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Differential gene expression between young and senescent, quiescent WI-38 cells.

To investigate age-related changes in gene expression in WI-38 cells, we isolated RNA from young and senescent, quiescent cultures and made subtracted cDNA libraries. Density-arrested cells were incubated in serum-free MCDB-104 for 3 days. RNA was then isolated and subtracted cDNA libraries were made in the phagemid vector pCDM8. Both by picking clones at random from these subtracted libraries and by differential hybridization screening with subtracted cDNA probes from young and senescent cells, we have identified a total of 11 genes for which RNA is expressed differentially in these quiescent young and senescent WI-38 cultures. Two genes, EPC-1 and EPC-A2, with elevated RNA levels in young cells, have sequences which have not previously been identified. Two of the genes with elevated RNA expression in the senescent cells are the mitochondria-coded genes for NADH dehydrogenase subunit 4 and for cytochrome b. We also identified seven other genes with elevated RNA levels in senescent cells. Three of these, LPC-1, LPC-14 and LPC-24, have been partially sequenced and have not previously been identified. These studies show that density-arrested, serum-deprived, quiescent young and senescent cells express a number of genes differentially. These differences are not growth-dependent, but are age-dependent. Our studies also show that the methods employed here, which include careful regulation of the cell cultures and subtraction of the libraries, result in libraries from which differentially expressed genes can be identified, either by random selection or by differential hybridization screening with subtracted probes.

Aging

Differential gene expression study in whole blood identifies candidate genes for psychosis in African American individuals.

Genome-wide association has identified regions of the genome that mediate risk for psychosis. It is possible that variants in these regions confer risk by altering gene expression. This work has predominantly been conducted in individuals of European descent and has focused narrowly on schizophrenia rather than psychosis as a syndrome. In the present study we investigated alterations in gene expression in African American individuals with a range of psychotic diagnoses to increase understanding of the etiology in an underserved population. We performed RNA-seq in whole bloody to survey the transcriptome in 126 patients with a psychosis-spectrum disorder and 217 healthy controls and applied differential gene expression analyses across the genome while controlling for age, sex, population stratification and batch. We found 18 differentially expressed genes (DEGs), some of the locations of the corresponding genes overlap with previously implicated regions for psychosis, but many of which were novel associations. Enrichment analysis of nominally significant genes (p&#xa0;<&#xa0;0.05) revealed overrepresentation of biological processes relating to platelet, immune and cellular function, and sensory perception. Weighted gene co-expression network analysis, applied to identify modules of co-expressed genes associated with psychosis, revealed 10 modules, one of which was significantly associated with psychosis. This module was significantly enriched for DEGs, and for platelet function. These results support the potential role of immune function in the etiology of psychosis, identify novel candidate gene expression phenotypes that correspond to both established and new genomic regions, in individuals of African American ancestry.

Humans

Enterococcus faecalis GP1764 induces an early differential gene expression in the intestine on key pathways related to cellular immune response and gut barrier function in chickens.

The aim of the present study was to elucidate the mode of action of Enterococcus faecalis GP1764 in improving performance traits during the starter phase by analyzing genome-wide gene expression and its interaction with microbial populations in the intestine of chickens challenged with an NSP-rich diet. At day 7, microbiota populations from ileal and cecal contents and transcriptomics from jejunal and cecal mucosa were analyzed between Control (Ctrl) and Enterococcus faecalis GP1764 (EntF) groups. Results from microbiota analysis demonstrated that EntF shifted &#x3b2;-diversity indices in ileum (neutral (p= 0.006) and phylogenetic (p= 0.006)) and caecum (phylogenetic (p= 0.017)). Transcriptomics revealed 43 differentially expressed genes for EntF vs. Ctrl in the jejunal mucosa. Of these, MHCY-36 (MHC-I-Related), RAG2 and MUC19-like genes were upregulated in EntF vs. Ctrl, protein-coding genes with immunomodulatory capacities as supported by GSEA and Cytoscape-ClueGo pathway analyses. Results suggest an intestinal immunomodulation induced through presentation of B vitamins metabolites, synthetized by EntF, to an undescribed subset of innate-like unconventional T lymphocytes in chickens, similar to MAIT cells in mammals. These cells could contribute to antibacterial responses and repair of damaged barrier tissue after inflammatory processes. The upregulation of the MUC19-like gene expression observed in the jejunal mucosa can protect gut integrity via the promotion of mucus production by goblet cells. Finally, RAG2, involved in V(D)J coding segments recombination in B- and T-cells may provide a greater recognition of foreign invaders, allowing the animals to efficiently fight against pathogenic infections. Collectively, these results suggest an important role of EntF in promoting the capacity of animals to rapidly act against pathogenic challenges, herein, inducing resilience towards dietary ingredients with anti-nutritional activity that impart moderate inflammation in chickens.

Enterococcus faecalis

PseudotimeDE-fast: fast testing of differential gene expression along cell pseudotime.

SUMMARY: Identifying differentially expressed (DE) genes along cell pseudotime is crucial for understanding dynamic biological processes captured by single-cell RNA sequencing. However, existing DE methods either produce invalid P-values by ignoring the uncertainty in pseudotime inference or struggle to scale with the growing size of modern datasets. To address these limitations, we introduce PseudotimeDE-fast, a scalable method for detecting DE genes along pseudotime with well-calibrated P-values. Through comprehensive simulations and real-data analyses, we demonstrate that PseudotimeDE-fast delivers comparable or superior performance to existing approaches while offering substantial improvements in computational efficiency. AVAILABILITY AND IMPLEMENTATION: PseudotimeDE-fast is implemented in R with Rcpp acceleration and released under the MIT license. The source code is available at: https://github.com/dsong-lab/PseudotimeDE.

Single-Cell Analysis

The prognostic value and molecular mechanisms of Porphyromonas gingivalis infection-associated differentially expressed genes in oral squamous cell carcinoma.

BACKGROUND: Increasing evidence suggests that Porphyromonas gingivalis (Pg) is associated with oral squamous cell carcinoma (OSCC) development and progression. This study aimed to identify Pg-associated genes with prognostic relevance in OSCC through integrated bioinformatics analysis. METHODS: OSCC-related differentially expressed genes (DEGs) were identified from the The Cancer Genome Atlas (TCGA)-OSCC cohort and intersected with Pg supernatant-associated DEGs from GSE192887. Raw count data were analyzed with DESeq2, whereas transcripts per million (TPM)-transformed expression values were used for downstream visualization and model construction. Weighted gene co-expression network analysis (WGCNA), univariate Cox regression, least absolute shrinkage and selection operator (LASSO) regression, and multivariable Cox modeling were used to develop a seven-gene prognostic signature, which was externally evaluated in GSE41613. Additional analyses examined treatment-associated expression changes in the seven model genes, pairwise correlations among the model genes, and correlations between Pg supernatant-associated differentially expressed gene (PgSDEG)-derived module eigengenes and immune-cell fractions. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was performed in eight paired OSCC and adjacent non-tumor tissues and in supplemented-brain heart infusion (BHI) vehicle-control and Pg culture-supernatant-treated HOK, HSC-3, and CAL-27 cells. RESULTS: A prognostic signature comprising CXCL8, GAST, HBQ1, PADI3, STC1, TEX19, and TMEM92 was established. The signature showed limited-to-moderate discrimination in the TCGA training cohort, with 1-, 3-, and 5-year areas under the curve (AUCs) of 0.68, 0.69, and 0.69, respectively, and limited discrimination in the GSE41613 external cohort (AUCs: 0.66, 0.67, and 0.61). Kaplan-Meier analysis showed poorer survival in the high-risk group in both cohorts. The GSE192887 analysis showed significant treatment-associated expression changes in all seven genes after Pg culture-supernatant exposure. In paired tissues, CXCL8 and TMEM92 were significantly higher in OSCC tissues, whereas STC1 was not significant after Holm correction. In CAL-27 cells, CXCL8, STC1, and TMEM92 increased significantly after culture-supernatant treatment, whereas the corresponding comparisons were not significant in HOK or HSC-3 cells after adjustment. CONCLUSIONS: This study developed a seven-gene Pg-associated prognostic signature for OSCC and provided complementary transcriptomic, immune-correlation, tissue, and cell-based evidence that placed the signature in biological context. The model showed limited-to-moderate discrimination and is not ready for clinical use. The enrichment, gene-correlation, and immune-correlation findings are hypothesis-generating rather than mechanistic evidence. Further independent validation and dedicated functional studies are required.

Oral squamous cell carcinoma (OSCC)

Differential gene expression during germination and after the induction of adventitious bud formation in Norway spruce embryos.

A pulse treatment of embryos of Norway spruce with cytokinin suppresses germinative development and induces the coordinate formation of adventitious buds from subepidermal cell layers. To analyse the patterns of gene expression associated with germination and the alterations induced by the bud induction treatment, we have isolated cDNA clones corresponding to genes that are differentially expressed in cytokinin-treated and untreated in vitro germinating embryos. One category of 14 clones hybridized to transcripts that were abundant specifically during germination. The expression of 8 of these genes was reduced by the bud induction treatment. Four clones, including one identified as a histone H2A gene, recognized transcripts that showed an increased abundance in bud-induced versus in vitro germinating embryos. A second category of 13 clones hybridized to transcripts that increased in abundance during post-germinative development of the seedling. Among these a subset of 8 clones, including an alpha-tubulin clone, corresponds to genes suppressed by the bud induction treatment, whereas 5 clones, including a gene with sequence similarity to polyubiquitin, were unaffected by the treatment. One clone hybridized to a message abundant in the seed, during early germination as well as in the vegetative bud, and showed 60% partial sequence identity to a barley (1----3)-beta-glucanase gene. Genes expressed exclusively in bud-induced or in vitro germinating embryos were not found. The results show that a major difference in gene expression between treated and untreated embryos is related to the shift from extensive cell proliferation to elongation and differentiation that occurs at the transition from germination to post-germinative development, and which is suppressed in the bud-induced embryos.

Amino Acid Sequence

Analysis of differentially expressed genes in schizophrenia based on bioinformatics and corresponding mRNA expression levels.

OBJECTIVE: This study aimed to use bioinformatics analysis to identify differentially expressed genes (DEGs) involved in the pathogenesis of schizophrenia and validate their mRNA expression levels through real-time quantitative PCR (qPCR). MATERIAL/METHODS: Datasets from the publicly available Gene Expression Omnibus (GEO) database were analyzed using R software to identify DEGs. Functional enrichment analyses, including Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, were conducted. A protein-protein interaction (PPI) network was constructed using Cytoscape software to identify key genes with notable expression changes. The expression levels of these key genes were subsequently validated in schizophrenia patients using qPCR to assess potential susceptibility genes. RESULTS: In total, 813 DEGs were identified, with six key genes highlighted through GO analysis and PPI network screening. Among these, HDAC1, UBA52, and FYN demonstrated statistically significant differences in mRNA expression between schizophrenia patients and healthy controls (P&#xa0;<&#xa0;0.05). CONCLUSIONS: This study identified several DEGs potentially linked to the pathogenesis of schizophrenia, suggesting that HDAC1, UBA52, and FYN could serve as candidate susceptibility genes and diagnostic biomarkers. These findings provide new insights and directions for future schizophrenia research.

Humans

Central role for differential gene expression in mammalian hibernation.

Mammalian hibernators experience dramatic reductions in body temperature, metabolic rate, respiratory rate, and heart rate during hibernation. These changes are precisely controlled and reversible with only internally driven mechanisms, suggesting specific biochemical regulation. We present a model that integrates our observations of differential liver gene expression during preparation for, and maintenance of, the hibernating state, with the known phylogenetic interspersion of hibernating species in several major mammalian lineages. This model predicts a major role for the differential expression of existing mammalian genes in the biochemical regulation of hibernation.

Animals

Integrated transcriptomic and metabolomic analysis of fluoride tolerance-related pathways and differentially expressed genes in silkworm strain XSKD.

XueSong KD (XSKD) silkworm strain exhibits prominent fluoride tolerance, yet the underlying molecular mechanisms of fluoride tolerance remains unclear. In the present study, fourth-instar pre-molting XSKD silkworms were used as experimental materials for integrated transcriptomic and untargeted metabolomic analyses. In total, 572 differentially expressed genes and 90 differential metabolites were screened. GO enrichment and KEGG enrichment based on the hypergeometric distribution model revealed that 13-Hydroxy-9Z,11E-octadecadienoic acid (13-(S)-HODE) acts as the core differential metabolite, which is significantly enriched in the linoleic acid metabolism pathway. Within this pathway, LOC101737302 and CYP338A1 display opposite expression trends and show correlations with pathway metabolites. Based on multi-omics data, this study preliminarily characterizes the lipid metabolic response under fluoride stress, providing omics dataset support for further in-depth exploration of the molecular mechanism of fluoride tolerance in silkworms.

Animals

Comprehensive Analysis of Differentially Expressed Genes and Immune Infiltration in Burn Injury: Key Biomarkers and Pathways.

BACKGROUND: Burn injuries trigger complex immune responses and gene expression changes, impacting wound healing and systemic inflammation. Understanding these changes is crucial for identifying biomarkers and therapeutic targets. METHODS: We analyzed two gene expression omnibus datasets (wound tissue [GSE8056] and blood [GSE37069]) to identify differentially expressed genes (DEGs) in burn injury samples versus controls. Immune cell proportions were assessed using CIBERSORT. Functional enrichment analyses (Gene Ontology and Kyoto Encyclopedia of Genes and Genomes) and protein-protein interaction networks were constructed to identify key genes and pathways. RESULTS: We identified 1170 upregulated and 1227 downregulated DEGs. Gene Ontology analysis revealed enrichment in neutrophil activation, inflammatory response, and extracellular matrix organization. Kyoto Encyclopedia of Genes and Genomes analysis highlighted cytokine-cytokine receptor interaction, TNF, and IL-17 signaling pathways. Immune infiltration analysis showed significant changes in neutrophils, macrophages (M1/M2), and T-cell subsets. Protein-protein interaction network analysis identified five hub genes: JUN, STAT1, Bcl2, MMP9, and TLR2. CONCLUSIONS: This study provides a comprehensive bioinformatic analysis of gene expression and immune responses in burn injuries. The identified DEGs, hub genes, and pathways offer insights into the immune response mechanisms and suggest potential targets for diagnostic and therapeutic interventions in burn injury management.

Burns

Differential gene expression in wild-type and X-ray-sensitive mutants of Chinese hamster ovary cell lines.

Complementary DNA cloning, differential screening and Northern hybridization techniques were used to study differential gene expression in the wild-type Chinese hamster ovary (CHO) K1 cell line and its two X-ray sensitive mutants, xrs-5 and xrs-6. 11 species of mRNAs were found underexpressed in the two independently isolated mutants. The steady-state levels of those mRNAs are 3-26-fold less in the two mutants, depending on the particular species. 6 of the underexpressed mRNAs have been identified by comparing the sequences of the cloned cDNAs to the known sequences in GenBank. 4 of them code for the structural proteins of ferritin heavy chain, nonmuscle myosin light chain 3nm, ribosomal protein S17 and L7, respectively. The other two have strong homology with mouse B2 or retroviral sequences. The remaining 5 mRNAs did not show significant homology with any of the known sequences and apparently represent newly isolated species. The effect of 137Cs gamma-rays on the expression of the 11 mRNAs has been studied. Radiation inhibited the expression of the B2-like gene in the mutants but not in the wild-type CHO cells. The levels of the other 10 mRNAs were not affected by radiation. The underexpression of this group of genes in both xrs-5 and xrs-6 mutants seems to be related to their radiation-sensitive phenotype, although the specific gene responsible has not been identified. Two models are proposed to explain the mechanism of underexpression. It is suggested that a cellular factor or/and chromosome structural changes are involved.

Amino Acid Sequence

PoweREST: Statistical Power Estimation for Spatial Transcriptomics Experiments to Detect Differentially Expressed Genes Between Two Conditions.

Recent advancements in Spatial Transcriptomics (ST) have significantly enhanced biological research in various domains. However, the high cost of current ST data generation techniques restricts its application in large-scale population studies. Consequently, there is a pressing need to maximize the use of available resources to achieve robust statistical power. One fundamental question in ST analysis is to detect differentially expressed genes (DEGs) among different conditions using ST data. Such DEG analysis is often performed but the associated power calculation is rarely discussed in the literature. To address this gap, we introduce, PoweREST (https://github.com/lanshui98/PoweREST), a power estimation tool designed to support power calculation of DEG detection with 10X Genomics Visium data. PoweREST enables power estimation both before any ST experiments or after preliminary data are collected, making it suitable for a wide variety of power analyses in ST studies. We also provide a user-friendly, program-free web application (https://lanshui.shinyapps.io/PoweREST/), allowing users to interactively calculate and visualize the study power along with relevant the parameters.

Differentially expressed genes

Differential gene expression in multilocus isozyme systmes of the developing green sunfish.

The patterns of expression of eight multilocous isozyme systems were investigated in the differentiated adult tissues and the early embryonic stages (0-210 hours after fertilization) of the green sunfish, Lepomis cyanellus. Enzymes encoded by approximately 23 gene loci were resolved by starch-gel electrophoresis and detected by specific histochemical staining. The developmental patterns of these isozyme systems appear to be the result of the diffential expression of the multiple gene loci. Isozymic forms of glucoseophosphate isomerase (GPI-A2), malate dehydrogenase (MDH-A2), and creatine kinase (CK-C2) were present in most differentiated tissues, in the unfertilized eggs, and in all stages of embryonic development. Closely homologous forms of these isozymes (GPI-B2, MDH-B2, and CK-A2) were expressed predominantly in skeletal muscle and were first detected at around the time of hatching (38-42 hours). The similar temporal and spatial patterns of gene expressions for the GPI, LDH, MDH, and CK loci suggest that the duplicates loci encoding enzymes, diverged in their regulation to patterns of differential gene expression which are similar for each enzyme system.

Adenylate Kinase

PoweREST: Statistical power estimation for spatial transcriptomics experiments to detect differentially expressed genes between two conditions.

Recent advancements in spatial transcriptomics (ST) have significantly enhanced biological research in various domains. However, the high cost for current ST data generation techniques restricts the large-scale application of ST. Consequently, maximization of the use of available resources to achieve robust statistical power for ST data is a pressing need. One fundamental question in ST analysis is detection of differentially expressed genes (DEGs) under different conditions using ST data. Such DEG analyses are performed frequently, but their power calculations are rarely discussed in the literature. To address this gap, we developed PoweREST, a power estimation tool designed to support the power calculation for DEG detection with 10X Genomics Visium data. PoweREST enables power estimation both before any ST experiments and after preliminary data are collected, making it suitable for a wide variety of power analyses in ST studies. We also provide a user-friendly, program-free web application that allows users to interactively calculate and visualize study power along with relevant parameters.

Gene Expression Profiling

Identification of genes differentially expressed in two types of v-myb-transformed avian myelomonocytic cells.

In an earlier study we found that different forms of the v-myb oncogene transform myeloid cells which resemble either monoblasts [when v-myb of avian myeloblastosis virus (AMV) was used] or promyelocytes [when a point mutant in v-myb of AMV was used; Introna, M., Golay, J., Frampton J., Nakano, T., Ness, S.A. & Graf, T. (1990). Cell, 63, 1287-1297]. In the present study we have searched for genes expressed in AMV mutant-transformed promyelocytes that are not expressed in AMV-transformed monoblasts using a differential screening approach. Eight different genes were identified among more than 500 differentially expressed clones. The most abundant of these was the previously identified myb-regulated mim-1 gene. The others were found to encode a small calcium-binding (MRP-like) protein; the p20K protein; goose-type lysozyme; a ribonuclease A/angiogenin-related protein; and three non-identified proteins. Although these genes appear to be rather lineage restricted, their expression varied in different subtypes of transformed myelomonocytic cells, and only two of them (goose lysozyme and ribonuclease) showed a similar expression pattern in normal promyelocytes and macrophages, suggesting an aberrant gene regulation in the transformed cells. Co-transfection experiments of a reporter construct containing the promoter of the ribonuclease A-related gene indicated that this promoter is regulated by the v-Myb oncoprotein without the involvement of Myb-specific binding sequences.

Amino Acid Sequence

Analysis of differentially expressed genes in retinitis pigmentosa retinas. Altered expression of clusterin mRNA.

The molecular and cellular processes underlying photoreceptor degeneration in retinitis pigmentosa (RP) are unknown. We have investigated gene expression in diseased retinas using differential hybridization screening of a retinal cDNA library with probes derived from normal and RP retinal RNA. Most differential clones detected corresponded to transcripts absent from the dystrophic state, including e.g. opsin. However, one clone was noticeably increased in RP in comparison with the control: partial sequencing showed it encoded clusterin. Increased expression of clusterin has been identified in several cases of tissues undergoing apoptosis (programmed cell death), and our finding suggests that the degenerative changes in advanced RP may represent another example of apoptosis, possibly with common causative mechanisms.

Adult

Hormonal control of gene expression: differential activation of rat bone marrow RNA polymerases by erythropoietin and testosterone.

Hormones play a role in the regulation of gene expression by inducing changes in enzyme patterns in target cells mediated by the synthesis of specific RNA molecules. Erythropoiesis has been used as a system for studying the molecular mechanism of regulation of gene action by means of two hormones: erythropoietin and testosterone. Experiments designed to correlate the biochemical action of both hormones on rat marrow cells are herein reported. Both factors seems to act at different biochemical and citological levels. Erythropoietin triggers the erythropoietic process acting on the erythropoietin sensitive cells (ESC), in which the hormone induces the synthesis of a high molecular weight RNA, which is the precursor of a functional 9 S messenger RNA. Testosterone seems to act on polychromatophilic erythroblasts, in which the synthesis of ribosomal RNA or its precursor is stimulated. The steroid enhances the nuclear ribonuclease activity, which could represent a control mechanism for the processing (maturation) of high molecular weight RNAs. The incorporation of 3H-GTP and 3H-UTP into RNA by isolated rat bone marrow nuclei is stimulated by erythropoietin and testosterone. Using alpha-amanitine and different ionic strength conditions it was found that erythropoietin enhances preferentially RNA polymerase II activity while testosterone increases RNA polymerase I activity. It is postulated that erythropoietin and testosterone act synergically to create the biochemical machinery for hemoglobin synthesis, the macromolecule that characterizes the erythropoietic process.

Amanitins

Isolation of cDNA clones corresponding to genes differentially expressed in two colon-carcinoma cell lines differing by their tumorigenicity.

In an effort to isolate genes involved in the progression of colonic cells leading to a carcinoma, we used as a model 2 rat colon-carcinoma cell lines selected from the same tumor, differing by their tumorigenicity. When soluble, Triton-X-100 extracted, or cytoskeletal proteins from the progressive PROb cells and the regressive REGb cells were analyzed by SDS-PAGE, minor differences were seen. Furthermore, mRNA-cDNA hybridization analyses showed extensive homology between the 2 mRNA populations. Thus, the homology between the 2 clones is high at both the protein and the mRNA levels. A PROb cDNA library was hybridized with 32P-cDNA synthesized from PROb or REGb mRNA. The clones giving a stronger signal when hybridized with the homologous PROb probe were isolated. The specificity of each clone was confirmed by RNA blotting. Most of the positive clones showed a 2- to 3-fold higher expression in PROb cells when compared with REGb cells. One clone (J 13) corresponded to an mRNA 7- to 10-fold more abundant in PROb cells, and was further studied. No gene amplification was detected by Southern blot analysis, indicating that the difference in mRNA content was most likely due to an increased transcription of this gene in PROb cells. Sequencing of the cDNA showed high homology with the rat ferritin light sub-unit. Over-expression of ferritin in PROb cells as compared with REGb cells was confirmed at the protein level using specific antibodies.

Adenocarcinoma