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(1) Background: Myelodysplastic neoplasms (MDSs) consist of a group of blood malignancies with a complex biological background. In this context, we investigated the role of autophagy and apoptosis in the pathogenesis and progression of MDSs. (2) Methods: To address this issue, we performed a systematic expression analysis on a total of 84 genes in patients with different types of MDSs (low/high risk of malignancy) versus healthy individuals. Furthermore, real-time quantitative PCR (qRT-PCR) was used to validate significantly upregulated or downregulated genes in a separate cohort of MDS patients and healthy controls. (3) Results: MDS patients were characterized by lower expression levels for a large series of genes involved in both processes compared to healthy individuals. Of importance, deregulation was more pronounced in patients with higher-risk MDS. Results from the qRT-PCR experiments displayed a high level of concordance with the PCR array, strengthening the relevance of our findings. (4) Conclusions: Our results indicate a clear effect of autophagy and apoptosis on MDS development, which becomes more pronounced as the disease progresses. The results from the present study are expected to assist in our understanding of the biological background of MDSs as well as in the identification of novel therapeutic targets.

Context: Active acromegaly is characterized by lipolysis-induced insulin resistance, which suggests adipose tissue (AT) as a primary driver of metabolic aberrations.Objective: To study the gene expression landscape in AT in patients with acromegaly before and after disease control in order to understand the changes and to identify disease-specific biomarkers.Methods: RNA sequencing was performed on paired subcutaneous adipose tissue (SAT) biopsies from six patients with acromegaly at time of diagnosis and after curative surgery. Clustering and pathway analyses were performed in order to identify disease activity-dependent genes. In a larger patient cohort (n = 23), the corresponding proteins were measured in serum by immunoassay. Correlations between growth hormone (GH), insulin-like growth factor I (IGF-I), visceral AT (VAT), SAT, total AT, and serum proteins were analyzed.Results: 743 genes were significantly differentially expressed (P-adjusted < .05) in SAT before and after disease control. The patients clustered according to disease activity. Pathways related to inflammation, cell adhesion and extracellular matrix, GH and insulin signaling, and fatty acid oxidation were differentially expressed.Serum levels of HTRA1, METRNL, S100A8/A9, and PDGFD significantly increased after disease control (P < .05). VAT correlated with HTRA1 (R = 0.73) and S100A8/A9 (R = 0.55) (P < .05 for both).Conclusion: AT in active acromegaly is associated with a gene expression profile of fibrosis and inflammation, which may corroborate the hyper-metabolic state and provide a means for identifying novel biomarkers. Context: Active acromegaly is characterized by lipolysis-induced insulin resistance, which suggests adipose tissue (AT) as a primary driver of metabolic aberrations. Objective: To study the gene expression landscape in AT in patients with acromegaly before and after disease control in order to understand the changes and to identify disease-specific biomarkers. Methods: RNA sequencing was performed on paired subcutaneous adipose tissue (SAT) biopsies from six patients with acromegaly at time of diagnosis and after curative surgery. Clustering and pathway analyses were performed in order to identify disease activity-dependent genes. In a larger patient cohort (n = 23), the corresponding proteins were measured in serum by immunoassay. Correlations between growth hormone (GH), insulin-like growth factor I (IGF-I), visceral AT (VAT), SAT, total AT, and serum proteins were analyzed. Results: 743 genes were significantly differentially expressed (P-adjusted < .05) in SAT before and after disease control. The patients clustered according to disease activity. Pathways related to inflammation, cell adhesion and extracellular matrix, GH and insulin signaling, and fatty acid oxidation were differentially expressed. Serum levels of HTRA1, METRNL, S100A8/A9, and PDGFD significantly increased after disease control (P < .05). VAT correlated with HTRA1 (R = 0.73) and S100A8/A9 (R = 0.55) (P < .05 for both). Conclusion: AT in active acromegaly is associated with a gene expression profile of fibrosis and inflammation, which may corroborate the hyper-metabolic state and provide a means for identifying novel biomarkers.

Craniofacial structures change dynamically in morphology during development through the coordinated regulation of various cellular molecules. However, it remains unclear how these complex mechanisms are regulated in a spatiotemporal manner. Here we applied natural cubic splines to model gene and microRNA (miRNA) expression from embryonic day (E) 10.5 to E14.5 in the proximal and distal regions of the maxillary processes to identify spatiotemporal patterns of gene and miRNA expression, followed by constructing corresponding regulatory networks. Three major groups of differentially expressed genes (DEGs) were identified, including 3,927 temporal, 314 spatial, and 494 spatiotemporal DEGs. Unsupervised clustering further resolved these spatiotemporal DEGs into 8 clusters with distinct expression patterns. Interestingly, we found 2 clusters of differentially expressed miRNAs: 1 had 80 miRNAs monotonically decreasing and the other had 97 increasing across developmental stages. To evaluate the phenotypic relevance of these DEGs during craniofacial development, we integrated data from the CleftGeneDB database and constructed the regulatory networks of genes related to orofacial clefts. Our analysis revealed 2 hub miRNAs, mmu-miR-325-3p and mmu-miR-384-5p, that repressed cleft-related genes Adamts3, Runx2, Fgfr2, Acvr1, and Edn2, while their expression increased over time. On the contrary, 2 hub miRNAs, mmu-miR-218-5p and mmu-miR-338-5p, repressed cleft-related genes Pbx2, Ermp1, Snai1, Tbx2, and Bmi1, while their expression decreased over time. Our experiments indicated that these miRNA mimics significantly inhibited cell proliferation in mouse embryonic palatal mesenchymal (MEPM) cells and O9-1 cells through the regulation of genes associated with cleft palate and validated the role of our regulatory networks in orofacial clefts. To facilitate interactive exploration of these data, we developed a user-friendly web tool to visualize the gene and miRNA expression patterns across developmental stages, as well as the regulatory networks ( https://fyan.shinyapps.io/facebase_shiny/ ). Taken together, our results provide a valuable resource that serves as a reference map for future research in craniofacial development.

Various cell types can be derived from stem cells. However, these cells are immature and do not match their adult counterparts in functional capabilities, limiting their use in disease modeling and cell therapies. Thus, it is crucial to understand the mechanisms of maturation in vivo. However, it is unknown if there are genes and pathways conserved across organs during maturation. To address this, we performed a time-series analysis of the transcriptome of the mouse heart, brain, liver, and kidney and analyzed their trajectories over time. In addition, gene regulatory networks were reconstructed to determine overlapping expression patterns. Based on these, we identified commonly upregulated and downregulated pathways across all four organs. Key upstream regulators were also predicted based on the temporal expression of downstream genes. These findings suggest the presence of universal regulators during organ maturation, which may help us develop a general strategy to mature stem cell-derived cells in vitro.

The cancer genome plays an increasingly large role in the care of patients with breast cancer. Commercially available gene-expression profiling assays are now a part of staging and treatment guidelines, and their use continues to be examined in large-scale studies. With the advent of next-generation sequencing, the cancer genome can now be examined more quickly, less invasively, and in much greater detail. These technologies have led to a more nuanced understanding of molecular pathways, allowing providers to better match patients to clinical trials. Furthermore, a new era of diagnostics based on liquid biopsies is expected to revolutionize disease detection and clinical care.

Lead (Pb) contamination has become an important abiotic stress that negatively influences crop biomass and yield, threatening human health via food chains. The excavation of causal genes for Pb tolerance in maize will contribute to the breeding of Pb-tolerant maize germplasms. This study aimed to demonstrate the effects of AKINbetagamma-1 protein kinase (ZmAKINβγ1) on maize tolerance to Pb and reveal its molecular mechanisms underlying Pb tolerance. ZmAKINβγ1 was identified using genome-wide association study and weighted gene co-expression network analysis for shoot dry weight (SDW) and root dry weight (RDW) under Pb treatment. The OE and RNAi experiments showed that ZmAKINβγ1 negatively regulated maize tolerance to Pb by reducing SDW and RDW and increasing Pb accumulation in maize. Comparative transcriptome analysis between the OE/RNAi and wild-type lines revealed that ZmAKINβγ1 participated in the pectin metabolism process and nitrogen compound response. Gene-based association analyses revealed that three variants located in ZmAKINβγ1 promoter induced changes in its expression and Pb tolerance among maize lines. The dual-luciferase reporter system verified that the two genotypes (AAT and CGG) of ZmAKINβγ1 promoter had contrasting transcriptional activities. Collectively, ZmAKINβγ1-mediated Pb tolerance provided new insights into the cultivation of Pb-tolerant maize varieties and phytoremediation of Pb-polluted soils.

The oriental fruit fly, Bactrocera dorsalis (Hendel), has very strong ecological adaptability and phenotypic plasticity. Here, the genome of B. dorsalis was assembled into 549.45 Mb sequences with a contig N50 length of 12.81 Mb. Among, 95.67 % assembled genome sequences were anchored on six chromosomes with an N50 length of 94.63 Mb. According to the basic characteristics of the sex chromosomes of Tephritidae, the X chromosome of B. dorsalis was identified. Significant gene expansions were detected in several important gene families related to adaptability. In particular, we annotated 50 histone modification enzymes (HMEs) in this genome. A comparative transcriptome analysis indicated that 12 HME genes were differentially expressed in two thermo-tolerant strains (heat and cold). Interestingly, four and seven of the 12 HME genes responded to heat shock or cold hardening, respectively. These evidences suggested that the histone modification as an epigenetic modification may be involved in the thermal tolerance of B. dorsalis, but with different regulation mechanisms in thermal acclimation and hardening. The high quality genome of B. dorsalis provides an invaluable resource for further functional genomic study. Moreover, comparative genomic analysis will shed insights on revealing the mechanisms of adaptive evolution in this fly.

Silver pomfret has been widely cultured in China due to its high economic value. Photobacterium damselae subsp. damselae (PDD) is a Gram-negative bacterium that has been shown to infect many fish species. To increase knowledge of the molecular mechanisms of the host defense against PDD, we conducted transcriptome analysis of head kidney in silver pomfret at 24 h and 72 h post-infection (hpi) via Illumina sequencing. The de novo assembly resulted in the identification of 79,063 unigenes, with 59,386 (75.11%) successfully annotated in public databases (NR, NT, KO, Swiss-Prot, Pfam, GO, and KOG databases). Comparison of gene expression profiles between PBS-injected fish (sham control) and PDD-challenged fish revealed 329 and 570 differentially expressed genes (DEGs) were screened at 24 hpi and 72 hpi, respectively. The DEGs were enriched in multiple immune-related pathways such as Hepatitis C, Gastric acid secretion, CAMs and Leukocyte transendothelial migration pathways, Primary immunodeficieny, ECM-receptor interaction, PI3K-Akt signaling pathway. The data obtained in the present study offers valuable information for acute immune response of silver pomfret challenged with PDD, which will facilitate further investigations on strategies against Photobacterium spp. infection in teleosts.

Molting is a basic physiological behavior of the Oriental river prawn (Macrobrachium nipponense), however, the gene expression patterns and immune mechanisms during the molting process of Oriental river prawn are unclear. In the current study, the gene expression levels of the hepatopancreas of the Oriental river prawn at different molting stages (pre-molting, Prm; mid-molting, Mm; and post-molting, Pom) were detected by mRNA sequencing. A total of 1721, 551, and 1054 differentially expressed genes (DEGs) were identified between the Prm hepatopancreas (PrmHe) and Mm hepatopancreas (MmHe), MmHe and Pom hepatopancreas (PomHe) and PrmHe and PomHe, respectively. The results showed that a total of 1151 DEGs were annotated into 316 signaling pathways, and the significantly enriched immune-related pathways were "Lysosome", "Hippo signaling pathway", "Apoptosis", "Autophagy-animal", and "Endocytosis". The qRT-PCR verification results of 30 randomly selected DEGs were consistent with RNA-seq. The expression patterns of eight immune related genes in different molting stages of the Oriental river prawn were analyzed by qRT-PCR. The function of Caspase-1 (CASP1) was further investigated by bioinformatics, qRT-PCR, and RNAi analysis. CASP1 has two identical conserved domains: histidine active site and pentapeptide motif, and the expression of CASP1 is the highest in ovary. The expression levels of triosephosphate isomerase (TPI), Cathepsin B (CTSB) and Hexokinase (HXK) were evaluated after knockdown of CASP1. This research provides a valuable basis to improve our understanding the immune mechanisms of Oriental river prawns at different molting stages. The identification of immune-related genes is of great significance for enhancing the immunity of the Oriental river prawn, or other crustaceans, by transgenic methods in the future.

Banded leaf and sheath blight (BLSB) is a devasting disease caused by the necrotrophic fungus Rhizoctonia solani that affects maize (Zea mays L.) fields worldwide, especially in China and Southeast Asia. Understanding how maize plants respond to R. solani infection is a key step towards controlling the spread of this fungal pathogen. In this study, we determined the transcriptome of maize plants infected by a low-virulence strain (LVS) and a high-virulence strain (HVS) of R. solani for 3 and 5 days by transcriptome deep-sequencing (RNA-seq). We identified 3,015 (for LVS infection) and 1,628 (for HVS infection) differentially expressed genes (DEGs). We confirmed the expression profiles of 10 randomly selected DEGs by quantitative reverse transcription PCR. We also performed a Gene Ontology (GO) enrichment analysis to establish which biological processes are associated with these DEGs, which revealed the enrichment of defense-related GO terms in LVS- and HVS-regulated genes. We selected 388 DEGs upregulated upon fungal infection as possible candidate genes. Among them, the overexpression of ZmNAC41 (encoding NAC transcription factor 41) or ZmBAK1 (encoding BRASSINOSTEROID INSENSITIVE 1-associated receptor kinase 1) in rice enhanced resistance to R. solani. In addition, overexpressing ZmBAK1 in rice also increased plant height, plant weight, thousand-grain weight, and grain length. The identification of 388 potential key maize genes related to resistance to R. solani provides significant insights into improving BLSB resistance.

Whole genome and transcriptome analysis of 213 Pseudomonas aeruginosa isolates resistant to antipseudomonal β-lactams collected in 30 countries was performed to evaluate resistance mechanisms against these agents.Isolates were susceptibility tested by reference broth microdilution. Whole genome and transcriptome sequencing were performed, and data were analysed using open-source tools. A statistical analysis of changes in the expression of5500 genes was compared to the expression of PAO1.The high-risk clones ST235 and ST111 were the most prevalent among90 sequence types (STs). Metallo-β-lactamase (MBLs) genes were detected in 40 isolates. AmpC and MexXY were the most common genes overexpressed in approximately 50% of the 173 isolates that did not carry MBLs. Isolates overexpressing pmrA and pmrB, the norspermidine production genes speD2 and speE2, and the operon arnBCADTEF-ugd were noted among strains resistant to ceftolozane-tazobactam and ceftazidime-avibactam, despite the lack of polymyxin resistance often associated to increased expression of these genes. Overexpression of MuxABC-OpmB, OprG, and OprE proteins were associated with resistance to ceftolozane-tazobactam in addition to the usual genes involved in cephalosporin, monobactam, and carbapenem resistance. Statistical analysis identified discrete mutations in ArmZ, OprD, and AmpC that correlated to antipseudomonal β-lactam resistance.P. aeruginosa resistance mechanisms are complex. This analysis suggests the role of multiple genes in resistance to antipseudomonal β-lactams, including some not commonly described.

One of the most widespread coral diseases linked to anthropogenic activities and recorded on reefs worldwide is characterized by anomalous growth formations in stony corals, referred to as coral growth anomalies (GAs). The biological functions of GA tissue include limited reproduction, reduced access to resources, and weakened ability to defend against predators. Transcriptomic analyses have revealed that, in some cases, disease progression can involve host genes related to oncogenesis, suggesting that the GA tissues may be malignant neoplasms such as those developed by vertebrates. The number of studies reporting the presence of GAs in common reef-forming species highlights the urgency of a thorough understanding of the pathology and causative factors of this disease and its parallels to higher organism malignant tissue growth. Here, we review the current state of knowledge on the etiology and holobiont features of GAs in reef-building corals.

Genomic profiling using short-read sequencing has utility in detecting disease-associated variation in both DNA and RNA. However, given the frequent occurrence of structural variation in cancer, molecular profiling using long-read sequencing improves the resolution of such events. For example, the Pacific Biosciences long-read RNA-sequencing (Iso-Seq) transcriptome protocol provides full-length isoform characterization, discernment of allelic phasing, and isoform discovery, and identifies expressed fusion partners. The Pacific Biosciences Fusion and Long Isoform Pipeline (PB_FLIP) incorporates a suite of RNA-sequencing software analysis tools and scripts to identify expressed fusion partners and isoforms. In addition, sequencing of a commercial reference (Spike-In RNA Variants) with known isoform complexity was performed and demonstrated high recall of the Iso-Seq and PB_FLIP workflow to benchmark our protocol and analysis performance. This study describes the utility of Iso-Seq and PB_FLIP analysis in improving deconvolution of complex structural variants and isoform detection within an institutional pediatric and adolescent/young adult translational cancer research cohort. The exemplar case studies demonstrate that Iso-Seq and PB_FLIP discover novel expressed fusion partners, resolve complex intragenic alterations, and discriminate between allele-specific expression profiles.

Oysters are commercially important intertidal filter-feeding species. Mass mortality events of oysters often occur due to environmental stresses, such as exposure to fluctuating temperatures, salinity, and air, as well as to metal pollution and pathogen infection. Here, RNA-seq data were used to identify shared and specific responsive genes by differential gene expression analysis and weighted gene co-expression network analysis. A total of 18 up-regulated and 10 down-regulated shared responsive genes were identified corresponding to five different stressors. Total 27 stressor-specific genes for temperature, 11 for salinity, 80 for air exposure, 51 for metal pollution, and 636 for Vibrio mediterranei pathogen stress were identified in oysters. Elongin-β was identified as a crucial gene for thermal stress response. Some HSP70s were determined to be shared responsive genes while others were specific to thermal tolerance. The proteins encoded by these stress-related genes should be further investigated to characterize their physiological functions. In addition, the uncharacterized proteins and ncRNAs that were identified may be involved in species-specific stress-response and regulatory mechanisms. This study identified specific genes related to stressors relevant to oyster cultivation. These findings provide useful information for new selective breeding strategies using a data driven method.

Neuropathic pain can cause significant physical and economic burden, and there are no effective long-term treatments. We conducted a bioinformatics analysis to identify mechanisms to determine strategies for more effective treatments of neuropathic pain.GSE24982 and GSE63442 microarray datasets were extracted from the Gene Expression Omnibus database to analyze transcriptome differences of neuropathic pain in the dorsal root ganglions (DRGs). We filtered the differentially expressed genes (DEGs) in the two datasets and conducted Gene Ontology (GO) functional analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of the shared DEGs. The Protein-Protein Interaction network was used to determine the hub genes, which were verified in the GSE30691 dataset. miRDB and miRWalk Databases were used to predict potential miRNA of the selected DEGs. We made the spinal nerve ligation (SNL) rat model and qPCR was used to verify the differential expression of hub genes.A total of 182 overlapped DEGs were found between GSE24982 and GSE63442 datasets. The GO and KEGG analysis showed that the selected DEGs were enriched in infection, transmembrane transport of ion channels, and synaptic transmission. We identified seven hub genes (Atf3, Aif1, Ctss, Gfap, Scg2, Jun, and Vgf). qPCR verified the expression differences of the hub genes in the DRGs after SNL model. Predicted miRNA targeting each selected hub genes were identified.Seven hub genes related to the pathogenesis of neuropathic pain and potential targeting miRNA were identified, expanding understanding of the mechanism of neuropathic pain and facilitating treatment development.

The apicomplexan cattle parasite Theileria parva is a major barrier to improving the livelihoods of smallholder farmers in Africa, killing over one million cattle on the continent each year. Although exotic breeds not native to Africa are highly susceptible to the disease, previous studies have illustrated that such breeds often show innate tolerance to infection by the parasite. The mechanisms underlying this tolerance remain largely unclear. To better understand the host response to T. parva infection we characterised the transcriptional response over 15 days in tolerant and susceptible cattle (n = 29) naturally exposed to the parasite. We identify key genes and pathways activated in response to infection as well as, importantly, several genes differentially expressed between the animals that ultimately survived or succumbed to infection. These include genes linked to key cell proliferation and infection pathways. Furthermore, we identify response expression quantitative trait loci containing genetic variants whose impact on the expression level of nearby genes changes in response to the infection. These therefore provide an indication of the genetic basis of differential host responses. Together these results provide a comprehensive analysis of the host transcriptional response to this under-studied pathogen, providing clues as to the mechanisms underlying natural tolerance to the disease.

Background: Formalin-fixed, paraffin-embedded (FFPE) tissues are a valuable resource for clinical and basic science research. Paraffin blocks and the resulting unstained sections (USS) are often st...

Gastrodia elata is an achlorophyllous and fully mycoheterotrophic orchid which obtains carbon and other nutrients from Armillaria species in its life cycle. Many researchers suggested that plant hormones, as signing molecules, play a central role in the plant-fungi interaction. In the process of Armillaria gallica 012 m cultivation, both exogenous indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) distinctly stimulated the growth of mycelia in solid media. The differential expression genes (DEGs) of A. gallica 012 m with IAA versus blank control (BK) and IBA versus BK were investigated. The results showed that more than 80% of DEGs of the IAA group were coincident with the DEGs of the IBA group, and more than half of upregulated DEGs and most of the downregulated DEGs of the IAA group coincided with those DEGs of the IBA group. Above research implied that A. gallica 012 m could perceive IAA and IBA, and possess similar responses and signaling pathways to IAA and IBA. The overlapping differential genes of the IAA group and IBA group were analyzed by GO term, and the results showed that several DEGs identified were related to biological processes including positive regulation of the biological process and biological process. The downregulated NmrA-like and FKBP_C genes might be benefit to the growth of mycelia. Those results can explain that exiguous IAA and IBA improved the growth of A. gallica to some extent. We speculate that IAA and IBA are signaling molecules, and regulate the expression of growth-related genes of A. gallica 012 m by the same signaling pathway.

Background. The prognostic value of tumor-infiltrating immune cells has been widely studied in nasopharyngeal carcinoma (NPC). However, the role of tumor-infiltrating immune cells in the diagnosis of NPC has not been fully elucidated. Thus, tumor-infiltrating immune cell-related biomarkers in the diagnosis of NPC patients were explored in the current study. Methods. Gene expression profiles of NPC patients were downloaded from the Gene Expression Omnibus (GEO) database. Differentially infiltrating immune cells (DDICs) between NPC and control samples were analyzed by the CIBERSORT algorithm. Weighted gene coexpression network analysis (WGCNA) was performed to screen hub genes significantly correlated with DDIC. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of hub genes were performed with R package clusterProfiler. The diagnostic value of hub genes was evaluated by receiver operating characteristic (ROC) curves. RT-qPCR was conducted to validate the expression patterns of diagnostic markers in NPC and adjacent control tissues. The correlations between diagnostic markers and immunomodulators were analyzed using the TISIDB. The protein-protein interaction (PPI) network based on immunomodulators significantly associated with diagnostic biomarkers was constructed and visualized by STRING. The functional enrichment analysis of genes in the PPI network was analyzed by the WebGestalt online tool. Results. The abundances of memory B cells, plasma cells, follicular helper T cells, activated NK cells, M0 macrophages, M1 macrophages, M2 macrophages, resting mast cells, and activated mast cells were significantly different between NPC and control samples. Dark orange was identified as the hub module, with a total of 371 genes associated with memory B cells, plasma cells, and M0 and M1 macrophages defined as hub genes, which were enriched into immune-related biological processes and pathways. FCER2, KHDRBS2, and IGSF9 were considered diagnostic biomarkers with areas under ROC curves as 0.985, 0.978, and 0.975, respectively. Moreover, real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) suggested that the expression patterns of FCER2, KHDRBS2, and IGSF9 were consistent with the results in GEO datasets. TISIDB analysis revealed that FCER2, KHDRBS2, and IGSF9 had a strong association with 8 immunoinhibitors (BTLA, CD160, CD96, LAG3, PDCD1, TIGIT, CD244, and TGFB1) and 11 immunostimulators (CD27, CD28, CD40LG, CD48, ICOS, KLRC1, KLRK1, TMIGD2, TNFRSF13C, CXCR4, and C10 or f54). The PPI network implied that these 19 immunomodulators had interactions with other 50 genes. WebGestalt analysis demonstrated that 69 genes in the PPI network were enriched into cytokine-cytokine receptor interaction, NF-kappa B signaling pathway, and pathways in cancer. Conclusion. Our study identified novel diagnostic biomarkers and revealed potential immune-related mechanisms in NPC. These findings enlighten the investigation of the molecular mechanisms of tumor-infiltrating immune cells regulating NPC.

Object. Osteosarcoma is an intractable malignant disease, and few therapeutic methods can thoroughly eradicate its focuses. This study attempted to investigate the related mechanism of osteosarcoma by bioinformatics methods. Methods. GSE70367 and GSE69470 were obtained from the GEO database. The differentially expressed genes (DEGs) and miRNAs were analyzed using the GEO2R tool and then visualized with R software. Moreover, the targets of the miRNAs in the DEGs were screened and then used for enrichment analysis. Besides, the STRING database and Cytoscape were applied to illustrate the protein-protein interaction network. RT-qPCR was performed to measure the expression of key genes and miRNAs. Western blot was applied to detect the signaling pathway. Results. 9 upregulated genes and 39 downregulated genes in GSE69470 were identified as the DEGs, and 31 upregulated genes and 56 downregulated genes in GSE70367 were identified as the DEGs. Moreover, 21 common genes were found in the DEGs of GSE70367 and GSE69470. The enrichment analysis showed that the common DEGs of GSE70367 and GSE69470 were related with cell development, covalent chromatin modification, and histone modification and involve in the regulation of MAPK, mTOR, and AMPK pathways. Besides, the miRNAs including miR-543, miR-495-3p, miR-433-3p, miR-381-3p, miR-301a-3p, miR-199b-5p, and miR-125b-5p were identified as the biomarkers of osteosarcoma. In addition, the target genes including HSPA5, PPARG, MAPK14, RAB11A, RAB5A, MAPK8, LEF1, HIF1A, CAV1, GS3KB, FOXO3, IGF1, and NFKBIA were identified as hub nodes. It was found that miR-301a-3p expression was decreased and mRNA expression of RAB5A and NFKBIA was increased in the pathological tissues. The AKT-PI3K-mTOR signaling pathway was activated in pathological tissues. Conclusion. In this study, 7 miRNAs and 13 hub genes were identified, which might be candidate markers. miR-301a-3p, RAB5A, and NFKBIA were abnormally expressed in osteosarcoma tissues.

As the most prevalent primary CNS tumor, glioma is characterized by high mortality and morbidity. This research aims to investigate glioma-associated microRNAs (miRNAs) and their target mRNAs, as well as to explore their biological functions in gliomas. The Gene Expression Omnibus (GEO) database was applied to acquire the GSE112264 miRNA microarray dataset and the GSE15824 mRNA dataset. We selected samples from the GSE112264 dataset and the GSE15824 to identify differently expressed miRNAs (DE-miRNAs) as well as differentially expressed mRNAs (DEGs), respectively. Next, the intersections of mRNA and target mRNAs of miRNA were selected, and we constructed miRNA-mRNA regulation networks. These DEGs were selected for Gene Oncology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses by conducting the package clusterProfiler. After conducting Cytoscape software, a protein-protein interaction (PPI) network was created. Next, survival analysis of the miR-423-3p was confirmed by conducting TCGA database. Subsequently, Quantitative real-time PCR (qRT-PCR) was conducted to verify miR-423-3p's expression. Finally, miR-423-3p's biological functions of in effecting the cell proliferative, migratory, and invasive capabilities of glioma were investigated by performing Cell Counting Kit-8 (CCK-8) and Transwell assays. Our analysis elucidated a novel miRNA-mRNA regulatory network related to glioma carcinogenesis, which may be considered as future therapeutic biomarkers for glioma.

Sepia esculenta is a common economic cephalopod that has received extensive attention due to the tender meat, rich protein content and certain medicinal value thereof. Over the past decade, multiple industries have discharged waste into the ocean in large quantities, thereby significantly increasing the concentration of heavy metals in the ocean. Copper (Cu) is a common heavy metal in the ocean. The increase of Cu content will affect numerous biological processes such as immunity and metabolism of marine organisms. High concentrations of Cu may inhibit S. esculenta growth, development, swimming, and other processes, which would significantly affect its culture. In this research, transcriptome analysis is used to initially explore Cu-exposed S. esculenta larval immune response mechanisms. And compared to control group with normally growing larvae, 2056 differentially expressed genes (DEGs) are identified in experimental group with Cu-exposed larvae. The results of DEGs functional enrichment analyses including GO and KEGG indicate that Cu exposure might promote inflammatory and innate immune responses in cuttlefish larvae. Then, 10 key genes that might regulate larval immunity are identified using a comprehensive analysis that combines protein-protein interaction (PPI) network and KEGG functional enrichment analyses, of which three genes with the highest number of protein interactions or involve in more KEGG signaling pathways are identified as hub genes that might significantly affect larval immune response processes. Comprehensive analysis of PPI network and KEGG signaling pathway are used for the first time to explore Cu-exposed S. esculenta larval immune response mechanisms. Our results preliminarily reveal immune response mechanisms of cephalopods exposed to heavy metals and provide valuable resources for further understanding mollusk immunity.

Aims Phenazines, such as phenazine-1-carboxylic acid (PCA), phenazine-1-carboxamide (PCN), 2-hydroxyphenazine-1-carboxylic acid (2-OH-PCA), 2-hydroxyphenazine (2-OH-PHZ), are a class of secondary metabolites secreted by plant-beneficial Pseudomonas. Ps. chlororaphis GP72 utilizes glycerol to synthesize PCA, 2-OH-PCA and 2-OH-PHZ, exhibiting broad-spectrum antifungal activity. Previous studies showed that the addition of dithiothreitol (DTT) could increase the phenazines production in Ps. chlororaphis GP72AN. However, the mechanism of high yield of phenazine by adding DTT is still unclear. Methods and Results In this study, untargeted and targeted metabolomic analysis were adopted to determine the content of metabolites. The results showed that the addition of DTT to GP72AN affected the content of metabolites of central carbon metabolism, shikimate pathway and phenazine competitive pathway. Transcriptome analysis was conducted to investigate the changed cellular process, and the result indicated that the addition of DTT affected the expression of genes involved in phenazine biosynthetic cluster and genes involved in phenazine competitive pathway, driving more carbon flux into phenazine biosynthetic pathway. Furthermore, genes involved in antioxidative stress, phosphate transport system and mexGHI-opmD efflux pump were also affected by adding DTT. Conclusion This study demonstrated that the addition of DTT altered the expression of genes related to phenazine biosynthesis, resulting in the change of metabolites involved in central carbon metabolism, shikimate pathway and phenazine competitive pathway. Significance and Impact of the Study This work expands the understanding of high yield of phenazine by the addition of DTT and provides several targets for increasing phenazine production.

Aim To reveal the antibacterial mechanism of protocatechuic acid (PCA) against Micrococcus luteus. Methods and Results M. luteus was exposed to PCA, and the antibacterial mechanism was revealed by measuring membrane potential, intracellular ATP and pH levels and transcriptome analysis. PCA induced the membrane potential depolarization of M. luteus, significantly decreased the intracellular ATP and pH levels of M. luteus and disrupted the integrity of the M. luteus cell membrane. Transcriptome analysis showed that PCA induced 782 differentially expressed genes (DEGs) of M. luteus. GO enrichment analysis revealed that the majority of DEGs are involved in pathways of metabolic process, cellular process, biological regulation and transport activity. In addition, PCA inhibited the growth of M. luteus in skimmed milk and extended the shelf life of skimmed milk. Conclusion PCA had good bactericidal activity against M. luteus through the mechanism of cell membrane disruption and metabolic process disorder. Significance and Impact of the Study PCA inhibits the growth of M. luteus in skimmed milk, suggesting that PCA is promising to be used as a novel preservative in food storage.

Teleosts have a unique immune system because their head kidney (HK) and trunk kidney (TK) are sites for hematopoiesis. However, the immune functions of the HK and TKs require further elucidation in yellow catfish (Pelteobagrus fulvidraco). In the present study, imprints of the HK and TK were examined using the Wright's-Giemsa staining method. Morphological characteristics of the blood cell lineages revealed that the HK and TK were hematopoietic organs. To explore its immune function, transcriptome sequencing was performed after infection with Aeromonas hydrophila. A total of 1139 genes showed significant alterations in their expression in the kidney; these genes included 737 upregulated and 402 downregulated genes. Furthermore, 1117 differentially expressed genes were observed in the HK, which included 784 upregulated and 333 downregulated genes. Both organs showed 357 upregulated genes and 85 downregulated genes. Some immune-related genes were only expressed in the TK, such as ATP-dependent RNA helicase DDX58, the gene encoding the immunoglobulin heavy chain and light chain. The immune responses in the HK and TK were differential and the TK played a critical role in the mechanism underlying the immune response. The purpose of the present study was to facilitate the elucidation of the immune defense mechanism of yellow catfish and other teleosts.

Hepatocellular carcinoma is one of the most frequent cancers with high mortality rate worldwide.TCGA LIHC HTseq counts were analyzed. GSEA was performed with GO BP gene sets. GO analysis was performed with differentially expressed genes. The subset of genes contributing most of the enrichment result of GO_BP_CHROMOSOME_SEGREGATION of GSEA were identified. Five genes have been selected in this subset of genes for further analysis. A microarray data set, GSE112790, was analyzed as a validation data set. Survival analysis was performed.According to GSEA and GO analysis several gene sets and processes related to chromosome segregation were enriched in LIHC. GO_BP_CHROMOSOME_SEGREGATION gene set from GSEA had the highest size of the genes contributing most of the enrichment. Five genes in this gene set; BRIP1, NSMCE2, ANAPC7, RAD18 and TTL, whose expressions and prognostic values have not been studied in hepatocellular carcinoma in detail, have been selected for further analyses. Expression of these five genes were identified as significantly upregulated in LIHC RNA-seq and HCC microarray data set. Survival analysis showed that high expression of the five genes was associated with poor overall survival in HCC patients.Selected genes were upregulated and had prognostic value in HCC.

The transcriptome of a cell constitutes an essential piece of cellular identity and contributes to the multifaceted complexity and heterogeneity of cell-types within the mammalian brain. Thus, while a wealth of studies have investigated transcriptomic alterations underlying the pathophysiology of diseases of the brain, their use of bulk-tissue homogenates makes it difficult to tease apart whether observed differences are explained by disease state or cellular composition. Cell-type-specific enrichment strategies are, therefore, crucial in the context of gene expression profiling. Laser capture microdissection (LCM) is one such strategy that allows for the capture of specific cell-types, or regions of interest, under microscopic visualization. In this review, we focus on using LCM for cell-type specific gene expression profiling in post-mortem human brain samples. We begin with a discussion of various LCM systems, followed by a walk-through of each step in the LCM to gene expression profiling workflow and a description of some of the limitations associated with LCM. Throughout the review, we highlight important considerations when using LCM with post-mortem human brain samples. Whenever applicable, commercially available kits that have proven successful in the context of LCM with post-mortem human brain samples are described.

Comparative transcriptome approaches assume that highly or dynamically expressed genes are important. This has led to the identification of many genes critical for cellular activity and organism development. However, while testes express the highest levels of long noncoding RNAs (lncRNAs), there is scarcely any evidence for lncRNAs with significant roles in fertility. This was explained by changes in chromatin structure during spermatogenesis that lead to 'promiscuous transcription' with no functional roles for the transcripts. Recent discoveries offer novel and surprising alternatives. Here, I review the current knowledge regarding the involvement of lncRNAs in fertility, why I find gametogenesis different from other developmental processes, offer models to explain why the experimental evidence did not meet theoretical predictions, and suggest possible approaches to test the models.

Vibrio is an important conditional pathogen in shrimp aquaculture. This research reported a dominant bacteria strain E1 isolated from a shrimp tank with the method of biofloc culture, which was further identified as Vibrio owensii. To understand the interaction between V. owensii and the host shrimp, we studied the pathogenicity of the V. owensii and the molecular mechanisms of the Fenneropenaeus merguiensis immunity during the Vibrio invasion. Drug susceptibility tests showed that V. owensii was resistant to antibiotics streptomycin oxacillin, tetracycline, minocycline, and aztreonam, but highly sensitive to cefazolin, cefotaxime, and ciprofloxacin, and moderately sensitive to cefotaxime, ampicillin, and piperacillin. Lethal concentration 50 (LC

Long non-coding RNA (lncRNA) play vital regulatory roles in various biological processes. Intestine is one of the most sensitive organs to environmental and homeostatic disruptions for fish. However, systematic profiles of lncRNAs in the intestine of teleost in responses to pathogen infections is still limited. Turbot (Scophthalmus maximus L.), an important commercial fish species in China, has been suffering with Vibrio anguillarum infection, resulted in dramatic economic loss. Hereinto, the intestinal tissues of turbot were sampled at 0 h, 2 h, 12 h, and 48 h following V. anguillarum infection. The histopathological analysis revealed that the pathological trauma was mainly present in intestinal tunica mucosal epithelium. After high-throughput sequencing and bioinformatic analysis, a total of 9722 lncRNAs and 21,194 mRNAs were obtained, and the average length and exon number of lncRNAs were both less than those of mRNAs. Among which, a set of 158 lncRNAs and 226 mRNAs were differentially expressed (DE-lncRNAs and DEGs) in turbot intestine at three time points, related to many immune-related genes such as complement, interleukin, chemokine, lysosome, and macrophage, indicating their potential critical roles in immune responses. In addition, 2803 and 1803 GO terms were enriched for DEGs and co-expressed target genes of DE-lncRNAs, respectively. Moreover, 127 and 50 KEGG pathways including cell adhesion molecules (CAMs), phagosome, JAK-STAT signaling pathway, cytokine-cytokine receptor interaction, and intestinal immune network for IgA production, were enriched for DEGs and co-expressed target genes of DE-lncRNAs, respectively. Finally, qRT-PCR was conducted to confirm the reliability of sequencing data. The present study will set the foundation for the future exploration of lncRNA functions in teleost in response to bacterial infection.

Membrane-associated RING-CH (MARCH) family, as Ring-type E3 ligases, have attracted extensive attention to their immune functions. MARCH4 plays an essential role in regulating immune response in mammal. In the present study, it is the first to report on MARCH4 characteristics and signal pathway in fish. MARCH4 in large yellow croaker Larimichthys crocea (named as LcMARCH4) encodes a RING-CH domain and two TM domains, as well as other function domains, including an N-terminal proline rich domain, an AxxxG-motif in TM1, a tyrosine-based YXXØ motif, and a C-terminal PDZ-binding domain. LcMARCH4 is a tissue-specific protein with highly significant expression in brain. The mRNA transcripts of LcMARCH4 were significantly induced in the main organs (skin, gill, spleen, and head-kidney) by C. irritans infection. Consistently, significant increase was observed in spleen and head-kidney after LPS, Poly I:C stimulation and V. parahaemolyticus infection. Subcellular localization analysis showed that LcMARCH4 was localized in the cytoplasm and membrane. Moreover, we found 46 DEGs in a comparative transcriptome analysis between the LcMARCH4 overexpression group and control vector group. The analysis showed that HSPA6, HSPA1B and DNAJB1 might play important regulatory roles to MARCH4 in fish. Notably, two noncoding RNA, both RN7SL1 and RN7SL2, the expression levels went up in MARCH4 overexpression cells. Taken together, this study will provide new insights into finfish MARCH4 and its potential regulatory signaling pathway as well.

Data integration is a critical step in the analysis of multiple single-cell RNA sequencing samples to account for heterogeneity due to both biological and technical variability. scINSIGHT is a new integration method for single-cell gene expression data, and can effectively use the information of biological condition to improve the integration of multiple single-cell samples. scINSIGHT is based on a novel non-negative matrix factorization model that learns common and condition-specific gene modules in samples from different biological or experimental conditions. Using these gene modules, scINSIGHT can further identify cellular identities and active biological processes in different cell types or conditions. Here we introduce the installation and main functionality of the scINSIGHT R package, including how to preprocess the data, apply the scINSIGHT algorithm, and analyze the output.

UNIFAN is an unsupervised cell type annotation tool for single-cell RNA sequencing data (scRNA-seq). Given single-cell expression data as input, UNIFAN outputs cell clusters as well as annotations for each cluster. The clustering process utilizes information on pathways and biological processes and these are also used to annotate the resulting clusters. In this software article, we focus on how to install UNIFAN and on the main steps involved in using UNIFAN for cell type annotations.

Background: The fetus grows in a sterile womb environment. After birth, the newborn immune system has two immediate hurdles to clear. First immediate suppression of the womb compatible immune system and turn on the immune system of the newborn that can counter the antigenic world. The underlying mechanism of immune fluctuation by milk microRNAs (miRNAs) can be crucial for the treatment of critical or premature newborn. Methods: We collected fourteen samples of each colostrum and mature milk from lactating mothers, four samples of each were used for microarray analysis, and the other ten were used for miRNA expression profiling by real-time PCR. Results: From the microarray, 154 differentially expressed miRNAs were identified, whereas 49 miRNAs were revealed as immune-related miRNAs based on a literature study. Among the 49 miRNAs, 33 were already shown as strongly validated immune-related miRNAs (validated by qPCR, Western Blot, and Luciferase assay) and were considered for further analysis. Twenty-two miRNA expressions were analysed by real-time PCR as their Ct values were within considerable limits. Twelve numbers of miRNAs were significantly downregulated in mature milk compared to colostrum, which were again subjected to bioinformatics analysis to predict the biological mechanisms behind the differentially expressed miRNAs. Conclusion: This study shed light on the human milk exosome miRNA expression dynamics during lactation and their possible role in the gradual skewing of the newborns' immune system. The information is crucial for the development and onset of sepsis in premature newborns in the NICU.

Vibrio parahaemolyticus causes serious economic losses to the shrimp farming industry. There is still a lack of adequate understanding of the changes in the overall response of N. denticulata sinensis caused by V. parahaemolyticus, particularly with respect to gill tissue, which is severely damaged by the pathogen. In this study, a total of 1358 differentially expressed genes were identified between the PBS control and Vibrio stimulation groups using transcriptome sequencing techniques. After further screening and analysis, many immune-related genes were obtained, involving lysosome pathway, metabolic process, chitin-binding protein, and serine protease family members. In addition, we randomly selected six DEGs in the lysosome pathway for qRT-PCR verification, and the results showed that their expression patterns were consistent with the RNA-seq. The results demonstrate the molecular regulation of the gill tissue response to V. parahaemolyticus infection in N. denticulata sinensis, contributing to the understand of the complex and efficient innate immune system and defense mechanisms in crustaceans.

By extracting molecular interactions identified by experiments, gene regulatory networks or gene circuits have documented in a large number of knowledge-based repositories. They provide systematic information and guidance of the functional connections between regulators, e.g., transcription factor proteins and miRNAs, and target genes. Network activity is defined as the degree of consistency between a regulatory network architecture and a specific cellular context of gene expression and can also be measured as a score of statistical significance. The gene network activities are closely related to the dynamics of cell states. To evaluate the activity of regulatory events in the form of network, we propose a network activity evaluation (NAE) framework by measuring the consistency between network architecture and gene expression data across specific states based on mathematical programming. NAE firstly employs the dynamic Bayesian network model to formulate the network structure with time series profiling data. For the constraints of prior knowledge about gene regulatory network, NAE introduces an interpretable general loss function with regularization penalties to calculate the degree of consistency between gene network and gene expression data. Moreover, we design a fast and convergent alternating direction method of multipliers algorithm to optimize the regularized constraint programming. The efficiency and advantage of the NAE framework is deduced through numerous experiments and comparison studies. It reflects the possibility and potential of the match between network and data, thereby helping to reveal the network activity and to explain the dynamic responds underlying the network structure caused by changes in molecular environment of living cells. The code of NAE is freely available for academic use (https://github.com/zpliulab/NAE).

A 31-gene expression profile (31-GEP) test that predicts metastatic risk in patients with cutaneous malignant melanoma (CMM) has previously been validated and is available for clinical use. The test dichotomizes patients into lower risk and higher risk groups based on differences that correspond to unique genetic expression patterns. Although the impact of such a test on dermatology providers' clinical decision-making has been studied, little is known about whether there exists an association between certain clinical features, such as dermoscopy, and 31-GEP results.In this retrospective analysis of 31-GEP test results ordered by dermatologists, we evaluated the frequency of dermoscopic features, using a modified dermoscopy three-point checklist, in 17 cases (n=17) and compared these findings to other key clinicopathologic features including tumor thickness, ulceration, and mitotic rate to 31-GEP results. Additionally, we evaluated the dermatologist's perspective and incorporation of GEP testing as part of patient discussion on melanoma management.31-GEP stratified patients into 4 groups; groups 1A and 1B are considered low risk of metastasis or recurrence, while 2A and 2B are considered high risk. Of the 17 cases, we had fifteen group 1A (88.23%), one 1B (5.88%), and one 2B (5.88%) result. Overall frequency of dermoscopic features is as follows; 100% of lesions presented with asymmetry, 47% with round structures, and 70.6% with blue-white color. The average time providers spent explaining and ordering the test was 15 minutes, with a range of 10 to 20 minutes.This study represents our experience and understanding of the dermatologistrsquo;s role ordering 31-GEP in the care pathway of melanoma patients and we recommend that dermatology providers consider ordering the test for newly diagnosed CMM patients. J Drugs Dermatol. 2022;21(12): doi:10.36849/JDD.6889.

Watercore is a common physiological disorder in pear and is closely related to excessive accumulation of sorbitol and sucrose. Our previous research found the watercore incidence of ‘Akibae’ (Pyrus pyrifolia cv. Akibae) fruit significantly decreased after boron application (BA). Moreover, foliar spray of boric acid also significantly improved fruit quality. To uncover the mechanisms underlying pear fruit response to BA, a comprehensive transcriptome analysis was performed in this study. Transcriptome results revealed a total of 3146 up-regulated and 1145 down-regulated differently expressed genes (DEGs) between control and treated fruits of ‘Akibae’ pear, respectively. BA significantly induced expression of sorbitol metabolism and sucrose metabolism genes. Besides, BA also increased the expression of starch degradation, fatty acid synthesis, IAA (indole-3-acetic acid) degradation, GA (gibberellin acid) synthesis and inhibit the expression of ethylene synthesis genes. Overall, these findings suggested that BA alleviated ‘Akibae’ watercore occurrence and improve fruit quality by regulating the decrease of sorbitol and sucrose, increased of fatty acid and a balance of plant hormone. Our results provided further information for understanding the molecular mechanism of the effect of boron application on pear fruit.

Backgrounds. Both exosome and circular RNA (circRNA) have been reported to participate in the pathogenesis of type 1 diabetes mellitus (T1DM). However, the exact role of exosomal circRNA in T1DM is largely unknown. Here, we identified the exosomal circRNA expression profiles in the plasma of T1DM patients and explored their potential function using bioinformatics analysis. Material and Methods. Exosomes were extracted by the size exclusion chromatography method from plasma of 10 T1DM patients and 10 age- and sex- matched control subjects. Illumina Novaseq6000 platform was used to detect the exosomal circRNA expression profiles. Multiple bioinformatics analysis was applied to investigate the potential biological functions of exosomal circRNAs. Results. A total of 784 differentially expressed exosomal circRNAs have been identified in T1DM patients, of which 528 were upregulated and 256 were downregulated. Gene Ontology analysis enriched terms such as protein ubiquitination involved in ubiquitin-dependent protein catabolic protein (GO:0042787), membrane (GO:0016020), and GTPase activator activity (GO:0005096). The most enriched pathway in Kyoto Encyclopedia of Genes and Genomes was ubiquitin-mediated proteolysis (ko04120). The miRNA-targeting prediction method was used to identify the miRNAs that bind to circRNAs, and circRNA-miRNA-mRNA pathways were constructed, indicating that interactions between circRNA, miRNA, and gene might be involved in the disease progression. Conclusions. The present study identified the exosomal circRNA expression profiles in T1DM for the first time. Our results threw novel insights into the molecular mechanisms of T1DM.

Screening new compounds for potential bioactivities against cellular targets is vital for drug discovery and chemical safety. Transcriptomics offers an efficient approach for assessing global gene expression changes, but interpreting chemical mechanisms from these data is often challenging. Connectivity mapping is a potential data-driven avenue for linking chemicals to mechanisms based on the observation that many biological processes are associated with unique gene expression signatures (gene signatures). However, mining the effects of a chemical on gene signatures for biological mechanisms is challenging because transcriptomic data contain thousands of noisy genes. New connectivity mapping approaches seeking to distinguish signal from noise continue to be developed, spurred by the promise of discovering chemical mechanisms, new drugs, and disease targets from burgeoning transcriptomic data. Here, we analyze these approaches in terms of diverse transcriptomic technologies, public databases, gene signatures, pattern-matching algorithms, and statistical evaluation criteria. To navigate the complexity of connectivity mapping, we propose a harmonized scheme to coherently organize and compare published workflows. We first standardize concepts underlying transcriptomic profiles and gene signatures based on various transcriptomic technologies such as microarrays, RNA-Seq, and L1000 and discuss the widely used data sources such as Gene Expression Omnibus, ArrayExpress, and MSigDB. Next, we generalize connectivity mapping as a pattern-matching task for finding similarity between a query (e.g., transcriptomic profile for new chemical) and a reference (e.g., gene signature of known target). Published pattern-matching approaches fall into two main categories: vector-based use metrics like correlation, Jaccard index, etc., and aggregation-based use parametric and nonparametric statistics (e.g., gene set enrichment analysis). The statistical methods for evaluating the performance of different approaches are described, along with comparisons reported in the literature on benchmark transcriptomic data sets. Lastly, we review connectivity mapping applications in toxicology and offer guidance on evaluating chemical-induced toxicity with concentration-response transcriptomic data. In addition to serving as a high-level guide and tutorial for understanding and implementing connectivity mapping workflows, we hope this review will stimulate new algorithms for evaluating chemical safety and drug discovery using transcriptomic data.

Triple-negative breast cancer (TNBC) is an aggressive disease that disproportionately affects African American (AA) women. Limited targeted therapeutic options exist for patients with TNBC. Here, we employ spatial transcriptomics to interrogate tissue from a racially diverse TNBC cohort to comprehensively annotate the transcriptional states of spatially resolved cellular populations. A total of 38,706 spatial features from a cohort of 28 sections from 14 patients were analyzed. Intratumoral analysis of spatial features from individual sections revealed heterogeneous transcriptional substructures. However, integrated analysis of all samples resulted in nine transcriptionally distinct clusters that mapped across all individual sections. Furthermore, novel use of join count analysis demonstrated nonrandom directional spatial dependencies of the transcriptionally defined shared clusters, supporting a conserved spatio-transcriptional architecture in TNBC. These findings were substantiated in an independent validation cohort comprising 17,861 spatial features representing 15 samples from 8 patients. Stratification of samples by race revealed race-associated differences in hypoxic tumor content and regions of immune-rich infiltrate. Overall, this study combined spatial and functional molecular analyses to define the tumor architecture of TNBC, with potential implications in understanding TNBC disparities. Significance: Spatial transcriptomics profiling of a diverse cohort of triple-negative breast cancers and innovative informatics approaches reveal a conserved cellular architecture across cancers and identify proportional differences in tumor cell composition by race.

Motivation Catastrophic transitions are ubiquitous in the dynamic progression of complex biological systems; that is, a critical transition at which complex systems suddenly shift from one stable state to another occurs. Identifying such a critical point or tipping point is essential for revealing the underlying mechanism of complex biological systems. However, it is difficult to identify the tipping point since few significant differences in the critical state are detected in terms of traditional static measurements. Results In this study, by exploring the dynamic changes in gene cooperative effects between the before-transition and critical states, we presented a model-free approach, the directed-network rank score (DNRS), to detect the early-warning signal of critical transition in complex biological systems. The proposed method is applicable to both bulk and single-cell RNA-sequencing (scRNA-seq) data. This computational method was validated by the successful identification of the critical or pre-transition state for both simulated and six real datasets, including three scRNA-seq datasets of embryonic development and three tumor datasets. In addition, the functional and pathway enrichment analyses suggested that the corresponding DNRS signaling biomarkers were involved in key biological processes. Availability and implementation The source code is freely available at https://github.com/zhongjiayuan/DNRS. Supplementary information Supplementary data are available at Bioinformatics online.

Background. Rheumatoid arthritis (RA) is recognized as a chronic inflammatory disease featured by pathological synovial inflammation. Currently, the underlying pathophysiological mechanisms of RA remain unclear. In the study, we attempted to explore the underlying mechanisms of RA and provide potential targets for the therapy of RA via bioinformatics analysis. Methods. We downloaded four microarray datasets (GSE77298, GSE55235, GSE12021, and GSE55457) from the GEO database. Firstly, GSE77298 and GSE55457 were identified DEGs by the “limma” and “sva” packages of R software. Then, we performed GO, KEGG, and GSEA enrichment analyses to further analyze the function of DEGs. Hub genes were screened using LASSO analysis and SVM-RFE analysis. To further explore the differences of the expression of hub genes in healthy control and RA patient synovial tissues, we calculated the ROC curves and AUC. The expression levels of hub genes were verified in synovial tissues of normal and RA rats by qRT-PCR and western blot. Furthermore, the CIBERSORTx was implemented to assess the differences of infiltration in 22 immune cells between normal and RA synovial tissues. We explored the association between hub genes and infiltrating immune cells. Results. CRTAM, CXCL13, and LRRC15 were identified as RA’s potential hub genes by machine learning and LASSO algorithms. In addition, we verified the expression levels of three hub genes in the synovial tissue of normal and RA rats by PCR and western blot. Moreover, immune cell infiltration analysis showed that plasma cells, T follicular helper cells, M0 macrophages, M1 macrophages, and gamma delta T cells may be engaged in the development and progression of RA. Conclusions. In brief, our study identified and validated that three hub genes CRTAM, CXCL13, and LRRC15 might involve in the pathological development of RA, which could provide novel perspectives for the diagnosis and treatment with RA.

With the advancement of single-cell separation techniques and high-throughput sequencing platforms, single-cell RNA-sequencing (scRNA-seq) has emerged as a vital technology for understanding tissue and organ systems at cellular resolution. Through transcriptional analysis, it is possible to characterize unique or rare cell types, interpret their interactions, and reveal novel functional states or shifts in developmental stages. As such, this technology is uniquely suited for studying the cells within the human ovary. The ovary is a cellularly heterogeneous organ that houses follicles, the reproductive and endocrine unit that consists of an oocyte surrounded by hormone-producing support cells, as well as many other cell populations constituting stroma, vasculature, lymphatic, and immune components. Here we review studies that have utilized scRNA-seq technology to analyze cells from healthy human ovaries and discuss the single-cell isolation techniques used. We identified two overarching applications for scRNA-seq in the human ovary. The first applies this technology to investigate transcriptional differences in oocytes/eggs from patients undergoing in vitro fertilization treatments to ultimately improve clinical outcomes. The second utilizes scRNA-seq for the pursuit of creating a comprehensive single-cell atlas of the human ovary. The knowledge gained from these studies underscores the importance of scRNA-seq technologies in unlocking a new biological understanding of the human ovary.

Motivation Gene expression imputation has been an essential step of the single-cell RNA-Seq data analysis workflow. Among several deep-learning methods, the debut of scGNN gained substantial recognition in 2021 for its superior performance and the ability to produce a cell–cell graph. However, the implementation of scGNN was relatively time-consuming and its performance could still be optimized. Results The implementation of scGNN 2.0 is significantly faster than scGNN thanks to a simplified close-loop architecture. For all eight datasets, cell clustering performance was increased by 85.02% on average in terms of adjusted rand index, and the imputation Median L1 Error was reduced by 67.94% on average. With the built-in visualizations, users can quickly assess the imputation and cell clustering results, compare against benchmarks and interpret the cell–cell interaction. The expanded input and output formats also pave the way for custom workflows that integrate scGNN 2.0 with other scRNA-Seq toolkits on both Python and R platforms. Availability and implementation scGNN 2.0 is implemented in Python (as of version 3.8) with the source code available at https://github.com/OSU-BMBL/scGNN2.0. Supplementary information Supplementary data are available at Bioinformatics online.

Tongue squamous cell carcinoma (TSCC) is considered the most common malignant tumor among the oral squamous cell carcinomas with a poor prognosis. Understanding the underlying molecular mechanisms that underpin TSCC and its treatments is the focus of the research. Deregulated expression of microRNAs (miRNAs) has recently been implicated in various biological processes linked to cancer. Therefore, in this study, we attempted to investigate miRNAs and their targets expressed in TSCC, which could be involved in its oncogenesis. We performed next-generation sequencing of small RNAs and transcriptomes in H357 TSCC cell line and human oral keratinocytes as a control to find miRNAs and mRNAs that are differentially expressed (DE), which were then supplemented with additional expression datasets from databases, yielding 269 DE miRNAs and 2094 DE genes. The target prediction followed by pathway and disease function analysis revealed that the DE targets were significantly associated with the key processes and pathways, such as apoptosis, epithelial–mesenchymal transition, endocytosis and vascular endothelial growth factor signaling pathways. Furthermore, the top 12 DE targets were chosen based on their involvement in more than one cancer-related pathway, of which 6 genes are targeted by miR-128-3p. Real-time quantitative PCR validation of this miRNA and its targets in H357 and SCC9 TSCC cells confirmed their possible targeting from their reciprocal expression, with MAP2K7 being a critical target that might be involved in oncogenesis and progression of TSCC by acting as a tumor suppressor. Further research is underway to understand how miR-128-3p regulates oncogenesis in TSCC via MAP2K7 and associated pathways.

Background: Hepatocellular Carcinoma (HCC) is one of the most common liver malignancies in the world. With highly invasive biological characteristics and lack of obvious clinical manifestations, hepatocellular Carcinoma usually has a poor prognosis and ranks fourth in cancer mortality. The aetiology and exact molecular mechanism of primary hepatocellular carcinoma are still unclear. This work aims to help identify biomarkers of early HCC diagnosis or prognosis. Results: In this study, Expression data and clinical information of HTSEQ-Counts were downloaded from The Cancer Genome Atlas (TCGA) database, and Gene Expression map GSE121248 was downloaded from Gene Expression Omnibus (GEO). By differentially expressed genes (DEGs) and Weighted Gene co-expression Network Analysis（WGCNA）searched for modules in the two databases that had the same effect on the biological characteristics of HCC, and extracted the module genes with the highest positive correlation with HCC from two databases, and finally obtained overlapping genes. Then, we performed functional enrichment analysis on the overlapping genes to understand their potential biological functions. The top ten hub genes were screened according to MCC through the String database and Cytoscape software. By survival analysis, high expression of CDK1, CCNA2, CDC20, KIF11, DLGAP5, KIF20A, ASPM, CEP55, TPX2 was associated with poorer overall survival (OS) of HCC patients. The DFS curve was plotted using the online website GEPIA2. Finally, based on the enrichment of these genes in the KEGG pathway, real hub genes were screened out, which were CDK1, CCNA2 and CDC20 respectively. Conclusions: High expression of these three genes was negatively correlated with survival time in HCC, and the expression of CDK1, CCNA2 and CDC20 were significantly higher in tumour tissues of HCC patients than in normal liver tissues as verified again by the HPA database. All in all, this provides a new feasible target for early and accurate diagnosis of HCC, clinical diagnosis, treatment and prognosis.