by HE staining in each group. Reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) and total antioxidant capacity (T-AOC) in the brain of mice were determined. The results showed that different doses of FGF-21 could reduce the time reaching the end (P < 0.01 or P < 0.05) and the number of touching blind side (P < 0.01 or P < 0.05) in the water maze comparing with the model group. It could also prolong the latency time (P < 0.05) and decrease the number of errors (P < 0.01 or P < 0.05) in the step down test. The result of HE staining showed that FGF-21 could significantly reduce brain cell damage in the hippocampus. The ROS and MDA levels of three different doses FGF-21 treatment group reduced significantly than that of the model group [(5.58 +/- 1.07), (7.78 +/- 1.92), (9.03 +/- 1.77) vs (12.75 +/- 2.02) pmol (DCF) x min(-1) x mg(-1), P < 0.01 or P < 0.05], [(2.92 +/- 0.71), (4.21 +/- 0.81), (4.41 +/- 0.97) vs (5.62 +/- 0.63) nmol x mg(-1) (protein), P < 0.01]. Comparing with the model group, the activities of SOD, GPx, CAT and T-AOC of the three different doses FGF-21 treatment groups were also improved in a dose-dependent manner. This study demonstrates that FGF-21 can ameliorate learning and memory abilities of D-galactose induced aging mice, improve the antioxidant abilities in brain tissue and delay brain aging. This finding provides a theoretical support for clinical application of FGF-21 as a novel therapeutics for preventing aging.

MOASL: Predicting drug mechanism of actions through similarity learning with transcriptomic signature.

Understanding the mechanisms of actions (MOAs) of compounds is crucial in drug discovery. A common step in drug MOAs annotation is to query the dysregulated gene signatures induced by drugs in a reference library of pre-defined signatures. However, traditional similarity-based computational strategies face challenges when dealing with high-dimensional and noisy transcriptional signature data. To address this issue, we introduce MOASL (MOAs prediction via Similarity Learning), a novel approach that contrastive to learn similarity embeddings among signatures with shared MOAs automatically. We evaluated the accuracy of signature matching on various transcriptional activity score (TAS) datasets and individual cell lines by using MOASL. The results show MOASL achieved higher performance over several statistical and machine learning methods. Furthermore, we provided the rationale of our model by visualizing the signature annotation procedure. Using MOASL, the MOAs label of query signature could be conveniently defined by calculating the similarity between the query embedding and the reference embeddings. Finally, we applied MOASL to repurpose thousands of compounds as glucocorticoid receptor (GR) agonists, accurately identifying 8 out of the top 10 compounds. MOASL is conveniently accessible on GitHub at https://github.com/jianglikun/MOASL, empowering researchers and practitioners in the field of drug discovery to predict the MOAs of drug.

EasyFlyTracker: A Simple Video Tracking Python Package for Analyzing Adult Drosophila Locomotor and Sleep Activity to Facilitate Revealing the Effect of Psychiatric Drugs.

The mechanism of psychiatric drugs (stimulant and non-stimulant) is still unclear. Precision medication of psychiatric disorders faces challenges in pharmacogenetics and pharmacodynamics research due to difficulties in recruiting human subjects because of possibility of substance abuse and relatively small sample sizes. Drosophila is a powerful animal model for large-scale studies of drug effects based on the precise quantification of behavior. However, a user-friendly system for high-throughput simultaneous tracking and analysis of drug-treated individual adult flies is still lacking. It is critical to quickly setup a working environment including both the hardware and software at a reasonable cost. Thus, we have developed EasyFlyTracker, an open-source Python package that can track single fruit fly in each arena and analyze Drosophila locomotor and sleep activity based on video recording to facilitate revealing the psychiatric drug effects. The current version does not support multiple fruit fly tracking. Compared with existing software, EasyFlyTracker has the advantages of low cost, easy setup and scaling, rich statistics of movement trajectories, and compatibility with different video recording systems. Also, it accepts multiple video formats such as common MP4 and AVI formats. EasyFlyTracker provides a cross-platform and user-friendly interface combining command line and graphic configurations, which allows users to intuitively understand the process of tracking and downstream analyses and automatically generates multiple files, especially plots. Users can install EasyFlyTracker, go through tutorials, and give feedback on http://easyflytracker.cibr.ac.cn. Moreover, we tested EasyFlyTracker in a study of Drosophila melanogaster on the hyperactivity-like behavior effects of two psychiatric drugs, methylphenidate and atomoxetine, which are two commonly used drugs treating attention-deficit/hyperactivity disorder (ADHD) in human. This software has the potential to accelerate basic research on drug effect studies with fruit flies.

An overview of posttraumatic stress disorder genetic studies by analyzing and integrating genetic data into genetic database PTSDgene.

Posttraumatic stress disorder (PTSD) is a debilitating psychiatric syndrome with complex etiology. Studies aiming to explore genetic susceptibility and environmental triggers of PTSD have been increasing. However, the results are limited and highly heterogeneous. To understand the genetic study status of PTSD and explore more reliable candidates, we obtained 105 PTSD related genetic studies by comprehensively literature searching and filtering 1762 studies. Detailed phenotype and sample information for each study and association results for each genetic marker were extracted. Based on the extracted data, we reviewed the PTSD genetic research status and further conducted bioinformatics analyses for the genetic data. Our analyses summarized the landscape of PTSD genetic studies, identified the genes with most genetic evidence, discovered the biological function of the candidate variants/genes and enlarged the overall candidates for future investigations. All the data were stored in the PTSDgene database (http://ptsdgene.psych.ac.cn). We hope PTSDgene could be a platform for the rapid growth of PTSD genetic data and provide new insights into the pathogenesis of PTSD.

Common variants near IKZF1 are associated with primary Sjögren's syndrome in Han Chinese.

Primary Sjögren's syndrome (pSS) is a systematic autoimmune disease with evidence of genetic predisposition. The IKZF1 (IKAROS family zinc finger 1 (Ikaros)) gene is located at 7p12.2, encodes a transcription factor related to chromatin remodeling, regulates lymphocyte differentiation, and has been reported to be associated with some autoimmune diseases. However, there have been no reports of an association between IKZF1 and pSS. To investigate the possibility of an association between the IKZF1 locus and pSS, we selected two single nucleotide polymorphisms (SNPs) in the IKZF1 locus, rs4917129 and rs4917014, based on a detailed analysis of genome-wide association study (GWAS) data and performed genotyping in 665 Han Chinese pSS patients and 863 healthy controls. The results of an association test showed significant association signals (rs4917129: P-value = 5.5e-4, OR (odds ratio) = 0.72, 95% CI (confidence interval) = 0.60-0.87; rs4917014: P-value = 1.2e-3, OR = 0.76, 95% CI = 0.64-0.89). A meta-analysis that combined the above results with data from previous GWAS, further confirmed these associations (rs4917129: Pmeta = 4.24e-8, ORmeta = 0.70, 95% CI = 0.61-0.79; rs4917014: Pmeta = 6.0e-8, ORmeta = 0.72, 95% CI = 0.64-0.81). A bioinformatics analysis indicated that both SNPs were located in a putative enhancer area in immune-related cell lines and tissues. A protein-protein interaction analysis found that IKZF1, together with GTF2I (an SS susceptibility gene newly identified through GWAS), could interact with histone deacetylase family proteins. In summary, this is the first study to report an association between IKZF1 and SS in Han Chinese.

A combined analysis of genome-wide expression profiling of bipolar disorder in human prefrontal cortex.

Numbers of gene expression profiling studies of bipolar disorder have been published. Besides different array chips and tissues, variety of the data processes in different cohorts aggravated the inconsistency of results of these genome-wide gene expression profiling studies. By searching the gene expression databases, we obtained six data sets for prefrontal cortex (PFC) of bipolar disorder with raw data and combinable platforms. We used standardized pre-processing and quality control procedures to analyze each data set separately and then combined them into a large gene expression matrix with 101 bipolar disorder subjects and 106 controls. A standard linear mixed-effects model was used to calculate the differentially expressed genes (DEGs). Multiple levels of sensitivity analyses and cross validation with genetic data were conducted. Functional and network analyses were carried out on basis of the DEGs. In the result, we identified 198 unique differentially expressed genes in the PFC of bipolar disorder and control. Among them, 115 DEGs were robust to at least three leave-one-out tests or different pre-processing methods; 51 DEGs were validated with genetic association signals. Pathway enrichment analysis showed these DEGs were related with regulation of neurological system, cell death and apoptosis, and several basic binding processes. Protein-protein interaction network further identified one key hub gene. We have contributed the most comprehensive integrated analysis of bipolar disorder expression profiling studies in PFC to date. The DEGs, especially those with multiple validations, may denote a common signature of bipolar disorder and contribute to the pathogenesis of disease.

Convergent evidence from systematic analysis of GWAS revealed genetic basis of esophageal cancer.

Recent genome-wide association studies (GWAS) have identified single nucleotide polymorphisms (SNPs) associated with risk of esophageal cancer (EC). However, investigation of genetic basis from the perspective of systematic biology and integrative genomics remains scarce.In this study, we explored genetic basis of EC based on GWAS data and implemented a series of bioinformatics methods including functional annotation, expression quantitative trait loci (eQTL) analysis, pathway enrichment analysis and pathway grouped network analysis.Two hundred and thirteen risk SNPs were identified, in which 44 SNPs were found to have significantly differential gene expression in esophageal tissues by eQTL analysis. By pathway enrichment analysis, 170 risk genes mapped by risk SNPs were enriched into 38 significant GO terms and 17 significant KEGG pathways, which were significantly grouped into 9 sub-networks by pathway grouped network analysis. The 9 groups of interconnected pathways were mainly involved with muscle cell proliferation, cellular response to interleukin-6, cell adhesion molecules, and ethanol oxidation, which might participate in the development of EC.Our findings provide genetic evidence and new insight for exploring the molecular mechanisms of EC.

Fibroblast growth factor 21 protects mouse brain against D-galactose induced aging via suppression of oxidative stress response and advanced glycation end products formation.

Fibroblast growth factor 21 (FGF21) is a hormone secreted predominantly in the liver, pancreas and adipose tissue. Recently, it has been reported that FGF21-Transgenic mice can extend their lifespan compared with wild type counterparts. Thus, we hypothesize that FGF21 may play some roles in aging of organisms. In this study d-galactose (d-gal)-induced aging mice were used to study the mechanism that FGF21 protects mice from aging. The three-month-old Kunming mice were subcutaneously injected with d-gal (180mg·kg(-1)·d(-1)) for 8weeks and administered simultaneously with FGF21 (1, 2 or 5mg·kg(-1)·d(-1)). Our results showed that administration of FGF21 significantly improved behavioral performance of d-gal-treated mice in water maze task and step-down test, reduced brain cell damage in the hippocampus, and attenuated the d-gal-induced production of MDA, ROS and advanced glycation end products (AGEs). At the same time, FGF21 also markedly renewed the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and total anti-oxidation capability (T-AOC), and decreased the enhanced total cholinesterase (TChE) activity in the brain of d-gal-treated mice. The expression of aldose reductase (AR), sorbitol dehydrogenase (SDH) and member-anchored receptor for AGEs (RAGE) declined significantly after FGF21 treatment. Furthermore, FGF21 suppressed inflamm-aging by inhibiting IκBα degradation and NF-κB p65 nuclear translocation. The expression levels of pro-inflammatory cytokines, such as TNF-α and IL-6, decreased significantly. In conclusion, these results suggest that FGF21 protects the aging mice brain from d-gal-induced injury by attenuating oxidative stress damage and decreasing AGE formation.