Gene regulatory roles of growth and differentiation factors in retinal development.

Retinal ganglion cell (RGC) differentiation is tightly controlled by extrinsic and intrinsic factors. Growth and differentiation factor 15 (GDF-15) promotes RGC differentiation, opposite to GDF-11 which inhibits RGC differentiation, both in the mouse retina and in human stem cells. To deepen our understanding of how these two closely related molecules confer opposing effects on retinal development, here we assess the transcriptional profiles of mouse retinal progenitors exposed to exogenous GDF-11 or -15. We find a dichotomous effect of GDF-15 on RGC differentiation, decreasing RGCs expressing residual pro-proliferative genes and increasing RGCs expressing non-proliferative genes, suggestive of greater RGC maturation. Furthermore, GDF-11 promoted the differentiation of photoreceptors and amacrine cells. These data enhance our understanding of the mechanisms underlying the differentiation of RGCs and photoreceptors from retinal progenitors and suggest new approaches to the optimization of protocols for the differentiation of these cell types.

The investigation of potential mechanism of Fuzhengkangfu Decoction against Diabetic myocardial injury based on a combined strategy of network pharmacology, transcriptomics, and experimental verification.

BACKGROUND AND OBJECTIVES: Diabetic cardiomyopathy (DCM) is a cardiac condition resulting from myocardial damage caused by diabetes mellitus (DM), currently lacking specific therapeutic interventions. Fuzhengkangfu decoction (FZK) plays an important role in the prevention and treatment of various cardiovascular diseases. However, the efficacy and potential mechanisms of FZK are not fully understood. This study aims to investigate the protective effect and mechanisms of FZK against DCM. METHODOLOGIES: Rats were given a high-calorie diet along with a low dosage of streptozotocin (STZ) to establish a rat model of DCM. The diabetic rats received FZK or normal saline subcutaneously for 12 weeks. Echocardiography was conducted to evaluate their heart function characteristics. Rat heart morphologies were assessed using Sirius Red staining and H&E staining. Transcriptome sequencing analysis and network pharmacology were used to reveal possible targets and mechanisms. Molecular docking was conducted to validate the association between the primary components of FZK and the essential target molecules. Finally, both in vitro and in vivo studies were conducted on the cardioprotective properties and mechanism of FZK. RESULTS: According to the results of network pharmacology, FZK may prevent DCM by reducing oxidative stress and preventing apoptosis. Transcriptomics confirmed that FZK protected against DCM-induced myocardial fibrosis and remodelling, as predicted by network pharmacology, and suggested that FZK regulated the expression of oxidative stress and apoptosis-related proteins. Integrating network pharmacology and transcriptome analysis results revealed that the AGE-RAGE signalling pathway-associated MMP2, SLC2A1, NOX4, CCND1, and CYP1A1 might be key targets. Molecular docking showed that Poricoic acid A and 5-O-Methylvisammioside had the highest docking activities with these targets. We further conducted in vivo experiments, and the results showed that FZK significantly attenuated left ventricular remodelling, reduced myocardial fibrosis, and improved cardiac contractile function. And, our study demonstrated that FZK effectively reduced oxidative stress and apoptosis of cardiomyocytes. The data showed that Erk, NF-κB, and Caspase 3 phosphorylation was significantly inhibited, and Bcl-2/Bax was significantly increased after FZK treatment. In vitro, FZK significantly reduced AGEs-induced ROS increase and apoptosis in cardiomyocytes. Furthermore, FZK significantly inhibited the phosphorylation of Erk and NF-κB proteins and decreased the expression of MMP2. All the results confirmed that FZK inhibited the activation of the Erk/NF-κB pathway in AGE-RAGE signalling and alleviated oxidative stress and apoptosis of cardiomyocytes. In summary, we verified that FZK protects against DCM by inhibiting myocardial apoptotic remodelling through the suppression of the AGE-RAGE signalling pathway. CONCLUSION: In conclusion, our research indicates that FZK demonstrates anti-cardiac dysfunction properties by reducing oxidative stress and cardiomyocyte apoptosis through the AGE-RAGE pathway in DCM, showing potential for therapeutic use.

Development of a cellular assay as a personalized model for testing chronic wound therapeutics.

Exudates of non-healing wounds contain drivers of pathogenicity. We utilized >800 exudates from non-healing and healing wounds of diverse etiologies, collected by three different methods, to develop a wound-specific, cell-based functional biomarker assay. Human dermal fibroblast proliferation served as readout to a) to differentiate between healing and non-healing wounds, b) follow the healing process of individual patients, and c) assess the effects of therapeutics for chronic wounds ex vivo. We observed a strong correlation between wound chronicity and inhibitory effects of individual exudates on fibroblast proliferation, with good diagnostic sensitivity (76-90%, depending on the sample collection method). Transition of a clinically non-healing to a healing phenotype restored fibroblast proliferation and extracellular matrix formation while reducing inflammatory cytokine production. Transcriptional analysis of fibroblasts exposed to ex vivo non-healing wound exudates revealed an induction of inflammatory cytokine- and chemokine pathways and the unfolded protein response, indicating that these changes may contribute to the pathology of non-healing wounds. Testing the wound therapeutics platelet derived growth factor and silver sulfadiazine yielded responses in line with clinical experience and indicate the usefulness of the assay to search for and profile new therapeutics.

Metabolic and mitochondria alterations induced by SARS-CoV-2 accessory proteins ORF3a, ORF9b, ORF9c and ORF10.

Antiviral signaling, immune response and cell metabolism are dysregulated by SARS-CoV-2, the causative agent of COVID-19. Here, we show that SARS-CoV-2 accessory proteins ORF3a, ORF9b, ORF9c and ORF10 induce a significant mitochondrial and metabolic reprogramming in A549 lung epithelial cells. While ORF9b, ORF9c and ORF10 induced largely overlapping transcriptomes, ORF3a induced a distinct transcriptome, including the downregulation of numerous genes with critical roles in mitochondrial function and morphology. On the other hand, all four ORFs altered mitochondrial dynamics and function, but only ORF3a and ORF9c induced a marked alteration in mitochondrial cristae structure. Genome-Scale Metabolic Models identified both metabolic flux reprogramming features both shared across all accessory proteins and specific for each accessory protein. Notably, a downregulated amino acid metabolism was observed in ORF9b, ORF9c and ORF10, while an upregulated lipid metabolism was distinctly induced by ORF3a. These findings reveal metabolic dependencies and vulnerabilities prompted by SARS-CoV-2 accessory proteins that may be exploited to identify new targets for intervention.

In-vivo neuronal dysfunction by Aß and tau overlaps with brain-wide inflammatory mechanisms in Alzheimer's disease.

The molecular mechanisms underlying neuronal dysfunction in Alzheimer's disease (AD) remain uncharacterized. Here, we identify genes, molecular pathways and cellular components associated with whole-brain dysregulation caused by amyloid-beta (Aß) and tau deposits in the living human brain. We obtained in-vivo resting-state functional MRI (rs-fMRI), Aß- and tau-PET for 47 cognitively unimpaired and 16 AD participants from the Translational Biomarkers in Aging and Dementia cohort. Adverse neuronal activity impacts by Aß and tau were quantified with personalized dynamical models by fitting pathology-mediated computational signals to the participant's real rs-fMRIs. Then, we detected robust brain-wide associations between the spatial profiles of Aß-tau impacts and gene expression in the neurotypical transcriptome (Allen Human Brain Atlas). Within the obtained distinctive signature of in-vivo neuronal dysfunction, several genes have prominent roles in microglial activation and in interactions with Aß and tau. Moreover, cellular vulnerability estimations revealed strong association of microglial expression patterns with Aß and tau's synergistic impact on neuronal activity (q < 0.001). These results further support the central role of the immune system and neuroinflammatory pathways in AD pathogenesis. Neuronal dysregulation by AD pathologies also associated with neurotypical synaptic and developmental processes. In addition, we identified drug candidates from the vast LINCS library to halt or reduce the observed Aß-tau effects on neuronal activity. Top-ranked pharmacological interventions target inflammatory, cancer and cardiovascular pathways, including specific medications undergoing clinical evaluation in AD. Our findings, based on the examination of molecular-pathological-functional interactions in humans, may accelerate the process of bringing effective therapies into clinical practice.

Prospective, International, Multisite Comparison of Platelet Isolation Techniques for Genome-Wide Transcriptomics: Communication from the SSC of the ISTH.

Genome-wide platelet transcriptomics is increasingly used to uncover new aspects of platelet biology and as a diagnostic and prognostic tool. Nevertheless, platelet isolation methods for transcriptomic studies are not standardized, introducing challenges for cross-study comparisons, data integration, and replication. In this prospective multicenter study, called "Standardizing Platelet Transcriptomics for Discovery, Diagnostics, and Therapeutics in the Thrombosis and Hemostasis Community (STRIDE)" by the ISTH SSCs, we assessed how three of the most commonly used platelet isolation protocols influence metrics from next-generation bulk RNA sequencing and functional assays. Compared with washing alone, more stringent removal of leukocytes by anti-CD45 beads or PALLTM filters resulted in a sufficient quantity of RNA for next-generation sequencing and similar quality of RNA sequencing metrics. Importantly, stringent removal of leukocytes resulted in the lower relative expression of known leukocyte-specific genes and the higher relative expression of known platelet-specific genes. The results were consistent across enrolling sites, suggesting the techniques are transferrable and reproducible. Moreover, all three isolation techniques did not influence basal platelet reactivity, but agonist-induced integrin αIIbß3 activation is reduced by anti-CD45 bead isolation compared to washing alone. In conclusion, the isolation technique chosen influences genome-wide transcriptional and functional assays in platelets. These results should help the research community make informed choices about platelet isolation techniques in their own platelet studies.

Integrating spatial transcriptomics and bulk RNA-seq: predicting gene expression with enhanced resolution through graph attention networks.

Spatial transcriptomics data play a crucial role in cancer research, providing a nuanced understanding of the spatial organization of gene expression within tumor tissues. Unraveling the spatial dynamics of gene expression can unveil key insights into tumor heterogeneity and aid in identifying potential therapeutic targets. However, in many large-scale cancer studies, spatial transcriptomics data are limited, with bulk RNA-seq and corresponding Whole Slide Image (WSI) data being more common (e.g. TCGA project). To address this gap, there is a critical need to develop methodologies that can estimate gene expression at near-cell (spot) level resolution from existing WSI and bulk RNA-seq data. This approach is essential for reanalyzing expansive cohort studies and uncovering novel biomarkers that have been overlooked in the initial assessments. In this study, we present STGAT (Spatial Transcriptomics Graph Attention Network), a novel approach leveraging Graph Attention Networks (GAT) to discern spatial dependencies among spots. Trained on spatial transcriptomics data, STGAT is designed to estimate gene expression profiles at spot-level resolution and predict whether each spot represents tumor or non-tumor tissue, especially in patient samples where only WSI and bulk RNA-seq data are available. Comprehensive tests on two breast cancer spatial transcriptomics datasets demonstrated that STGAT outperformed existing methods in accurately predicting gene expression. Further analyses using the TCGA breast cancer dataset revealed that gene expression estimated from tumor-only spots (predicted by STGAT) provides more accurate molecular signatures for breast cancer sub-type and tumor stage prediction, and also leading to improved patient survival and disease-free analysis. Availability: Code is available at https://github.com/compbiolabucf/STGAT.

A comparison of the TempO-Seq and Affymetrix microarray platform using RTqPCR validation.

Next-generation risk assessment relies on mechanistic data from new approach methods, including transcriptome data. Various technologies, such as high-throughput targeted sequencing methods and microarray technologies based on hybridization with complementary probes, are used to determine differentially expressed genes (DEGs). The integration of data from different technologies requires a good understanding of the differences arising from the use of various technologies.To better understand the differences between the TempO-Seq platform and Affymetrix chip technology, whole-genome data for the volatile compound dimethylamine were compared. Selected DEGs were also confirmed using RTqPCR validation. Although the overlap of DEGs between TempO-Seq and Affymetrix was no higher than 37%, a comparison of the gene regulation in terms of log2fold changes revealed a very high concordance. RTqPCR confirmed the majority of DEGs from either platform in the examined dataset. Only a few conflicts were found (11%), while 22% were not confirmed, and 3% were not detected.Despite the observed differences between the two platforms, both can be validated using RTqPCR. Here we highlight some of the differences between the two platforms and discuss their applications in toxicology.

Integrating single-nucleus RNA sequencing and spatial transcriptomics to elucidate a specialized subpopulation of astrocytes, microglia and vascular cells in brains of mouse model of lipopolysaccharide-induced sepsis-associated encephalopathy.

BACKGROUND: Understanding the mechanism behind sepsis-associated encephalopathy (SAE) remains a formidable task. This study endeavors to shed light on the complex cellular and molecular alterations that occur in the brains of a mouse model with SAE, ultimately unraveling the underlying mechanisms of this condition. METHODS: We established a murine model using intraperitoneal injection of lipopolysaccharide (LPS) in wild type and Anxa1-/- mice and collected brain tissues for analysis at 0-hour, 12-hour, 24-hour, and 72-hour post-injection. Utilizing advanced techniques such as single-nucleus RNA sequencing (snRNA-seq) and Stereo-seq, we conducted a comprehensive characterization of the cellular responses and molecular patterns within the brain. RESULTS: Our study uncovered notable temporal differences in the response to LPS challenge between Anxa1-/- (annexin A1 knockout) and wild type mice, specifically at the 12-hour and 24-hour time points following injection. We observed a significant increase in the proportion of Astro-2 and Micro-2 cells in these mice. These cells exhibited a colocalization pattern with the vascular subtype Vas-1, forming a distinct region known as V1A2M2, where Astro-2 and Micro-2 cells surrounded Vas-1. Moreover, through further analysis, we discovered significant upregulation of ligands and receptors such as Timp1-Cd63, Timp1-Itgb1, Timp1-Lrp1, as well as Ccl2-Ackr1 and Cxcl2-Ackr1 within this region. In addition, we observed a notable increase in the expression of Cd14-Itgb1, Cd14-Tlr2, and Cd14-C3ar1 in regions enriched with Micro-2 cells. Additionally, Cxcl10-Sdc4 showed broad upregulation in brain regions containing both Micro-2 and Astro-2 cells. Notably, upon LPS challenge, there was an observed increase in Anxa1 expression in the mouse brain. Furthermore, our study revealed a noteworthy increase in mortality rates following Anxa1 knockdown. However, we did not observe substantial differences in the types, numbers, or distribution of other brain cells between Anxa1-/- and wildtype mice over time. Nevertheless, when comparing the 24-hour post LPS injection time point, we observed a significant decrease in the proportion and distribution of Micro-2 and Astro-2 cells in the vicinity of blood vessels in Anxa1-/- mice. Additionally, we noted reduced expression levels of several ligand-receptor pairs including Cd14-Tlr2, Cd14-C3ar1, Cd14-Itgb1, Cxcl10-Sdc4, Ccl2-Ackr1, and Cxcl2-Ackr1. CONCLUSIONS: By combining snRNA-seq and Stereo-seq techniques, our study successfully identified a distinctive cellular colocalization, referred to as a special pathological niche, comprising Astro-2, Micro-2, and Vas-1 cells. Furthermore, we observed an upregulation of ligand-receptor pairs within this niche. These findings suggest a potential association between this cellular arrangement and the underlying mechanisms contributing to SAE or the increased mortality observed in Anxa1 knockdown mice.

A single-cell transcriptomic study of heterogeneity in human embryonic tanycytes.

Disruptions in energy homeostasis can lead to diseases like obesity and diabetes, affecting millions of people each year. Tanycytes, the adult stem cells in the hypothalamus, play crucial roles in assisting hypothalamic neurons in maintaining energy balance. Although tanycytes have been extensively studied in rodents, our understanding of human tanycytes remains limited. In this study, we utilized single-cell transcriptomics data to explore the heterogeneity of human embryonic tanycytes, investigate their gene regulatory networks, analyze their intercellular communication, and examine their developmental trajectory. Our analysis revealed the presence of two clusters of ß tanycytes and three clusters of α tanycytes in our dataset. Surprisingly, human embryonic tanycytes displayed significant similarities to mouse tanycytes in terms of marker gene expression and transcription factor activities. Trajectory analysis indicated that α tanycytes were the first to be generated, giving rise to ß tanycytes in a dorsal-ventral direction along the third ventricle. Furthermore, our CellChat analyses demonstrated that tanycytes generated earlier along the developmental lineages exhibited increased intercellular communication compared to those generated later. In summary, we have thoroughly characterized the heterogeneity of human embryonic tanycytes from various angles. We are confident that our findings will serve as a foundation for future research on human tanycytes.

Temperatures above 37°C increase virulence of a convergent Klebsiella pneumoniae sequence type 307 strain.

Background: Convergence of Klebsiella pneumoniae (KP) pathotypes has been increasingly reported in recent years. These pathogens combine features of both multidrug-resistant and hypervirulent KP. However, clinically used indicators for hypervirulent KP identification, such as hypermucoviscosity, appear to be differentially expressed in convergent KP, potential outbreak clones are difficult to identify. We aimed to fill such knowledge gaps by investigating the temperature dependence of hypermucoviscosity and virulence in a convergent KP strain isolated during a clonal outbreak and belonging to the high-risk sequence type (ST)307. Methods: Hypermucoviscosity, biofilm formation, and mortality rates in Galleria mellonella larvae were examined at different temperatures (room temperature, 28°C, 37°C, 40°C and 42°C) and with various phenotypic experiments including electron microscopy. The underlying mechanisms of the phenotypic changes were explored via qPCR analysis to evaluate plasmid copy numbers, and transcriptomics. Results: Our results show a temperature-dependent switch above 37°C towards a hypermucoviscous phenotype, consistent with increased biofilm formation and in vivo mortality, possibly reflecting a bacterial response to fever-like conditions. Furthermore, we observed an increase in plasmid copy number for a hybrid plasmid harboring carbapenemase and rmpA genes. However, transcriptomic analysis revealed no changes in rmpA expression at higher temperatures, suggesting alternative regulatory pathways. Conclusion: This study not only elucidates the impact of elevated temperatures on hypermucoviscosity and virulence in convergent KP but also sheds light on previously unrecognized aspects of its adaptive behavior, underscoring its resilience to changing environments.

Decoding heterogeneous and coordinated tissue architecture in glioblastoma using spatial transcriptomics.

Glioblastoma multiforme (GBM) is one of the most lethal brain tumors, characterized by profound heterogeneity. While single-cell transcriptomic studies have revealed extensive intra-tumor heterogeneity, shed light on intra-tumor diversity, spatial intricacies remain largely unexplored. Leveraging clinical GBM specimens, this study employs spatial transcriptomics technology to delve into gene expression heterogeneity. Our investigation unveils a significant enrichment of tissue stem cell signature in regions bordering necrosis and the peritumoral area, positively correlated with the mesenchymal subtype signature. Moreover, upregulated genes in these regions are linked with extracellular matrix (ECM)-receptor interaction, proteoglycans, as well as vascular endothelial growth factor (VEGF) and angiopoietin-Tie (ANGPT) signaling pathways. In contrast, signatures related to glycogen metabolism and oxidative phosphorylation show no relevance to pathological zoning, whereas creatine metabolism signature is notably exclusive to vascular-enriched areas. These spatial profiles not only offer valuable references but also pave the way for future in-depth functional and mechanistic investigations into GBM progression.

The alarmin IL-33 exacerbates pulmonary inflammation and immune dysfunction in SARS-CoV-2 infection.

Dysregulated host immune responses contribute to disease severity and worsened prognosis in COVID-19 infection and the underlying mechanisms are not fully understood. In this study, we observed that IL-33, a damage-associated molecular pattern molecule, is significantly increased in COVID-19 patients and in SARS-CoV-2-infected mice. Using IL-33-/- mice, we demonstrated that IL-33 deficiency resulted in significant decreases in bodyweight loss, tissue viral burdens, and lung pathology. These improved outcomes in IL-33-/- mice also correlated with a reduction in innate immune cell infiltrates, i.e., neutrophils, macrophages, natural killer cells, and activated T cells in inflamed lungs. Lung RNA-seq results revealed that IL-33 signaling enhances activation of inflammatory pathways, including interferon signaling, pathogen phagocytosis, macrophage activation, and cytokine/chemokine signals. Overall, these findings demonstrate that the alarmin IL-33 plays a pathogenic role in SARS-CoV-2 infection and provides new insights that will inform the development of effective therapeutic strategies for COVID-19.

Abnormal metabolism in melanocytes participates in the activation of dendritic cell in halo nevus.

A comprehensive analysis of spatial transcriptomics was carried out to better understand the progress of halo nevus. We found that halo nevus was characterized by overactive immune responses, triggered by chemokines and dendritic cells (DCs), T cells, and macrophages. Consequently, we observed abnormal cell death, such as apoptosis and disulfidptosis in halo nevus, some were closely related to immunity. Interestingly, we identified aberrant metabolites such as uridine diphosphate glucose (UDP-G) within the halo nevus. UDP-G, accompanied by the infiltration of DCs and T cells, exhibited correlations with certain forms of cell death. Subsequent experiments confirmed that UDP-G was increased in vitiligo serum and could activate DCs. We also confirmed that oxidative response is an inducer of UDP-G. In summary, the immune response in halo nevus, including DC activation, was accompanied by abnormal cell death and metabolites. Especially, melanocyte-derived UDP-G may play a crucial role in DC activation.

RNA expression profiling of peritoneal metastasis from pancreatic cancer treated with Pressurized Intraperitoneal Aerosol Chemotherapy (PIPAC).

Objectives: Pressurized Intraperitoneal Aerosol Chemotherapy (PIPAC) is an experimental treatment option in peritoneal metastasis from pancreatic cancer (PM-PC). Aims were to examine mRNA profile of fibrosis due to response after systemic chemotherapy and PIPAC (Regression) compared to treatment-naïve PM-PC and chronic cholecystitis-related peritoneal fibrosis (Controls). Methods: Peritoneal biopsies (PBs) from PM-PC patients who had undergone systemic chemotherapy and PIPAC were evaluated with Peritoneal Regression Grading Score (PRGS). We extracted RNA from PBs with Regression (PRGS 1, n=11), treatment-naïve PM-PC (n=10), and Controls (n=10). Profiling of 800 mRNAs was performed (NanoString nCounter, PanCancer Immuno-Oncology 360 (IO-360) and 30 additional stroma-related mRNAs). Results: Regression vs. PM-PC identified six up-regulated and 197 down-regulated mRNAs (FDR≤0.05), linked to TNF-α signaling via NF-kB, G2M checkpoint, epithelial-mesenchymal transition, estrogen response, and coagulation. Regression vs. Controls identified 43 significantly up-regulated mRNAs, linked to interferon-α response, and down-regulation of 99 mRNAs, linked to TNF-α signaling via NF-kB, inflammatory response, epithelial-mesenchymal transition, KRAS signaling, and hypoxia (FDR≤0.05). Conclusions: In regressive fibrosis of PM-PC after systemic chemotherapy and PIPAC (Regression), downregulation of mRNAs related to key tumor biological pathways was identified. Regression also showed transcriptional differences from unspecific, benign fibrosis (Controls). Future studies should explore whether mRNA profiling of PBs with PM from PC or other primaries holds prognostic or predictive value.

Comparative transcriptome profiling reveals distinct regulatory responses of secondary defensive metabolism in Datura species (Solanaceae) under plant development and herbivory-mediated stress.

Differential expression of genes is key to mediating developmental and stress-related plant responses. Here, we addressed the regulation of plant metabolic responses to biotic stress and the developmental variation of defense-related genes in four species of the genus Datura with variable patterns of metabolite accumulation and development. We combine transcriptome profiling with phylogenomic techniques to analyze gene expression and coexpression in plants subjected to damage by a specialist folivore insect. We found (1) common overall gene expression in species of similar chemical profiles, (2) species-specific responses of proteins involved in specialized metabolism, characterized by constant levels of gene expression coupled with transcriptional rearrangement, and (3) induction of transcriptional rearrangement of major terpene and tropane alkaloid genes upon herbivory. Our results indicate differential modulation of terpene and tropane metabolism linked to jasmonate signaling and specific transcription factors to regulate developmental variation and stress programs, and suggest plastic adaptive responses to cope with herbivory. The transcriptional profiles of specialized metabolism shown here reveal complex genetic control of plant metabolism and contribute to understanding the molecular basis of adaptations and the physiological variation of significant ecological traits.

Comparative transcriptomics analysis on Senecavirus A-infected and non-infected cells.

Senecavirus A (SVA) is an emerging virus that causes the vesicular disease in pigs, clinically indistinguishable from other high consequence vesicular diseases. This virus belongs to the genus Senecavirus in the family Picornaviridae. Its genome is a positive-sense, single-stranded RNA, approximately 7,300 nt in length, with a 3' poly(A) tail but without 5'-end capped structure. SVA can efficiently propagate in different cells, including some non-pig-derived cell lines. A wild-type SVA was previously rescued from its cDNA clone using reverse genetics in our laboratory. In the present study, the BSR-T7/5 cell line was inoculated with the passage-5 SVA. At 12 h post-inoculation, SVA-infected and non-infected cells were independently collected for the analysis on comparative transcriptomics. The results totally showed 628 differentially expressed genes, including 565 upregulated and 63 downregulated ones, suggesting that SVA infection significantly stimulated the transcription initiation in cells. GO and KEGG enrichment analyses demonstrated that SVA exerted multiple effects on immunity-related pathways in cells. Furthermore, the RNA sequencing data were subjected to other in-depth analyses, such as the single-nucleotide polymorphism, transcription factors, and protein-protein interactions. The present study, along with our previous proteomics and metabolomics researches, provides a multi-omics insight into the interaction between SVA and its hosts.

Examining effects of a novel estrogenic perfluoro-alcohol, 1H,1H,8H,8H-Perfluorooctane-1,8-diol (FC8-diol), using the fathead minnow EcoToxChip.

In a previous in vivo study, adult male fathead minnows (Pimephales promelas) were exposed via water for 4 days to 1H,1H,8H,8H-perfluorooctane-1,8-diol (FC8-diol). The present study expands on the evaluation of molecular responses to this perfluoro-alcohol by analyzing 26 male fathead minnow liver RNA samples from that study (five from each test concentration: 0, 0.018, 0.051, 0.171, and 0.463 mg FC8-diol/L) using fathead minnow EcoToxChips Ver. 1.0. EcoToxChips are a quantitative polymerase chain reaction array that allows for simultaneous measurement of >375 species-specific genes of toxicological interest. Data were analyzed with the online tool EcoToxXplorer. Among the genes analyzed, 62 and 96 were significantly up- and downregulated, respectively, by one or more FC8-diol treatments. Gene expression results from the previous study were validated, showing an upregulation of vitellogenin mRNA (vtg) and downregulation of insulin-like growth factor 1 mRNA (igf1). Additional genes related to estrogen receptor activation including esr2a (estrogen receptor 2a) and esrrb (estrogen related receptor beta) were also affected, providing further confirmation of the estrogenic nature of FC8-diol. Furthermore, genes involved in biological pathways related to lipid and carbohydrate metabolism, innate immune response, endocrine reproduction, and endocrine thyroid were significantly affected. These results both add confidence in the use of the EcoToxChip tool for inferring chemical mode(s) of action and provide further insights into the possible biological effects of FC8-diol. Environ Toxicol Chem 2024;00:1-9. © 2024 SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

SmuMYB113 is the determinant of fruit color in pepino (Solanum muricatum).

Pepino (Solanum muricatum) is an herbaceous crop phylogenetically related to tomato and potato. Pepino fruit vary in color, size and shape, and are eaten fresh. In this study, we use pepino as a fruit model to understand the transcriptional regulatory mechanisms controlling fruit quality. To identify the key genes involved in anthocyanin biosynthesis in pepino, two genotypes were studied that contrasted in foliar and fruit pigmentation. Anthocyanin profiles were analyzed, as well as the expression of genes that encode enzymes for anthocyanin biosynthesis and transcriptional regulators using both RNA-seq and quantitative PCR. The differential expression of the transcription factor genes R2R3 MYB SmuMYB113 and R3MYB SmuATV suggested their association with purple skin and foliage phenotype. Functional analysis of these genes in both tobacco and pepino showed that SmuMYB113 activates anthocyanins, while SmuATV suppresses anthocyanin accumulation. However, despite elevated expression in all tissues, SmuMYB113 does not significantly elevate flesh pigmentation, suggesting a strong repressive background in fruit flesh tissue. These results will aid understanding of the differential regulation controlling fruit quality aspects between skin and flesh in other fruiting species.

Single-cell sequencing of immune cells from the coronary sinus reveals immune mechanisms of the progression of persistent atrial fibrillation.

Identifying the atlas of immune cells from coronary sinus circulation (CSC) of patients with persistent atrial fibrillation (PerAF) may provide new insights into the role of immune cells in the progression of AF. Single-cell sequencing revealed substantial alterations in immune cells from CSCs of patients with PerAF, especially a markedly elevated abundance of T cells, after which we identified a T cell subset: FGFBP2(+)TRDC(-)CD4(-) T cells (Ftc-T cells), which can promote the proliferation of cardiac fibroblasts (CFs),and the proportion of Ftc-T had a positive linear with AF recurrence post catheter ablation (CA). Moreover, IFI27 was found to be highly enriched in Ftc-T cells and promoted CFs proliferation and collagen expression. Altogether, our findings represent a unique resource providing in-depth insights into the heterogeneity of the immune cell from CSC of patients with PerAF and highlight the potential role of Ftc-T cells and IFI27 for AF progression.