Leveraging the antiviral type I interferon system as a first line of defense against SARS-CoV-2 pathogenicity.

The emergence and spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in significant global morbidity, mortality, and societal disruption. A better understanding of virus-host interactions may potentiate therapeutic insights toward limiting this infection. Here we investigated the dynamics of the systemic response to SARS-CoV-2 in hamsters by histological analysis and transcriptional profiling. Infection resulted in consistently high levels of virus in the upper and lower respiratory tracts and sporadic occurrence in other distal tissues. A longitudinal cohort revealed a wave of inflammation, including a type I interferon (IFN-I) response, that was evident in all tissues regardless of viral presence but was insufficient to prevent disease progression. Bolstering the antiviral response with intranasal administration of recombinant IFN-I reduced viral disease, prevented transmission, and lowered inflammation in vivo. This study defines the systemic host response to SARS-CoV-2 infection and supports use of intranasal IFN-I as an effective means of early treatment.

Cactus: A user-friendly and reproducible ATAC-Seq and mRNA-Seq analysis pipeline for data preprocessing, differential analysis, and enrichment analysis.

The ever decreasing cost of Next-Generation Sequencing coupled with the emergence of efficient and reproducible analysis pipelines has rendered genomic methods more accessible. However, downstream analyses are basic or missing in most workflows, creating a significant barrier for non-bioinformaticians. To help close this gap, we developed Cactus, an end-to-end pipeline for analyzing ATAC-Seq and mRNA-Seq data, either separately or jointly. Its Nextflow-, container-, and virtual environment-based architecture ensures efficient and reproducible analyses. Cactus preprocesses raw reads, conducts differential analyses between conditions, and performs enrichment analyses in various databases, including DNA-binding motifs, ChIP-Seq binding sites, chromatin states, and ontologies. We demonstrate the utility of Cactus in a multi-modal and multi-species case study as well as by showcasing its unique capabilities as compared to other ATAC-Seq pipelines. In conclusion, Cactus can assist researchers in gaining comprehensive insights from chromatin accessibility and gene expression data in a quick, user-friendly, and reproducible manner.

A radiomics signature derived from CT imaging to predict MSI status and immunotherapy outcomes in gastric cancer: a multi-cohort study.

BACKGROUND: Accurate microsatellite instability (MSI) testing is essential for identifying gastric cancer (GC) patients eligible for immunotherapy. We aimed to develop and validate a CT-based radiomics signature to predict MSI and immunotherapy outcomes in GC. METHODS: This retrospective multicohort study included a total of 457 GC patients from two independent medical centers in China and The Cancer Imaging Archive (TCIA) databases. The primary cohort (n = 201, center 1, 2017-2022), was used for signature development via Least Absolute Shrinkage and Selection Operator (LASSO) and logistic regression analysis. Two independent immunotherapy cohorts, one from center 1 (n = 184, 2018-2021) and another from center 2 (n = 43, 2020-2021), were utilized to assess the signature's association with immunotherapy response and survival. Diagnostic efficiency was evaluated using the area under the receiver operating characteristic curve (AUC), and survival outcomes were analyzed via the Kaplan-Meier method. The TCIA cohort (n = 29) was included to evaluate the immune infiltration landscape of the radiomics signature subgroups using both CT images and mRNA sequencing data. RESULTS: Nine radiomics features were identified for signature development, exhibiting excellent discriminative performance in both the training (AUC: 0.851, 95%CI: 0.782, 0.919) and validation cohorts (AUC: 0.816, 95%CI: 0.706, 0.926). The radscore, calculated using the signature, demonstrated strong predictive abilities for objective response in immunotherapy cohorts (AUC: 0.734, 95%CI: 0.662, 0.806; AUC: 0.724, 95%CI: 0.572, 0.877). Additionally, the radscore showed a significant association with PFS and OS, with GC patients with a low radscore experiencing a significant survival benefit from immunotherapy. Immune infiltration analysis revealed significantly higher levels of CD8 + T cells, activated CD4 + B cells, and TNFRSF18 expression in the low radscore group, while the high radscore group exhibited higher levels of T cells regulatory and HHLA2 expression. CONCLUSION: This study developed a robust radiomics signature with the potential to serve as a non-invasive biomarker for GC's MSI status and immunotherapy response, demonstrating notable links to post-immunotherapy PFS and OS. Additionally, distinct immune profiles were observed between low and high radscore groups, highlighting their potential clinical implications.

Costs of reproduction are present but latent in eusocial bumblebee queens.

BACKGROUND: The standard evolutionary theory of ageing proposes that ageing occurs because of a trade-off between reproduction and longevity. Eusocial insect queens exhibit positive fecundity-longevity associations and so have been suggested to be counter-examples through not expressing costs of reproduction and through remodelling conserved genetic and endocrine networks regulating ageing and reproduction. If so, eusocial evolution from solitary ancestors with negative fecundity-longevity associations must have involved a stage at which costs of reproduction were suppressed and fecundity and longevity became positively associated. Using the bumblebee (Bombus terrestris), we experimentally tested whether queens in annual eusocial insects at an intermediate level of eusocial complexity experience costs of reproduction, and, using mRNA-seq, the extent to which they exhibit a remodelling of relevant genetic and endocrine networks. Specifically, we tested whether costs of reproduction are present but latent, or whether a remodelling of relevant genetic and endocrine networks has already occurred allowing queens to reproduce without costs. RESULTS: We experimentally increased queens' costs of reproduction by removing their eggs, which caused queens to increase their egg-laying rate. Treatment queens had significantly reduced longevity relative to control queens whose egg-laying rate was not increased. Reduced longevity in treatment queens was not caused by increased worker-to-queen aggression or by increased overall activity in queens. In addition, treatment and control queens differed in age-related gene expression based on mRNA-seq in both their overall expression profiles and the expression of ageing-related genes. Remarkably, these differences appeared to occur principally with respect to relative age, not chronological age. CONCLUSIONS: This study represents the first simultaneously phenotypic and transcriptomic experimental test for a longevity cost of reproduction in eusocial insect queens. The results support the occurrence of costs of reproduction in annual eusocial insects of intermediate social complexity and suggest that reproductive costs are present but latent in queens of such species, i.e. that these queens exhibit condition-dependent positive fecundity-longevity associations. They also raise the possibility that a partial remodelling of genetic and endocrine networks underpinning ageing may have occurred in intermediately eusocial species such that, in unmanipulated conditions, age-related gene expression depends more on chronological than relative age.

A single cell RNA sequencing resource for early sea urchin development.

Identifying cell states during development from their mRNA profiles provides insight into their gene regulatory network. Here, we leverage the sea urchin embryo for its well-established gene regulatory network to interrogate the embryo using single cell RNA sequencing. We tested eight developmental stages in Strongylocentrotus purpuratus, from the eight-cell stage to late in gastrulation. We used these datasets to parse out 22 major cell states of the embryo, focusing on key transition stages for cell type specification of each germ layer. Subclustering of these major embryonic domains revealed over 50 cell states with distinct transcript profiles. Furthermore, we identified the transcript profile of two cell states expressing germ cell factors, one we conclude represents the primordial germ cells and the other state is transiently present during gastrulation. We hypothesize that these cells of the Veg2 tier of the early embryo represent a lineage that converts to the germ line when the primordial germ cells are deleted. This broad resource will hopefully enable the community to identify other cell states and genes of interest to expose the underpinning of developmental mechanisms.

Dissecting heterogeneous cell populations across drug and disease conditions with PopAlign.

Single-cell measurement techniques can now probe gene expression in heterogeneous cell populations from the human body across a range of environmental and physiological conditions. However, new mathematical and computational methods are required to represent and analyze gene-expression changes that occur in complex mixtures of single cells as they respond to signals, drugs, or disease states. Here, we introduce a mathematical modeling platform, PopAlign, that automatically identifies subpopulations of cells within a heterogeneous mixture and tracks gene-expression and cell-abundance changes across subpopulations by constructing and comparing probabilistic models. Probabilistic models provide a low-error, compressed representation of single-cell data that enables efficient large-scale computations. We apply PopAlign to analyze the impact of 40 different immunomodulatory compounds on a heterogeneous population of donor-derived human immune cells as well as patient-specific disease signatures in multiple myeloma. PopAlign scales to comparisons involving tens to hundreds of samples, enabling large-scale studies of natural and engineered cell populations as they respond to drugs, signals, or physiological change.

Genome-Wide Analysis of Stress-Responsive Genes and Alternative Splice Variants in Arabidopsis Roots under Osmotic Stresses.

Plant roots show distinct gene-expression profiles from those of shoots under abiotic stress conditions. In this study, we performed mRNA sequencing (mRNA-Seq) to analyze the transcriptional profiling of Arabidopsis roots under osmotic stress conditions-high salinity (NaCl) and drought (mannitol). The roots demonstrated significantly distinct gene-expression changes from those of the aerial parts under both the NaCl and the mannitol treatment. We identified 68 closely connected transcription-factor genes involved in osmotic stress-signal transduction in roots. Well-known abscisic acid (ABA)-dependent and/or ABA-independent osmotic stress-responsive genes were not considerably upregulated in the roots compared to those in the aerial parts, indicating that the osmotic stress response in the roots may be regulated by other uncharacterized stress pathways. Moreover, we identified 26 osmotic-stress-responsive genes with distinct expressions of alternative splice variants in the roots. The quantitative reverse-transcription polymerase chain reaction further confirmed that alternative splice variants, such as those for ANNAT4, MAGL6, TRM19, and CAD9, were differentially expressed in the roots, suggesting that alternative splicing is an important regulatory mechanism in the osmotic stress response in roots. Altogether, our results suggest that tightly connected transcription-factor families, as well as alternative splicing and the resulting splice variants, are involved in the osmotic stress response in roots.

Fcγ receptor-mediated phagocytosis pathway was involved in phagocytosis of mIgM+ B lymphocytes from largemouth bass (Micropterus salmoides).

The potential for phagocytosis has been proven in teleost B cells, but the research on the regulatory mechanism of phagocytosis remains lacking. In this study, three largemouth bass (Micropterus salmoides) (15 ± 5 g) were injected intraperitoneally with Nocardia seriolae (105 CFU/100 µl/fish) in vivo, and their spleen was collected at 72 h post-infection for mRNA-seq. After the de novo assembly of the paired-end reads, 73,622 unigenes were obtained. Gene expression profiling revealed that 2043 unigenes were differentially expressed after N. seriolae infection, comprising 1285 upregulated and 758 downregulated unigenes (q-value <0.05, log2FC > |2|) of which 181 genes were involved in phagocytosis. The Kyoto Encyclopaedia of Genes and Genomes (KEGG) analysis demonstrated that 12 differentially expressed genes (DEG) associated with phagocytosis were enriched in the Fcγ receptor-mediated phagocytosis signalling pathway. In vitro, the phagocytic ability of mIgM+ B lymphocytes was validated using indirect immunofluorescence assay (IIFA) and fluorescence activating cell sorter (FACS), and the phagocytosis rates of the mIgM+ B lymphocytes incubated with a Lyn inhibitor had decreased from 18.533 ± 6.00% to 11.610 ± 4.236% compared with the unblocked group. These results suggested that the Fcγ receptor-mediated phagocytosis signalling pathway had participated in the phagocytosis of B cells and provide further insight into the role of B cells in innate immunology.

Animal regeneration in the era of transcriptomics.

Animal regeneration, the ability to restore a lost body part, is a process that has fascinated scientists for centuries. In this review, we first present what regeneration is and how it relates to development, as well as the widespread and diverse nature of regeneration in animals. Despite this diversity, animal regeneration includes three common mechanistic steps: initiation, induction and activation of progenitors, and morphogenesis. In this review article, we summarize and discuss, from an evolutionary perspective, the recent data obtained for a variety of regeneration models which have allowed to identify key shared mechanisms that control these main steps of animal regeneration. This review also synthesizes the wealth of high-throughput mRNA sequencing data (bulk mRNA-seq) concerning regeneration which have been obtained in recent years, highlighting the major advances in the regeneration field that these studies have revealed. We stress out that, through a comparative approach, these data provide opportunities to further shed light on the evolution of regeneration in animals. Finally, we point out how the use of single-cell mRNA-seq technology and integration with epigenomic approaches may further help researchers to decipher mechanisms controlling regeneration and their evolution in animals.

Transcriptomic Response of the Diazotrophic Bacteria Gluconacetobacter diazotrophicus Strain PAL5 to Iron Limitation and Characterization of the fur Regulatory Network.

Gluconacetobacter diazotrophicus has been the focus of several studies aiming to understand the mechanisms behind this endophytic diazotrophic bacterium. The present study is the first global analysis of the early transcriptional response of exponentially growing G. diazotrophicus to iron, an essential cofactor for many enzymes involved in various metabolic pathways. RNA-seq, targeted gene mutagenesis and computational motif discovery tools were used to define the G. diazotrophicusfur regulon. The data analysis showed that genes encoding functions related to iron homeostasis were significantly upregulated in response to iron limitations. Certain genes involved in secondary metabolism were overexpressed under iron-limited conditions. In contrast, it was observed that the expression of genes involved in Fe-S cluster biosynthesis, flagellar biosynthesis and type IV secretion systems were downregulated in an iron-depleted culture medium. Our results support a model that controls transcription in G. diazotrophicus by fur function. The G. diazotrophicusfur protein was able to complement an E. colifur mutant. These results provide new insights into the effects of iron on the metabolism of G. diazotrophicus, as well as demonstrate the essentiality of this micronutrient for the main characteristics of plant growth promotion by G. diazotrophicus.

Comparative impact of dietary carbohydrates on the liver transcriptome in two strains of mice.

Excessive long-term consumption of dietary carbohydrates, including glucose, sucrose, or fructose, has been shown to have significant impact on genome-wide gene expression, which likely results from changes in metabolic substrate flux. However, there has been no comprehensive study on the acute effects of individual sugars on the genome-wide gene expression that may reveal the genetic changes altering signaling pathways, subsequent metabolic processes, and ultimately physiological/pathological responses. Considering that gene expressions in response to acute carbohydrate ingestion might be different in nutrient sensitive and insensitive mammals, we conducted comparative studies of genome-wide gene expression by deep mRNA sequencing of the liver in nutrient sensitive C57BL/6J and nutrient insensitive BALB/cJ mice. Furthermore, to determine the temporal responses, we compared livers from mice in the fasted state and following ingestion of standard laboratory mouse chow supplemented with plain drinking water or water containing 20% glucose, sucrose, or fructose. Supplementation with these carbohydrates induced unique extents and temporal changes in gene expressions in a strain specific manner. Fructose and sucrose stimulated gene changes peaked at 3 h postprandial, whereas glucose effects peaked at 12 h and 6 h postprandial in C57BL/6J and BABL/cJ mice, respectively. Network analyses revealed that fructose changed genes were primarily involved in lipid metabolism and were more complex in C57BL/6J than in BALB/cJ mice. These data demonstrate that there are qualitative and antitative differences in the normal physiological responses of the liver between these two strains of mice and C57BL/6J is more sensitive to sugar intake than BALB/cJ.

Regulation of gene expression during the fasting-feeding cycle of the liver displays mouse strain specificity.

The liver maintains metabolic homeostasis by integrating the regulation of nutrient status with both hormonal and neural signals. Many studies on hepatic signaling in response to nutrients have been conducted in mice. However, no in-depth study is currently available that has investigated genome-wide changes in gene expression during the normal physiological fasting-feeding cycle in nutrient-sensitive and -insensitive mice. Using two strains of mice, C57BL/6J and BALB/cJ, and deploying deep RNA-Seq complemented with quantitative RT-PCR, we found that feeding causes substantial and transient changes in gene expression in the livers of both mouse strains. The majority of significantly changed transcripts fell within the areas of biological regulation and cellular and metabolic processes. Among the metabolisms of three major types of macronutrients (i.e. carbohydrates, proteins, and lipids), feeding affected lipid metabolism the most. We also noted that the C57BL/6J and BALB/cJ mice significantly differed in gene expression and in changes in gene expression in response to feeding. In both fasted and fed states, both mouse strains shared common expression patterns for about 10,200 genes, and an additional 400-600 genes were differentially regulated in one strain but not the other. Among the shared genes, more lipogenic genes were induced upon feeding in BABL/cJ than in C57BL/6J mice. In contrast, in the population of differentially enriched genes, C57BL/6J mice expressed more genes involved in lipid metabolism than BALB/cJ mice. In summary, these results reveal that the two mouse strains used here exhibit several differences in feeding-induced hepatic responses in gene expression, especially in lipogenic genes.

Application of the 3' mRNA-Seq using unique molecular identifiers in highly degraded RNA derived from formalin-fixed, paraffin-embedded tissue.

BACKGROUND: Archival formalin-fixed, paraffin-embedded (FFPE) tissue samples with clinical and histological data are a singularly valuable resource for developing new molecular biomarkers. However, transcriptome analysis remains challenging with standard mRNA-seq methods as FFPE derived-RNA samples are often highly modified and fragmented. The recently developed 3' mRNA-seq method sequences the 3' region of mRNA using unique molecular identifiers (UMI), thus generating gene expression data with minimal PCR bias. In this study, we evaluated the performance of 3' mRNA-Seq using Lexogen QuantSeq 3' mRNA-Seq Library Prep Kit FWD with UMI, comparing with TruSeq Stranded mRNA-Seq and RNA Exome Capture kit. The fresh-frozen (FF) and FFPE tissues yielded nucleotide sizes range from 13 to > 70% of DV200 values; input amounts ranged from 1 ng to 100 ng for validation. RESULTS: The total mapped reads of QuantSeq 3' mRNA-Seq to the reference genome ranged from 99 to 74% across all samples. After PCR bias correction, 3 to 56% of total sequenced reads were retained. QuantSeq 3' mRNA-Seq data showed highly reproducible data across replicates in Universal Human Reference RNA (UHR, R > 0.94) at input amounts from 1 ng to 100 ng, and FF and FFPE paired samples (R = 0.92) at 10 ng. Severely degraded FFPE RNA with ≤30% of DV200 value showed good concordance (R > 0.87) with 100 ng input. A moderate correlation was observed when directly comparing QuantSeq 3' mRNA-Seq data with TruSeq Stranded mRNA-Seq (R = 0.78) and RNA Exome Capture data (R > 0.67). CONCLUSION: In this study, QuantSeq 3' mRNA-Seq with PCR bias correction using UMI is shown to be a suitable method for gene quantification in both FF and FFPE RNAs. 3' mRNA-Seq with UMI may be applied to severely degraded RNA from FFPE tissues generating high-quality sequencing data.

Deploying new generation sequencing for the study of flesh color depletion in Atlantic Salmon (Salmo salar).

BACKGROUND: The flesh pigmentation of farmed Atlantic salmon is formed by accumulation of carotenoids derived from commercial diets. In the salmon gastrointestinal system, the hindgut is considered critical in the processes of carotenoids uptake and metabolism. In Tasmania, flesh color depletion can noticeably affect farmed Atlantic salmon at different levels of severity following extremely hot summers. In this study, RNA sequencing (RNA-Seq) was performed to investigate the reduction in flesh pigmentation. Library preparation is a key step that significantly impacts the effectiveness of RNA sequencing (RNA-Seq) experiments. Besides the commonly used whole transcript RNA-Seq method, the 3' mRNA-Seq method is being applied widely, owing to its reduced cost, enabling more repeats to be sequenced at the expense of lower resolution. Therefore, the output of the Illumina TruSeq kit (whole transcript RNA-Seq) and the Lexogen QuantSeq kit (3' mRNA-Seq) was analyzed to identify genes in the Atlantic salmon hindgut that are differentially expressed (DEGs) between two flesh color phenotypes. RESULTS: In both methods, DEGs between the two color phenotypes were associated with metal ion transport, oxidation-reduction processes, and immune responses. We also found DEGs related to lipid metabolism in the QuantSeq method. In the TruSeq method, a missense mutation was detected in DEGs in different flesh color traits. The number of DEGs found in the TruSeq libraries was much higher than the QuantSeq; however, the trend of DEGs in both library methods was similar and validated by qPCR. CONCLUSIONS: Flesh coloration in Atlantic salmon is related to lipid metabolism in which apolipoproteins, serum albumin and fatty acid-binding protein genes are hypothesized to be linked to the absorption, transport and deposition of carotenoids. Our findings suggest that Grp could inhibit the feeding behavior of low color-banded fish, resulting in the dietary carotenoid shortage. Several SNPs in genes involving in carotenoid-binding cholesterol and oxidative stress were detected in both flesh color phenotypes. Regarding the choice of the library preparation method, the selection criteria depend on the research design and purpose. The 3' mRNA-Seq method is ideal for targeted identification of highly expressed genes, while the whole RNA-Seq method is recommended for identification of unknown genes, enabling the identification of splice variants and trait-associated SNPs, as we have found for duox2 and duoxa1.

Gene expression during larval caste determination and differentiation in intermediately eusocial bumblebees, and a comparative analysis with advanced eusocial honeybees.

The queen-worker caste system of eusocial insects represents a prime example of developmental polyphenism (environmentally-induced phenotypic polymorphism) and is intrinsic to the evolution of advanced eusociality. However, the comparative molecular basis of larval caste determination and subsequent differentiation in the eusocial Hymenoptera remains poorly known. To address this issue within bees, we profiled caste-associated gene expression in female larvae of the intermediately eusocial bumblebee Bombus terrestris. In B. terrestris, female larvae experience a queen-dependent period during which their caste fate as adults is determined followed by a nutrition-sensitive period also potentially affecting caste fate but for which the evidence is weaker. We used mRNA-seq and qRT-PCR validation to isolate genes differentially expressed between each caste pathway in larvae at developmental stages before and after each of these periods. We show that differences in gene expression between caste pathways are small in totipotent larvae, then peak after the queen-dependent period. Relatively few novel (i.e., taxonomically-restricted) genes were differentially expressed between castes, though novel genes were significantly enriched in late-instar larvae in the worker pathway. We compared sets of caste-associated genes in B. terrestris with those reported from the advanced eusocial honeybee, Apis mellifera, and found significant but relatively low levels of overlap of gene lists between the two species. These results suggest both the existence of low numbers of shared toolkit genes and substantial divergence in caste-associated genes between Bombus and the advanced eusocial Apis since their last common eusocial ancestor.

Qi-Long-Tian formula extract alleviates symptoms of acute high-altitude diseases via suppressing the inflammation responses in rat.

BACKGROUND: Chinese Yunnan Province, located in the Yunnan-Guizhou Plateau, is a famous tourist paradise where acute high-altitude illness common occurs among lowland people visitors due to non-acclimatization to the acute hypobaric hypoxia (AHH) conditions. Traditional Chinese medicine, such as Qi-Long-Tian (QLT) formula, has shown effectiveness and safety in the treatment of acute high-altitude diseases. The aim of this study was to clarify the therapeutic mechanisms of this traditional formula using a rat model in a simulated plateau environment. METHODS: Following testing, lung tissue samples were evaluated by hematoxylin-eosin staining and for biochemical characteristics. mRNA-Seq was used to compare differentially expressed genes in control rats, and in rats exposed to AHH and AHH with QLT treatment. RESULTS: Inflammation-related effectors induced following QLT treatment for AHH included MMP9 and TIMP1, and involved several phosphorylation signaling pathways implicated in AHH pathogenesis such as PI3K/AKT and MAPK signaling. CONCLUSION: This study provides insights into the major signaling pathways induced by AHH and in the protective mechanisms involved in QLT formula activity.

Alternative polyadenylation events differ dramatically between Tongcheng and Large White pigs in response to PRRSV infection.

Alternative polyadenylation (APA) is a widespread post-transcriptional regulation mechanism that increases the biological complexity of transcriptome and proteome. However, it is unclear whether APA regulation plays a role in genetic resistance to porcine reproductive and respiratory syndrome virus (PRRSV). Here, we reported genome-wide APA regulation of porcine alveolar macrophages in PRRSV-resistant Tongcheng (TC) pigs and PRRSV-susceptible Large White (LW) pigs upon PRRSV infection. Using 3' mRNA sequencing strategy, we detected 75 981 high-quality APA sites in porcine alveolar macrophages of TC and LW pigs. Furthermore, 1202 and 1089 differentially expressed APA sites, as well as 79 and 117 untranslated region-APA switching genes were identified in TC pigs and LW pigs upon PRRSV infection respectively. The APA events in TC pigs and LW pigs were involved in different biological pathways, while APA events in TC pigs are directly associated with the immune response to PRRSV infection. In addition, we identified genetic variations affecting polyadenylation signal between TC pigs and LW pigs. These findings would provide helpful information on APA regulation for further understanding of genetic resistance to PRRSV.

The Effects of Naringenin on miRNA-mRNA Profiles in HepaRG Cells.

Naringenin, a natural flavonoid widely found in citrus fruits, has been reported to possess anti-oxidant, anti-inflammatory, and hepatoprotective properties as a natural dietary supplement. However, the regulatory mechanism of naringenin in human liver remains unclear. In the present study, messenger RNA sequencing (mRNA-seq), microRNA sequencing (miRNA-seq), and real-time qPCR were used to distinguish the expression differences between control and naringenin-treated HepaRG cells. We obtained 1037 differentially expressed mRNAs and 234 miRNAs. According to the target prediction and integration analysis in silico, we found 20 potential miRNA-mRNA pairs involved in liver metabolism. This study is the first to provide a perspective of miRNA-mRNA interactions in the regulation of naringenin via an integrated analysis of mRNA-seq and miRNA-seq in HepaRG cells, which further characterizes the nutraceutical value of naringenin as a food additive.

Isolation and Characterization of Human Colon Adenocarcinoma Stem-Like Cells Based on the Endogenous Expression of the Stem Markers.

BACKGROUND: Cancer stem cells' (CSCs) self-maintenance is regulated via the pluripotency pathways promoting the most aggressive tumor phenotype. This study aimed to use the activity of these pathways for the CSCs' subpopulation enrichment and separating cells characterized by the OCT4 and SOX2 expression. METHODS: To select and analyze CSCs, we used the SORE6x lentiviral reporter plasmid for viral transduction of colon adenocarcinoma cells. Additionally, we assessed cell chemoresistance, clonogenic, invasive and migratory activity and the data of mRNA-seq and intrinsic disorder predisposition protein analysis (IDPPA). RESULTS: We obtained the line of CSC-like cells selected on the basis of the expression of the OCT4 and SOX2 stem cell factors. The enriched CSC-like subpopulation had increased chemoresistance as well as clonogenic and migration activities. The bioinformatic analysis of mRNA seq data identified the up-regulation of pluripotency, development, drug resistance and phototransduction pathways, and the downregulation of pathways related to proliferation, cell cycle, aging, and differentiation. IDPPA indicated that CSC-like cells are predisposed to increased intrinsic protein disorder. CONCLUSION: The use of the SORE6x reporter construct for CSCs enrichment allows us to obtain CSC-like population that can be used as a model to search for the new prognostic factors and potential therapeutic targets for colon cancer treatment.

Transcriptional profiling of genes in tongue epithelial tissue from immature and adult rats by the RNA-Seq technique.

Children are more sensitive than adults to bitterness and thus dislike bitter tastes more than adults do. However, why children are more sensitive to bitterness has never been revealed. To elucidate the effects of age on taste perception, a double-bottle preference test was first performed with immature and adult rats. Then, RNA-Seq analysis was performed on tongues obtained from rats of the same ages as those in the double-bottle test. The immature rats exhibited a lower consumption rate of bitter solution than the adult rats. Bioinformatics analysis yielded 1,347 differentially expressed genes (DEGs) between male adult rats (MARs, 80 days old) and male immature rats (MIRs, 20 days old) and 380 DEGs between female adult rats (FARs, 80 days old) and female immature rats (FIRs, 20 days old). These DEGs were mainly associated with growth, development, differentiation, and extracellular processes, among other mechanisms. According to Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, the DEGs were enriched for bitter taste transduction. Specifically, the Gnb3 and TRPM5 genes were downregulated in FARs compared with FIRs and in MARs compared with MIRs, and the protein expression of TRPM5 was significantly downregulated in MARs compared with MIRs. The data presented herein suggest that transcriptional regulation of taste-associated signal transduction occurs differently in tongue epithelial tissue of rats at different ages, although additional analyses are needed to confirm this conclusion.