Up-regulated Differentially Expressed Genes Research Articles

In vitro induction of tetraploids and their phenotypic and transcriptome analysis in Glehnia littoralis

BackgroundGlehnia littoralis is a medicinal and edible plant species having commercial value and has several hundred years of cultivation history. Polyploid breeding is one of the most important and fastest ways to generate novel varieties. To obtain tetraploids of G. littoralis in vitro, colchicine treatment was given to the seeds and then were screened based on morphology, flow cytometry, and root tip pressing assays. Furthermore, transcriptome analysis was performed to identity the differentially expressed genes associated with phenotypic changes in tetraploid G. littoralis.ResultsThe results showed that 0.05% (w/v) colchicine treatment for 48 h was effective in inducing tetraploids in G. littoralis. The tetraploid G. littoralis (2n = 4x = 44) was superior in leaf area, leaf thickness, petiole diameter, SPAD value (Chl SPAD), stomatal size, epidermal tissues thickness, palisade tissues thickness, and spongy tissues thickness to the diploid ones, while the stomatal density of tetraploids was significantly lower. Transcriptome sequencing revealed, a total of 1336 differentially expressed genes (DEGs) between tetraploids and diploids. Chromosome doubling may lead to DNA content change and gene dosage effect, which directly affects changes in quantitative traits, with changes such as increased chlorophyll content, larger stomata and thicker tissue of leaves. Several up-regulated DEGs were found related to growth and development in tetraploid G. littoralis such as CKI, PPDK, hisD and MDP1. KEGG pathway enrichment analyses showed that most of DEGs were enriched in metabolic pathways.ConclusionsThis is the first report of the successful induction of tetraploids in G. littoralis. The information presented in this study facilitate breeding programs and molecular breeding of G. littoralis varieties.

Pathogenicity and transcriptomic profiling reveals immunology molecular hallmarks after CA10 virus infection.

Hand, foot and mouth disease (HFMD) is a common infectious disease caused by viral infection by a variety of enteroviruses, with coxsackievirus A 10 (CA10) having become more prevalent in recent years. In this study, models of CA10 infection were established in 7-day-old Institute of Cancer Research (ICR) mice by intraperitoneal injection to analyze the pathogenicity of the virus. RNA sequencing analysis was used to screen the differentially expressed genes (DEGs) after CA10 infection. Coxsackievirus A 16 (CA16) and enterovirus 71 (EV71) infections were also compared with CA10. After CA10 virus infection, the mice showed paralysis of the hind limbs at 3 days post infection and weight loss at 5 days post infection. We observed viral replication in various tissues and severe inflammatory cell infiltration in skeletal muscle. The RNA-sequencing analysis showed that the DEGs in blood, muscle, thymus and spleen showed heterogeneity after CA10 infection and the most up-regulated DEGs in muscle were enriched in immune-related pathways. Compared with CA16 and EV71 infection, CA10 may have an inhibitory effect on T helper (Th) cell differentiation and cell growth. Additionally, the common DEGs in the three viruses were most enriched in the immune system response, including the Toll-like receptor pathway and the nucleotide-binding and oligomerization domain (NOD)-like pathway. Our findings revealed a group of genes that coordinate in response to CA10 infection, which increases our understanding of the pathological mechanism of HFMD.

Cardiac transcriptomic changes induced by early CKD in mice reveal novel pathways involved in the pathogenesis of Cardiorenal syndrome type 4

BackgroundCardiorenal syndrome (CRS) type 4 is prevalent among the chronic kidney disease (CKD) population, with many patients dying from cardiovascular complications. However, limited data regarding cardiac transcriptional changes induced early by CKD is available. MethodsWe used a murine unilateral ureteral obstruction (UUO) model to evaluate renal damage, cardiac remodeling, and transcriptional regulation at 21 days post-surgery through histological analysis, RT-qPCR, RNA-seq, and bioinformatics. ResultsUUO leads to significant kidney injury, low uremia, and pathological cardiac remodeling, evidenced by increased collagen deposition and smooth muscle alpha-actin 2 expression. RNA-seq analysis identified 76 differentially expressed genes (DEGs) in UUO hearts. Upregulated DEGs were significantly enriched in cell cycle and cell division pathways, immune responses, cardiac repair, inflammation, proliferation, oxidative stress, and apoptosis. Gene Set Enrichment Analysis further revealed mitochondrial oxidative bioenergetic pathways, autophagy, and peroxisomal pathways are downregulated in UUO hearts. Vimentin was also identified as an UUO-upregulated transcript. ConclusionsOur results emphasize the relevance of extensive transcriptional changes, mitochondrial dysfunction, homeostasis deregulation, fatty-acid metabolism alterations, and vimentin upregulation in CRS type 4 development.

P148 Longitudinal single-cell data informs deterministic modelling of inflammatory bowel disease

Abstract Background Crucial contributions to our understanding of the inflammatory process in inflammatory bowel disease (IBD) have been made using single cell level approaches. Extracting mechanistic details of cell and gene dynamics would be critical to understanding the mechanisms mediating mucosal healing and predicting response to specific therapies. However, this remains challenging. Therefore, our aim was to use ordinary differential equations (ODEs) for mathematical modelling of cell abundance and prediction of biological processes in IBD. Methods We used mathematical modelling of published longitudinal single-cell mRNA sequencing (scRNA-seq) data from DSS colitis in mice to simulate cellular dynamics during IBD. We validated our predictions using pseudo-bulk gene expression analysis and flow cytometry. We also analysed a published microarray dataset from colonic biopsies of 51 IBD patients prior to infliximab treatment. Results Based on the scRNA-seq data, we used our mathematical model to predict the immune dynamics of a different mouse colitis protocol. We validated these predictions experimentally using flow cytometry and found good agreement between model simulation and experimental data (predicted vs. observed, Pearson's rho=0.92). We found that the estimated turnover rates predicted the protein level of the proliferation marker KI-67 (rho=0.88) in DSS colitis. Finally, we wanted to investigate whether our mathematical model reflected processes in human IBD. We reasoned that genes correlated with epithelial cell dynamics might be indicative of mucosal healing after DSS retrieval. We found more highly correlated genes when using simulated epithelial abundance compared to experimentally measured epithelial abundance. Many genes associated with tight junctions (Cldn8, Cftr, Crb3, Jam2) were among the highly positively correlated genes and genes associated with inflammation (Il1b, Jak2, Il10ra) were among the negatively correlated genes with epithelial cell abundance. We then looked for differentially expressed (DE) genes in pre-treatment colon biopsies between responders and non-responders to infliximab. We found a significant overlap of 25 positively correlated genes with upregulated DE genes in responders, i.e. associated with tight junctions, and 73 negatively correlated genes with upregulated genes in non-responders, i.e. associated with inflammatory pathways. Conclusion We demonstrate that mathematical modelling of colonic cell dynamics provides insight into IBD progression and treatment response. Furthermore, our results suggest that pre-treatment barrier integrity may predict and influence response to infliximab treatment.

Transcriptomic characterization of the human segmental endotoxin challenge model

Segmental instillation of lipopolysaccharide (LPS) by bronchoscopy safely induces transient airway inflammation in human lungs. This model enables investigation of pulmonary inflammatory mechanisms as well as pharmacodynamic analysis of investigational drugs. The aim of this work was to describe the transcriptomic profile of human segmental LPS challenge with contextualization to major respiratory diseases. Pre-challenge bronchoalveolar lavage (BAL) fluid and biopsies were sampled from 28 smoking, healthy participants, followed by segmental instillation of LPS and saline as control. Twenty-four hours post instillation, BAL and biopsies were collected from challenged lung segments. Total RNA of cells from BAL and biopsy samples were sequenced and analysed for differentially expressed genes (DEGs). After challenge with LPS compared with saline, 6316 DEGs were upregulated and 241 were downregulated in BAL, but only one DEG was downregulated in biopsy samples. Upregulated DEGs in BAL were related to molecular functions such as “Inflammatory response” or “chemokine receptor activity”, and upregulated pro-inflammatory pathways such as “Wnt-"/“Ras-"/“JAK-STAT” “-signaling pathway”. Furthermore, the segmental LPS challenge model resembled aspects of the five most prevalent respiratory diseases chronic obstructive pulmonary disease (COPD), asthma, pneumonia, tuberculosis and lung cancer and featured similarities with acute exacerbations in COPD (AECOPD) and community-acquired pneumonia. Overall, our study provides extensive information about the transcriptomic profile from BAL cells and mucosal biopsies following LPS challenge in healthy smokers. It expands the knowledge about the LPS challenge model providing potential overlap with respiratory diseases in general and infection-triggered respiratory insults such as AECOPD in particular.

Transcriptomics analysis reveals distinct mechanism of breast cancer stem cells regulation in mammospheres from MCF-7 and T47D cells

Luminal A breast cancer, constituting 70 % of breast cancer cases, presents a challenge due to the development of resistance and recurrence caused by breast cancer stem cells (BCSC). Luminal breast tumors are characterized by TP53 expression, a tumor suppressor gene involved in maintaining stem cell attributes in cancer. Although a previous study successfully developed mammospheres (MS) from MCF-7 (with wild-type TP53) and T47D (with mutant TP53) luminal breast cancer cells for BCSC enrichment, their transcriptomic profiles remain unclear. We aimed to elucidate the transcriptomic disparities between MS of MCF-7 and T47D cells using bioinformatics analyses of differentially expressed genes (DEGs), including the KEGG pathway, Gene Ontology (GO), drug-gene association, disease-gene association, Gene Set Enrichment Analysis (GSEA), DNA methylation analysis, correlation analysis of DEGs with immune cell infiltration, and association analysis of genes and small-molecule compounds via the Connectivity Map (CMap). Upregulated DEGs were enriched in metabolism-related KEGG pathways, whereas downregulated DEGs were enriched in the MAPK signaling pathway. Drug-gene association analysis revealed that both upregulated and downregulated DEGs were associated with fostamatinib. The KEGG pathway GSEA results indicated that the DEGs were enriched for oxidative phosphorylation, whereas the downregulated DEGs were negatively enriched for the p53 signaling pathway. Examination of DNA methylation revealed a noticeable disparity in the expression patterns of the PKM2, ERO1L, SLC6A6, EPAS1, APLP2, RPL10L, and NEDD4 genes when comparing cohorts with low- and high-risk breast cancer. Furthermore, a significant positive correlation was identified between SLC6A6 expression and macrophage presence, as well as MSN, and AKR1B1 expression and neutrophil and dentritic cell infiltration. CMap analysis unveiled SA-83851 as a potential candidate to counteract the effects of DEGs, specifically in cells harbouring mutant TP53. Further research, including in vitro and in vivo validations, is warranted to develop drugs targeting BCSCs.

Different Transcriptome Features of Peripheral Blood Mononuclear Cells in Non-Emphysematous Chronic Obstructive Pulmonary Disease.

Non-emphysematous chronic obstructive pulmonary disease (COPD), which is defined based on chest computed tomography findings, presented different transcriptome features of peripheral blood mononuclear cells (PBMCs) compared with emphysematous COPD. Enrichment analysis of transcriptomic data in COPD demonstrated that the "Hematopoietic cell lineage" pathway in Kyoto Encyclopedia of Genes and Genomes pathway analysis was highly upregulated, suggesting that cellular dynamic dysregulation in COPD lungs is affected by pathologically modified PBMCs. The differentially expressed genes (DEGs) upregulated in PBMCs reflected the disease state of non-emphysematous COPD. Upregulated DEGs such as XCL1, PRKCZ, TMEM102, CD200R1, and AQP1 activate T lymphocytes and eosinophils. Upregulating keratan sulfate biosynthesis and metabolic processes is associated with protection against the destruction of the distal airways. ITGA3 upregulation augments interactions with extracellular matrix proteins, and COL6A1 augments the profibrotic mast cell phenotype during alveolar collagen VI deposition. Upregulating HSPG2, PDGFRB, and PAK4 contributes to the thickening of the airway wall, and upregulating SERPINF1 expression explains the better-preserved vascular bed. Therefore, gene expression and pathway analysis in PBMCs in patients with non-emphysematous COPD represented type 2 immune responses and airway remodeling features. Therefore, these patients have asthmatic potential despite no clinical signs of asthma, in contrast to those with emphysematous COPD.

Identification of novel candidate biomarkers related to immune cell infiltration in peri‐implantitis

AbstractObjectiveThe present study was performed to identify key biomarkers associated with immune cell infiltration in peri‐implantitis through bioinformatic analyses.MethodsSix peri‐implantitis soft tissue samples and six healthy gingiva samples were obtained from GSE106090, and were used to identify immune‐associated differentially expressed genes (DEGs) in peri‐implantitis. The candidate biomarkers associated with immune cell infiltration were examined by immunohistochemical staining.ResultsWe identified 2089 upregulated and 2173 downregulated genes. Upregulated DEGs were significantly associated with immune response. Ten key candidate biomarkers were identified in the PPI network, including IL1B, TLR2, TLR4, CCL4, CXCL8, IL10, IL6, CD4, CCL3, and PTPRC. The expression level of the 10 genes increased in peri‐implantitis soft tissue samples compared with healthy gingiva samples. The proportion of CD4+ T cells, iTreg, and Tfh in infiltration immune cells increased in peri‐implantitis soft tissue samples and were positively correlated with the expression level of candidate biomarkers TLR4, CCL3, CXCL8, and IL1B. Immunohistochemistry showed that there were more lymphocytes in peri‐implantitis soft tissue samples, with an increased expression level of TLR4, CCL3, CXCL8, and IL1B.ConclusionIdentification of four novel diagnostic biomarkers was helpful for revealing the molecular mechanisms and could serve as a risk predictor for the immune microenvironment in peri‐implantitis.

CD39+CD55- Fb Subset Exhibits Myofibroblast-Like Phenotype and Is Associated with Pain in Osteoarthritis of the Knee.

Recent studies utilizing single-cell analysis have unveiled the presence of various fibroblast (Fb) subsets within the synovium under inflammatory conditions in osteoarthritis (OA), distinguishing them from those in rheumatoid arthritis (RA). Moreover, it has been reported that pain in knee OA patients is linked to specific fibroblast subsets. Single-cell expression profiling methods offer an incredibly detailed view of the molecular states of individual cells. However, one limitation of these methods is that they require the destruction of cells during the analysis process, rendering it impossible to directly assess cell function. In our study, we employ flow cytometric analysis, utilizing cell surface markers CD39 and CD55, in an attempt to isolate fibroblast subsets and investigate their relationship with OA pathology. Synovial tissues were obtained from 25 knee OA (KOA) patients. Of these, six samples were analyzed by RNA-seq (n = 3) and LC/MS analysis (n = 3). All 25 samples were analyzed to estimate the proportion of Fb (CD45-CD31-CD90+) subset by flow cytometry. The proportion of Fb subsets (CD39+CD55- and CD39-CD55+) and their association with osteoarthritis pathology were evaluated. CD39+CD55- Fb highly expressed myogenic markers such as CNN1, IGFBP7, MYH11, and TPM1 compared to CD39-CD55+ Fb. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of upregulated differentially expressed genes (DEGs) in CD39+CD55- Fb identified the Apelin pathway and cGMP-PKC-signaling pathway as possibly contributing to pain. LC/MS analysis indicated that proteins encoded by myogenic marker genes, including CNN1, IGFBP7, and MYH11, were also significantly higher than in CD39-CD55+ Fb. CD39-CD55+ Fb highly expressed PRG4 genes and proteins. Upregulated DEGs were enriched for pathways associated with proinflammatory states ('RA', 'TNF signaling pathway', 'IL-17 signaling pathway'). The proportion of CD39+CD55- Fb in synovium significantly correlated with both resting and active pain levels in knee OA (KOA) patients (resting pain, ρ = 0.513, p = 0.009; active pain, ρ = 0.483, p = 0.015). There was no correlation between joint space width (JSW) and the proportion of CD39+CD55- Fb. In contrast, there was no correlation between the proportion of CD39-CD55+ Fb and resting pain, active pain, or JSW. In conclusion, CD39+CD55- cells exhibit a myofibroblast phenotype, and its proportion is associated with KOA pain. Our study sheds light on the potential significance of CD39+CD55- synovial fibroblasts in osteoarthritis, their myofibroblast-like phenotype, and their association with joint pain. These findings provide a foundation for further research into the mechanisms underlying fibrosis, the impact of altered gene expression on osteoarthritic joints, and potential therapeutic strategies.

Mechanisms of ag85a/b DNA vaccine conferred immunotherapy and recovery from Mycobacterium tuberculosis-induced injury.

Our previous research developed a novel tuberculosis (TB) DNA vaccine ag85a/b that showed a significant therapeutic effect on the mouse tuberculosis model by intramuscular injection (IM) and electroporation (EP). However, the action mechanisms between these two vaccine immunization methods remain unclear. In a previous study, 96 Mycobacterium tuberculosis (MTB) H37 Rv-infected BALB/c mice were treated with phosphate-buffered saline, 10, 50, 100, and 200 μg ag85a/b DNA vaccine delivered by IM and EP three times at 2-week intervals, respectively. In this study, peripheral blood mononuclear cells (PBMCs) from three mice in each group were isolated to extract total RNA. The gene expression profiles were analyzed using gene microarray technology to obtain differentially expressed (DE) genes. Finally, DE genes were validated by real-time reverse transcription-quantitive polymerase chain reaction and the GEO database. After MTB infection, most of the upregulated DE genes were related to the digestion and absorption of nutrients or neuroendocrine (such as Iapp, Scg2, Chga, Amy2a5), and most of the downregulated DE genes were related to cellular structural and functional proteins, especially the structure and function proteins of the alveolar epithelial cell (such as Sftpc, Sftpd, Pdpn). Most of the abnormally upregulated or downregulated DE genes in the TB model group were recovered in the 100 and 200 μg ag85a/b DNA IM groups and four DNA EP groups. The pancreatic secretion pathway downregulated and the Rap1 signal pathway upregulated had particularly significant changes during the immunotherapy of the ag85a/b DNA vaccine on the mouse TB model. The action targets and mechanisms of IM and EP are highly consistent. Tuberculosis infection causes rapid catabolism and slow anabolism in mice. For the first time, we found that the effective dose of the ag85a/b DNA vaccine immunized whether by IM or EP could significantly up-regulate immune-related pathways and recover the metabolic disorder and the injury caused by MTB.

Transcriptome analysis identifies differentially expressed genes involved in lignin biosynthesis in barley

Lignin is an essential metabolite for plant growth but negatively affects the quality of forage barley. Genetic modification of quality traits to improve the forage digestibility requires an understanding of the molecular mechanism of lignin biosynthesis. RNA-Seq was used to quantify transcripts differentially expressed among leaf, stem and spike tissues from two barley genotypes. A total of 13,172 differentially expressed genes (DEGs) were identified, of which much more up-regulated DEGs were detected from the contrasting groups of leaf vs spike (L-S) and stem vs spike (S-S), and down-regulated DEGs were dominant in the group of stem vs leaf (S-L). 47 DEGs were successfully annotated to the monolignol pathway and six of them were candidate genes regulating the lignin biosynthesis. The qRT-PCR assay verified the expression profiles of the six candidate genes. Among them, four genes might positively regulate the lignin biosynthesis during forage barley development in terms of the consistency of their expression levels and changes of lignin content among the tissues, while the other two genes may have the reverse effects. These findings provide target genes for further investigations on molecular regulatory mechanisms of lignin biosynthesis and genetic resources for improvement of forage quality in barley molecular breeding programme.

Abstract C044: A case and control genetic profile of tissue and serum in African American men

Abstract Prostate cancer is the most common non-cutaneous cancer among men. A man with 1,2 or 3 first degree relatives with prostate cancer, has a 2,5, and 11-fold increased risk of developing prostate cancer. The heritability rate of prostate cancer is 58%. African American men have the highest incidence and mortality rate of prostate cancer in American. Men of African descent, globally, are more likely to die from prostate cancer than any other ancestral groups. We performed Immunohistochemistry(IHC) and RNA seq analysis on African American tissue and serum samples, with a focus on HMGA2, which has been shown promote EMT, invasion, and metastasis in cancer. Acknowledgements: These studies were supported by NIH/NIMHD U54MD007590 and U54MD013376. Our sample set included 3 benign and 2 prostate cancer tumors from African Americans. IHC markers showed HMGA2 staining within epithelial cells of the prostate cancer tissue. This result validates studies that show that distinct subtypes of prostate cancer may arise from luminal and basal epithelial cell types. RNA seq results showed HMGA to be non-significantly down-regulated in tumor samples. The most differentially expressed (DE) gene, however, was SNORD116-18. Studies have shown SNORD116-18 expression to be associated with chronic lymphocytic leukemia in distinguishing prognostic groups. This finding may be a pathway of interest for further studies. RNA seq of serum gave homogeneous results, however, gene set enrichment analysis of the most differentially expressed genes within the serum showed overlap with inflammation pathways gene for upregulated DE genes and Methylation Pathway for downregulated DE genes. Further analysis on more samples from African Americans tissue and serum will be used to validate findings. Citation Format: Maxine S. Harlemon, Roni Bollag, Martha Terris, Ana Cecilia Millena, Nathan Bowen, Valerie Odero-Marah. A case and control genetic profile of tissue and serum in African American men [abstract]. In: Proceedings of the 15th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2022 Sep 16-19; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2022;31(1 Suppl):Abstract nr C044.

Transcriptome response of pubertal male Clarias macrocephalus to dietary 17α-methyltestosterone

We studied the mechanism of the effects of 17α-methyltestosterone (MT) on previously reported enhanced growth and maturation of the Asian catfish Clarias macrocephalus. A total of 18 pubertal male (3 fish·tank-1) catfish (ABW=125.97 g) were stocked into six experimental circular tanks and fed with a basal diet (control) or a basal diet containing 60 mg MT kg-1 diet for 90 days. At the termination of the experiment, livers were excised, and total RNA was extracted, evaluated, and reverse transcribed to cDNA. Six libraries of cDNA (3 for the control and 3 for the MT-treated group) were subjected to bioinformatics. Differentially expressed genes (DEGs) were identified using similarity scores of the de novo assembled transcript with those in the seven public databases. There were 141 up- and 72 down-regulated DEGs, mostly belonging to uncharacterized/not yet identified proteins. Thus, DEGs in the top enriched KEGG pathways were used to characterize the effects of MT. Upregulated DEGs included tyrosine hydroxylase (TH), Cytoplasmic Polyadenylation Element Binding (CPEB), Fas-associated death domain (FADD), Major Histocompatibility Complex class 1 (MHC-1), other immune-related genes, among others. Down-regulated DEGs included Cytochrome P450 family one subfamily A member 1 (CYP1A1), Alternative oxidase (AOX), UDP-glycosyltransferase (UGT), Acyl-CoA Synthase Bubblegum Family Member 1 (ACSBG), Stearoyl-CoA Desaturase (SCD), among others. In conclusion, MT-treated male Clarias macrocephalus exhibited up-regulation of some pathways protecting cellular conditions like cell proliferation, survival, development and homeostasis processes, for development and homeostasis processes. MT also affected changes in bile acid production and the inhibition of the production/conversion of testosterone. MT resulted in the down-regulation of a serotonergic system that possibly affected gonadal development, inhibition of the retinoid enzyme that would otherwise diminish the full effects of MT or its metabolite, and inhibition of the formation of estrogen, leading to down-regulation of several estrogen-related KEGG pathways.

Upregulated mitosis-associated genes CENPE, CENPF, and DLGAP5 predict poor prognosis and chemotherapy resistance of Acute Myeloid Leukemia.

Mitosis-associated genes are dysregulated in many types of cancers and play important roles in disease progression and chemotherapy resistance. However, their expression and functions in chemotherapy-resistant Acute Myeloid Leukemia (AML) are still largely undetermined. This study aims to explore the roles of spindle assembly checkpoint (SAC) genes CENPE, CENPF, and DLGAP5 in chemotherapy-resistant AML. RNA-sequencing (RNA-seq) was performed in patients with chemotherapy-resistant AML and chemotherapy-sensitive AML. AML mRNA data from 151 patients with recurrence were downloaded from TCGA. Integrated analysis of the differentially expressed genes (DEGs), GO and KEGG pathways. CENPE, CENPF, or DLGAP5 knockdown cell lines were used to analyse proliferation, apoptosis and cell cycle alterations. A total of 87 DEGs (48 upregulated and 39 downregulated) were obtained through gene analysis of R/R-AML and a total of 329 DEGs (202 upregulated and 127 downregulated) were obtained in refractory S-AML. Upregulated DEGs were mainly enriched in cell cycle (GO: 0007049, hsa04110) and mitotic cell cycle (GO: 0000278) processes and pathway. Venn diagram analysis identified the most upregulated DEGs (including CENPE, CENPF, and DLGAP5) in chemoresistant AML. The expression of CENPE, CENPF and DLGAP5 in R-AML (TCGA) was significantly higher than that of primary AML (GEO). The proliferation of K562 cells after CENPE and DLGAP5 knockdown was significantly decreased (P= 0.0001 and P= 0.0006). In THP-1 cells, the CCK-8 values after CENPE, CENPF and DLGAP5 knockdown were significantly decreased (P= 0.01, P= 0.0395 and P= 0.0362). Knockdown of CENPE, CENPF and DLGAP5 significantly increased cell apoptosis by regulating Caspase-9, BAX, TP-53 and bcl-2, and induced cell cycle arrested by regulating CDK1, CDK2, CDKN1A, and CyclinD1. In conclusion, the mitotic cell cycle-associated genes CENPE, CENPF, and DLGAP5 were upregulated in chemotherapy-resistant AML patients and might be useful for predicting poor prognosis.

Transcriptomic signatures responding to PKM2 activator TEPP-46 in the hyperglycemic human renal proximal epithelial tubular cells.

Pyruvate kinase M2 (PKM2), as the terminal and last rate-limiting enzyme of the glycolytic pathway, is an ideal enzyme for regulating metabolic phenotype. PKM2 tetramer activation has shown a protective role against diabetic kidney disease (DKD). However, the molecular mechanisms involved in diabetic tubular have not been investigated so far. In this study, we performed transcriptome gene expression profiling in human renal proximal tubular epithelial cell line (HK-2 cells) treated with 25 mM high D-glucose (HG) for 7 days before the addition of 10 μM TEPP-46, an activator of PKM2 tetramerization, for a further 1 day in the presence of HG. Afterwards, we analyzed the differentially expressed (DE) genes and investigated gene relationships based on weighted gene co-expression network analysis. The results showed that 2,902 DE genes were identified (adjusted P-value ≤ 0.05), where 2,509 DE genes (86.46%) were co-expressed in the key module. Four extremely downregulated DE genes (HSPA8, HSPA2, HSPA1B, and ARRB1) and three extremely upregulated DE genes (GADD45A, IGFBP3, and SIAH1) enriched in the downregulated endocytosis (hsa04144) and upregulated p53 signaling pathway (hsa04115), respectively, were validated by qRT-PCR experiments. The qRT-PCR results showed that the relative expression levels of HSPA8 [adjusted P-value = 4.45 × 10-34 and log2(FC) = -1.12], HSPA2 [adjusted P-value = 6.09 × 10-14 and log2(FC) = -1.27], HSPA1B [adjusted P-value = 1.14 × 10-11 and log2(FC) = -1.02], and ARRB1 [adjusted P-value = 2.60 × 10-5 and log2(FC) = -1.13] were significantly different (P-value < 0.05) from the case group to the control group. Furthermore, the interactions and predicted microRNAs of the key genes (HSPA8, HSPA2, HSPA1B, and ARRB1) were visualized in networks. This study identified the key candidate transcriptomic biomarkers and biological pathways in hyperglycemic HK-2 cells responding to the PKM2 activator TEPP-46 that can highlight a possibility of PKM2 tetramerization reshaping the interplay among endocytic trafficking through the versatile networks of Hsp70s and rewiring the crosstalk between EGFR signal transduction circuits and metabolic stress to promote resilience, which will be valuable for further research on PKM2 in DKD.

Exposure to Contaminated River Water is Associated with Early Hatching and Dysregulation of Gene Expression in Early Life Stages of the Endangered Copper Redhorse (Moxostoma hubbsi)

The copper redhorse (Moxostoma hubbsi) is an endangered fish that spawns exclusively in the Richelieu River (Quebec, Canada). Tributaries of the Richelieu are contaminated with high levels of current-use pesticides, which may impact early-life stage (ELS) copper redhorse and other native fishes. We assessed the effects of exposure to contaminated river water on ELS copper redhorse and river redhorse (Moxostoma carinatum), a related fish that shares the copper redhorse's spawning grounds and nursery habitat. A riverside flow-through system was used to expose copper and river redhorse embryos (1000 each) to Richelieu River water or laboratory water as a control. Fish were maintained until 14 days posthatch, and water samples were taken daily for chemical analysis. Following a heavy rain event, concentrations of two neonicotinoid pesticides, clothianidin and thiamethoxam, exceeded water quality guidelines for aquatic life (20 ng/L). Using nontargeted screening, we tentatively identified an additional 24 pharmaceutical and personal care products and 23 pesticides in river water. Effects of river water on ELS fish were observed in both species, but the copper redhorse appeared to be more sensitive. Fish exposed to river water hatched 10.7 (copper redhorse) and 2.4 (river redhorse) cumulative degree days earlier than controls. Copper redhorse survival was significantly lower in river water (73 ± 16%) compared to laboratory water (93 ± 3%), whereas river redhorse survival was similar between treatments (84 ± 6% and 89 ± 4%, respectively). Sequencing of copper redhorse larvae RNA revealed 18 differentially expressed genes (DEGs) following 14 days of exposure to river water. Eight up-regulated DEGs were linked to immune function and injury response, and seven down-regulated DEGs were involved with digestion and nutrient absorption. The present study provided valuable data on the effects of ELS exposure to a real-world mixture of contaminants in two fish species of concern. Environ Toxicol Chem 2022;41:1950-1966. © 2022 SETAC.

Genes and Diseases: Insights from Transcriptomics Studies.

Results of expression studies can be useful to clarify the genotype-phenotype relationship. However, according to data from recent literature, there is a large group of genes that are revealed as differentially expressed (DE) in many studies, regardless of the biological context. Additional analyses could shed more light on the relationships between genes, their differential expression, and diseases. We generated a set of 9972 disease genes from five gene-phenotype databases (OMIM, ORPHANET, DDG2P, DisGeNet and MalaCards) and a report of the International Union of Immunological Societies. To study transcriptomics of disease and non-disease genes in healthy tissues, we obtained data from the Human Protein Atlas (HPA) website. We analyzed the dependency between expression in healthy tissues and gene occurrence in Gene Expression Omnibus series using tools within the Enrichr libraries. The results of expression studies were annotated with Gene Ontology (GO) and Human Phenotype Ontology (HPO) terms. Using transcriptomics analysis of healthy tissues, we validated the previous findings of higher expression levels of disease genes in pathologically linked tissues compared to other tissues. Preferentially DE genes were generally highly expressed in one or multiple tissues and were enriched for disease genes. According to the results of GO enrichment analyses, both down- and up-regulated DE genes most often took part in immune response, translation and tissue-specific processes. A connection between DE-related pathology and the diversity of HPO terms was found. Investigating a link between expression and phenotype contributes to understanding the mode of development and progression of human diseases.

Combined Analysis of Pharmaceutical Active Ingredients and Transcriptomes of Glycyrrhiza uralensis Under PEG6000-Induced Drought Stress Revealed Glycyrrhizic Acid and Flavonoids Accumulation via JA-Mediated Signaling.

Glycyrrhiza uralensis contains many secondary metabolites with a wide range of pharmacological activities. Drought stress acts as a positive regulator to stimulate the production of medicinal active component in G. uralensis, however, the underlying mechanism remains unclear. The aim of this work is to investigate the accumulation and regulatory mechanism of pharmaceutical active ingredients in G. uralensis under drought stress. The materials of the aerial and underground parts of G. uralensis seedlings treated by 10% PEG6000 for 0, 2, 6, 12, and 24 h were used for RNA sequencing and determination of phytohormones and pharmaceutical active ingredients. PEG6000, ibuprofen (IBU), and jasmonic acid (JA) were utilized to treat G. uralensis seedlings for content detection and gene expression analysis. The results showed that, the contents of glycyrrhizic acid, glycyrrhetinic acid, and flavonoids (licochalcone A, glabridin, liquiritigenin, isoliquiritigenin, and liquiritin) were significantly accumulated in G. uralensis underground parts under drought stress. Kyoto Encyclopedia of Genes and Genomes analysis of the transcriptome data of drought-treated G. uralensis indicated that up-regulated differentially expressed genes (UDEGs) involved in glycyrrhizic acid synthesis in the underground parts and flavonoids synthesis in both aerial and underground parts were significantly enriched. Interestingly, the UDEGs participating in jasmonic acid (JA) signal transduction in both aerial and underground parts were discovered. In addition, JA content in both aerial and underground parts under drought stress showed the most significantly accumulated. And drought stress stimulated the contents of JA, glycyrrhizic acid, and flavonoids, coupled with the induced expressions of genes regulating the synthesis and transduction pathway. Moreover, In PEG6000- and JA-treated G. uralensis, significant accumulations of glycyrrhizic acid and flavonoids, and induced expressions of corresponding genes in these pathways, were observed, while, these increases were significantly blocked by JA signaling inhibitor IBU. JA content and expression levels of genes related to JA biosynthesis and signal transduction were also significantly increased by PEG treatment. Our study concludes that drought stress might promote the accumulation of pharmaceutical active ingredients via JA-mediated signaling pathway, and lay a foundation for improving the medicinal component of G. uralensis through genetic engineering technology.

Gene Expression Microarray Data Identify Hub Genes Involved in Osteoarthritis

The present study was performed to explore the underlying molecular mechanisms and screen hub genes of osteoarthritis (OA) via bioinformatics analysis. In total, twenty-five OA synovial tissue samples and 25 normal synovial tissue samples were derived from three datasets, namely, GSE55457, GSE55235, and GSE1919, and were used to identify the differentially expressed genes (DEGs) of OA by R language. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of DEGs were conducted using the Database for Annotation, Visualization, and Integrated Discovery (DAVID). A Venn diagram was built to show the potential hub genes identified in all three datasets. The STRING database was used for constructing the protein–protein interaction (PPI) networks and submodules of DEGs. We identified 507 upregulated and 620 downregulated genes. Upregulated DEGs were significantly involved in immune response, MHC class II receptor activity, and presented in the extracellular region, while downregulated DEGs were mainly enriched in response to organic substances, extracellular region parts, and cadmium ion binding. Results of KEGG analysis indicated that the upregulated DEGs mainly existed in cell adhesion molecules (CAMs), while downregulated DEGs were significantly involved in the MAPK signaling pathway. A total of eighteen intersection genes were identified across the three datasets. These include Nell-1, ATF3, RhoB, STC1, and VEGFA. In addition, 10 hub genes including CXCL12, CXCL8, CCL20, and CCL4 were found in the PPI network and module construction. Identification of DEGs and hub genes associated with OA may be helpful for revealing the molecular mechanisms of OA and further promotes the development of relevant biomarkers and drug targets.

Interaction between live seaweed and various Vibrio species by co-culture: Antibacterial activity and seaweed microenvironment

The antibacterial activity of live seaweed (Ulva fasciata, Gracilaria blodgettii and Pelvetia siliguosa) were examined on the three Vibrio (V.) strains (V. parahaemolyticus J1 and V. alginolyticus R1 from maricultural areas, a standard strain of V. parahaemolyticus ATCC17802). Three seaweed species all showed strong inhibition on the culturability of Vibrio strains. Ulva (U.) fasciata as representative seaweed was further studied, which also exhibited inhibition on the swimming behavior, twitching behavior and biofilm formation of three strains. Much more up-regulated differentially expressed genes (DEGs) of U. fasciata occurred in response to V. alginolyticus (5213) than those to both two strains V. parahaemolyticus (3860, 4167), and more than 90% DEGs related to the biosynthesis of secondary metabolites (SMs) were all up-regulated (128–161 DEGs), the least of which and their related pathways occurred by the stress of the standard strain. The stress of two strains from mariculture areas caused richer and more similar profile of up-regulated DEGs in the 30–36 types of SMs biosynthesis pathways, especially for ubiquinone and other terpenoid-quinone biosynthesis with five more up-regulated DEGs than those by the stress of the standard strain. However, the most up-regulated DEGs in the pathways of biosynthesis of unsaturated fatty acids and steroid biosynthesis occurred under the stress of the standard strain and V. alginolyticus, respectively. In addition, Vibrio strains largely promoted the enrichment of endophytic bacteria (EB), mainly for phylum Proteobacteria and genera Alteromonas, Fretibacter, Ruegeria, etc., which favored the U. fasciata against Vibrio. The more significant variation of EB community was also caused by the Vibrio stress isolated from mariculture areas than that by standard strain.