Expression Of NFKBIA Research Articles

Different expression patterns of inflammation-related genes and serum microRNAs in young-onset ischemic stroke

Brain ischemia results in the activation of a cascade of inflammatory responses, contributing to the pathogenesis of stroke. This study aimed to assess the patterns of possible changes in the expression of specific inflammation-associated protein-encoding genes and miRNAs in the peripheral blood between the acute and chronic phase of young-onset cryptogenic (Cryp) and large-artery atherosclerotic (LAA) stroke. Blood and serum were collected from patients with cryptogenic and large-artery atherosclerotic stroke at the stroke onset and 1-year follow-up. The relative expression of inflammation-related genes was analysed at the mRNA and miRNA levels using real-time quantitative PCR. Moreover, the relationship between the relative gene expression levels and clinical data was assessed using several different statistical approaches. Seventy-three patients were included in this study, with a median age of 47 (IQR, 9) years. Approximately 72% were men. In patients with cryptogenic stroke, at the mRNA level, ICAM1, CXCL8, TNF, NFKBIA, PYCARD, IL1B, and IL18 were observed to be upregulated at the stroke onset compared to the 1-year follow-up. In patients with LAA stroke, only the expression of NFKBIA was significantly higher during acute stroke. Further, the miRNA serum levels of miR-21, miR-122, and miR-155 were higher at the onset of stroke in patients with cryptogenic stroke but not in those with LAA stroke. The differences between the relative gene expression levels during acute stroke and at the 1-year follow-up were more pronounced in patients with cryptogenic stroke with no cardiovascular risk factors. The expression changes of inflammatory genes in whole blood and miRNAs in the serum differ in patients with cryptogenic and LAA stroke.

Higher expression of BRCA1, and CHUK and lower expression of NFKBIA, ESR1, PIK3R1, and PPARG in HAM/TSP compared to ATLL, a diverse pathological consequence

Higher expression of BRCA1, and CHUK and lower expression of NFKBIA, ESR1, PIK3R1, and PPARG in HAM/TSP compared to ATLL, a diverse pathological consequence

A bird's eye view of the potential role of NFKBIA in pan-cancer

In the 21st century, cancer remains a serious threat to people's health and has become a prominent public health problem. NFKBIA is involved in the pathological process of many diseases including cancer, but its specific role in pan-cancer has not yet been fully elucidated. This study aims to deepen the understanding of cancer pathology by analyzing the potential functions of NFKBIA in pan-cancer. We used TCGA data to analyze differences of expression of NFKBIA in pan-cancer. We explored the prognostic value, clinical relevance, immune relevance, potential biological function, and diagnosis and treatment value of NFKBIA in pan-cancer through bioinformatics analysis. This study found that in pan-cancer, NFKBIA exhibits differences in expression, which correlate with the prognosis, diagnosis, treatment value and clinical and immune parameters. We have identified that Aspirin, Astaxanthin and Bardoxolone methyl are expected to play a potential therapeutic role in pan-cancer. The results of this study will help to improve our understanding of the role and potential mechanism of NFKBIA in cancer pathology, which may provide guidance for cancer-related research and clinical diagnosis and treatment.

Formononetin inhibits IgE by huPlasma/PBMCs and mast cells/basophil activation via JAK/STAT/PI3-Akt pathways.

Food allergy is a prevalent disease in the U.S., affecting nearly 30 million people. The primary management strategy for this condition is food avoidance, as limited treatment options are available. The elevation of pathologic IgE and over-reactive mast cells/basophils is a central factor in food allergy anaphylaxis. This study aims to comprehensively evaluate the potential therapeutic mechanisms of a small molecule compound called formononetin in regulating IgE and mast cell activation. In this study, we determined the inhibitory effect of formononetin on the production of human IgE from peripheral blood mononuclear cells of food-allergic patients using ELISA. We also measured formononetin's effect on preventing mast cell degranulation in RBL-2H3 and KU812 cells using beta-hexosaminidase assay. To identify potential targets of formononetin in IgE-mediated diseases, mast cell disorders, and food allergies, we utilized computational modeling to analyze mechanistic targets of formononetin from various databases, including SEA, Swiss Target Prediction, PubChem, Gene Cards, and Mala Cards. We generated a KEGG pathway, Gene Ontology, and Compound Target Pathway Disease Network using these targets. Finally, we used qRT-PCR to measure the gene expression of selected targets in KU812 and U266 cell lines. Formononetin significantly decreased IgE production in IgE-producing human myeloma cells and PBMCs from food-allergic patients in a dose-dependent manner without cytotoxicity. Formononetin decreased beta-hexosaminidase release in RBL-2H3 cells and KU812 cells. Formononetin regulates 25 targets in food allergy, 51 in IgE diseases, and 19 in mast cell diseases. KEGG pathway and gene ontology analysis of targets showed that formononetin regulated disease pathways, primary immunodeficiency, Epstein-Barr Virus, and pathways in cancer. The biological processes regulated by formononetin include B cell proliferation, differentiation, immune response, and activation processes. Compound target pathway disease network identified NFKB1, NFKBIA, STAT1, STAT3, CCND1, TP53, TYK2, and CASP8 as the top targets regulated at a high degree by formononetin. TP53, STAT3, PTPRC, IL2, and CD19 were identified as the proteins mostly targeted by formononetin. qPCR validated genes of Formononetin molecular targets of IgE regulation in U266 cells and KU812 cells. In U266 cells, formononetin was found to significantly increase the gene expression of NFKBIA, TP53, and BCL-2 while decreasing the gene expression of BTK TYK, CASP8, STAT3, CCND1, STAT1, NFKB1, IL7R. In basophils KU812 cells, formononetin significantly increased the gene expression of NFKBIA, TP53, and BCL-2 while decreasing the gene expression of BTK, TYK, CASP8, STAT3, CCND1, STAT1, NFKB1, IL7R. These findings comprehensively present formononetin's mechanisms in regulating IgE production in plasma cells and degranulation in mast cells.

Comparative Transcriptome Analyses in Mantle Cell Lymphoma (MCL): Evidence of E2F1 As a Major Target in Aggressive Blastoid MCL Model

Background Mantle cell lymphoma (MCL) is a highly aggressive B-cell lymphoma characterized by the translocation t(11;14)(q13;q32) which triggers the dysregulation of the cyclin D1 by placing its coding gene CC1ND1 under the control of an immunoglobulin promoter. One of the major consequences of this molecular event is the disturbance of the cyclin D/Rb pathway leading to Rb inactivation and activation the elongation factor E2F1 mainly implicated in cell cycle progression. The mainstay of MCL therapies remains the chemo-immunotherapy followed by autologous transplantation but recently several novel therapies have been developed such as BTK inhibitors, Venetoclax, CDK4/6 inhibitors and more recently CAR-T therapies. However, patients with aggressive forms of MCL exhibit low response rates, requiring the identification of novel therapeutic targets. Materials and Methods We have previously generated an MCL cell line (UPN-1) from a patient with a blastoid variant of MCL presenting unique morphological and cytogenetics characteristics (M'kacher et al, Oncogene 2003). This cell line has been shown to harbor a TP53 mutation and a highly increased sensitivity to radiation despite the absence of functional p53. The molecular basis of this phenomenon has not been explored. In this work, we further characterized UPN-1 cell line using the Clarium S human microarray which has been compared to the gene expression profile of other existing MCL cell lines (GRANTA-519, JEKO-1, JVM-2, MAVEK-1, MINO, REC-1, SP-49, SP-53, Z-138). After mathematical correction, a supervised analysis was performed between triplicate samples of UPN-1 and a group constituted with all other MCL cell lines. We have then set up in vitro assays to determine the effect of a direct E2F1 inhibitor on the proliferative potential of UPN-1. Results The comparative transcriptome analyses revealed a list of 47 genes specifically upregulated in UPN-1. Unsupervised classification allowed to discriminate UPN-1 samples from other MCL cell lines based on the expression of these 47 genes. These data suggested a specific expression profile of UPN-1 as compared to other cell lines. Among the upregulated genes, E2F1 elongation factor was found to be highly upregulated in UPN-1 as compared to other MCL cell lines along with positive enrichment of E2F targets which are mainly implicated in cell cycle progression (Figure 1). Interestingly, UPN-1 has a low level of BCL-2 as compared to other MCL lines that could explain its radio sensitivity. However, other signaling pathways involved in the inhibition of apoptosis were found to be present in UPN-1 by geneset enrichment analysis. The expression of CDKN1A (p21), major cell cycle regulator and known as a marker of aggressive MCL, was found to be low in UPN-1. A repression of immune response pathway such as NFKB with low expression of NFKBIA was identified. Finally, the expression of PIK3CD which encodes for a therapeutic target in MCL (such as PI3KDelta inhibitor Idelalisib) was also significantly repressed in UPN-1 as compared to nine other MCL cell lines. To evaluate the effect of the direct inhibition of E2F1, we treated UPN-1 cells with a targeted E2F1 inhibitor 5‘-Deoxy-5‘-(methylthio)adenosine (MTA) which is a protein methylation inhibitor. MTT assays revealed a 50% reduction of proliferation at + 4 hours in comparison to the vehicle-treated conditions (P<0.0001).Annexin assays showed an increase in early apoptosis (36.6 %) at + 8 hours with subsequent rise in late apoptotic cells (64.5 %) observed after 24 hours of treatment. Interestingly, qRT-PCR analyses of MTA-treated cells exhibited a decrease in the expression of downstream factors associated with E2F1, such as RAD51, BRCA1, CDK2, CCNB2, RANBP1. Conclusions We report here for the first time the comparative transcriptome analysis of the unique aggressive blastoid MCL line UPN-1 as compared to other MCL lines described. This cell line exhibited a major upregulation of E2F1 expression and a down regulation of the cell cycle inhibitor CDKN1A(p21). The potential effects of the direct inhibition of E2F1 using the targeted inhibitor MTA demonstrated a reduction in cell proliferation and induction of apoptosis, suggesting the possibility of associating clinically acceptable E2F1 inhibition strategies to other current therapies.

Novel diagnostic biomarkers of oxidative stress, immunological characterization and experimental validation in Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative condition causing cognitive decline. Oxidative stress (OS) is believed to contribute to neuronal death and dysfunction in AD. We conducted a study to identify differentially expressed OS-related genes (DEOSGs) through bioinformatics analysis and experimental validation, aiming to develop a diagnostic model for AD. We analyzed the GSE33000 dataset to identify OS regulator expression profiles and create molecular clusters (C1 and C2) associated with immune cell infiltration using 310 AD samples. Cluster analysis revealed significant heterogeneity in immune infiltration. The 'WGCNA' algorithm identified cluster-specific and disease-specific differentially expressed genes (DGEs). Four machine learning models (random forest (RF), support vector machine (SVM), generalized linear model (GLM) and extreme gradient boosting (XGB)) were compared, with GLM performing the best (AUC = 0.812). Five DEOSGs (NFKBIA, PLCE1, CLIC1, SLCO4A1, TRAF3IP2) were identified based on the GLM model. AD subtype prediction accuracy was validated using nomograms and calibration curves. External datasets (GSE122063 and GSE106241) confirmed the expression levels and clinical significance of important genes. Experimental validation through RT-qPCR showed increased expression of NFKBIA, CLIC1, SLCO4A1, TRAF3IP2, and decreased expression of PLCE1 in the temporal cortex of AD mice. This study provides insights for AD research and treatment, particularly focusing on the five model-related DEOSGs.

MiR-196b-5p activates NF-κB signaling in non-small cell lung cancer by directly targeting NFKBIA

BackgroundOur recent study found that QKI-5 regulated miRNA, miR-196b-5p, promotes non-small cell lung cancer (NSCLC) progression by directly targeting GATA6, TSPAN12 and FAS. However, the biological functions of miR-196b-5p in NSCLC progression and metastasis still remain elusive. MethodsCell proliferation, migration, colony formation, cell cycle assays were used to investigate cellular phenotypic changes. Quantitative real-time PCR (qRT-PCR) and western blot analyses were used to measure expressions of relative gene and protein. Interaction between QKI-5 and miR-196b-5p was determined by RNA immunoprecipitation (RIP) assay. Luciferase reporter assay was used to determine direct binding between miR-196b-5p and NFKBIA 3′-UTR. ELISA assay was used to measure secreted IL6 proteins. Mice xenograft model was used to assess the functions of NFKBIA on in vivo tumor growth. ResultsWe demonstrated that the miR-196b-5p facilitates lung cancer cell proliferation, migration, colony formation, and cell cycle by directly targeting NFKBIA, a negative regulator of NFκB signaling. Knocking down NFKBIA increases IL6 mediated phosphorylation of signal transducer and activator of transcription 3 (STAT3) to promote lung cancer cell growth by activating NFκB signaling. The expression of NFKBIA was significantly downregulated in NSCLC tissue samples, and was negatively correlated with the expression miR-196b-5p. In addition, we found that downregulated QKI-5 expression was associated with the elevated miR-224 expression in NSCLC. ConclusionsOur findings indicated that the miR-224/QKI-5/miR-196b-5p/NFKBIA signaling pathway might play important functions in the progression of NSCLC, and suggested that targeting this pathway might be an effective therapeutic strategy in treating NSCLC.

Identification of biomarkers related to copper metabolism in patients with pulmonary arterial hypertension

BackgroundThe pathogenesis of pulmonary arterial hypertension (PAH) and associated biomarkers remain to be studied. Copper metabolism is an emerging metabolic research direction in many diseases, but its role in PAH is still unclear.MethodsPAH-related datasets were downloaded from the Gene Expression Omnibus database, and 2067 copper metabolism-related genes (CMGs) were obtained from the GeneCards database. Differential expression analysis and the Venn algorithm were used to acquire the differentially expressed CMGs (DE-CMGs). DE-CMGs were then used for the coexpression network construction to screen candidate key genes associated with PAH. Furthermore, the predictive performance of the model was verified by receiver operating characteristic (ROC) analysis, and genes with area under the curve (AUC) values greater than 0.8 were selected as diagnostic genes. Then support vector machine, least absolute shrinkage and selection operator regression, and Venn diagrams were applied to detect biomarkers. Moreover, gene set enrichment analysis was performed to explore the function of the biomarkers, and immune-related analyses were utilized to study the infiltration of immune cells. The drug-gene interaction database was used to predict potential therapeutic drugs for PAH using the biomarkers. Biomarkers expression in clinical samples was verified by real-time quantitative PCR.ResultsFour biomarkers (DDIT3, NFKBIA, OSM, and PTGER4) were screened. The ROC analysis showed that the 4 biomarkers performed well (AUCs > 0.7). The high expression groups for the 4 biomarkers were enriched in protein activity-related pathways including protein export, spliceosome and proteasome. Furthermore, 8 immune cell types were significantly different between the two groups, including naive B cells, memory B cells, and resting memory CD4 T cells. Afterward, a gene-drug network was constructed. This network illustrated that STREPTOZOCIN, IBUPROFEN, and CELECOXIB were shared by the PTGER4 and DDIT3. Finally, the results of RT-qPCR in clinical samples further confirmed the results of the public database for the expression of NFKBIA and OSM.ConclusionIn conclusion, four biomarkers (DDIT3, NFKBIA, OSM, and PTGER4) with considerable diagnostic values were identified, and a gene-drug network was further constructed. The results of this study may have significant implications for the development of new diagnostic biomarkers and actionable targets to expand treatment options for PAH patients.

The eighth International Forum on Stem Cells: virtual meeting, October 20-21, 2022.

The eighth International Forum on Stem Cells: virtual meeting, October 20-21, 2022.

Identification of Key Genes and miRNAs Affecting Osteosarcoma Based on Bioinformatics.

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

Single-cell transcriptome profiling reveals heterogeneous neutrophils with prognostic values in sepsis

SummaryNeutrophils constitute the largest proportion of nucleated peripheral blood cells, and neutrophils have substantial heterogeneity. We profiled nearly 300,000 human peripheral blood cells in this study using single-cell RNA sequencing. A large proportion (>50%) of these cells were annotated as neutrophils. Neutrophils were further clustered into four subtypes, including Neu1, Neu2, Neu3, and Neu4. Neu1 is characterized by high expression of MMP9, HP, and RGL4. Neu1 was associated with septic shock and significantly correlated with the sequential organ failure assessment (SOFA) score. A gene expression module in Neu1 named Neu1_C (characterized by expression of NFKBIA, CXCL8, G0S2, and FTH1) was highly predictive of septic shock with an area under the curve of 0.81. The results were extensively validated in external bulk datasets by using single-cell deconvolution methods. In summary, our study establishes a general framework for studying neutrophil-related mechanisms, prognostic biomarkers, and potential therapeutic targets for septic shock.

Computational and Mechanistic Study of the Therapeutic Potential of Compound Formononetin in Preventing Mast Cell Activation and IgE Production in Food Anaphylaxis

The prevalence of allergic diseases such as Asthma, Rhinitis, Anaphylaxis, Eczema, Urticaria, and Angioedema have been rising dramatically in low‐ and middle‐income countries around the world. According to American College of Allergy, Asthma & Immunology, allergies are the 6th leading cause of chronic illness in the U.S. and more than 50 million Americans suffer from allergies each year. Extensive data suggest that food allergies affect up to 10% of the world population and have been increasing in the last two to three decades. Globally 300 million people have asthma and about 200 to 250 million people suffer from food allergies. Previously we have shown that a small molecule compound, formononetin have shown decreases IgE production from B cells. We hypothesize that formononetin will be a potential candidate for preventing food anaphylaxis in food allergy by regulating molecular pathways that involved in IgE production in B cells, mast cell activation and degranulation. The objective of this study is to reveal potential therapeutic mechanisms of formononetin in prevention and treatment of food anaphylaxis from food allergy, by using computational modeling, including target mining, gene ontology enrichment, pathway analyses, protein‐protein interaction analyses and in silico molecular docking to identify gene and protein targets that are regulated in food allergy and mast cells diseases. The targets identified were further validated using qRT‐PCR in B cell multiple myeloma cell line. U266 cells were cultured to 1.0 × 106 cells/mL and then incubated with formononetin at different concentrations of 20 μg/mL for 72 hours. Supernatants were collected for measuring IgE levels by ELISA and cell viability was determined using Trypan blue dye. mRNA expression was determined for the top targets identified and compared with GAPDH. Mining formononetin targets into obtained disease targets uncovers 25 targets for food allergy, 51 targets for IgE diseases and 19 targets for mast cell diseases. This targets where further used for gene ontology, KEGG pathway analysis, CTPD network and protein‐protein interaction. Top regulated KEGG pathways include the primary immunodeficiency pathway, Epstein Barr virus infection pathway, Th17 cell differentiation, PI3K‐Akt pathway. Regulated gene ontology biological processes include B cell proliferation, B cell differentiation, B cell activation and apoptotic process. Formononetin decreased IgE at after 72 hours, P < 0.05 without cytotoxicity. Formononetin decreased mRNA expression of P53, TYK2, CASP8 significantly (P < 0.05) and IgEH very significantly (P < 0.01) and increased the mRNA expression of BCL2, NFKBIA and BTK significantly (P < 0.05) after 72 hours in the culture group compared to the untreated group. These findings suggest that formononetin may be a novel therapeutic candidate for treatment and prevention of IgE/mast cell mediated food allergy and other allergic diseases.

Porphyromonas gingivalis W83 Membrane Components Induce Distinct Profiles of Metabolic Genes in Oral Squamous Carcinoma Cells.

Periodontitis, a chronic inflammatory disease is caused by a bacterial biofilm, affecting all periodontal tissues and structures. This chronic disease seems to be associated with cancer since, in general, inflammation intensifies the risk for carcinoma development and progression. Interactions between periodontal pathogens and the host immune response induce the onset of periodontitis and are responsible for its progression, among them Porphyromonas gingivalis (P. gingivalis), a Gram-negative anaerobic rod, capable of expressing a variety of virulence factors that is considered a keystone pathogen in periodontal biofilms. The aim of this study was to investigate the genome-wide impact of P. gingivalis W83 membranes on RNA expression of oral squamous carcinoma cells by transcriptome analysis. Human squamous cell carcinoma cells (SCC-25) were infected for 4 and 24 h with extracts from P. gingivalis W83 membrane, harvested, and RNA was extracted. RNA sequencing was performed, and differential gene expression and enrichment were analyzed using GO, KEGG, and REACTOME. The results of transcriptome analysis were validated using quantitative real-time PCR with selected genes. Differential gene expression analysis resulted in the upregulation of 15 genes and downregulation of 1 gene after 4 h. After 24 h, 61 genes were upregulated and 278 downregulated. GO, KEGG, and REACTONE enrichment analysis revealed a strong metabolic transcriptomic response signature, demonstrating altered gene expressions after 4 h and 24 h that mainly belong to cell metabolic pathways and replication. Real-time PCR of selected genes belonging to immune response, signaling, and metabolism revealed upregulated expression of CCL20, CXCL8, NFkBIA, TNFAIP3, TRAF5, CYP1A1, and NOD2. This work sheds light on the RNA transcriptome of human oral squamous carcinoma cells following stimulation with P. gingivalis membranes and identifies a strong metabolic gene expression response to this periodontal pathogen. The data provide a base for future studies of molecular and cellular interactions between P. gingivalis and oral epithelium to elucidate the basic mechanisms of periodontitis and the development of cancer.

Single-Cell Transcriptome Analysis of Chronic Antibody-Mediated Rejection After Renal Transplantation.

Renal transplantation is currently the most effective treatment for end-stage renal disease. However, chronic antibody-mediated rejection (cABMR) remains a serious obstacle for the long-term survival of patients with renal transplantation and a problem to be solved. At present, the role and mechanism underlying immune factors such as T- and B- cell subsets in cABMR after renal transplantation remain unclear. In this study, single-cell RNA sequencing (scRNA-seq) of peripheral blood monocytes (PBMCs) from cABMR and control subjects was performed to define the transcriptomic landscape at single-cell resolution. A comprehensive scRNA-seq analysis was performed. The results indicated that most cell types in the cABMR patients exhibited an intense interferon response and release of proinflammatory cytokines. In addition, we found that the expression of MT-ND6, CXCL8, NFKBIA, NFKBIZ, and other genes were up-regulated in T- and B-cells and these genes were associated with pro-inflammatory response and immune regulation. Western blot and qRT-PCR experiments also confirmed the up-regulated expression of these genes in cABMR. GO and KEGG enrichment analyses indicated that the overexpressed genes in T- and B-cells were mainly enriched in inflammatory pathways, including the TNF, IL-17, and Toll-like receptor signaling pathways. Additionally, MAPK and NF-κB signaling pathways were also involved in the occurrence and development of cABMR. This is consistent with the experimental results of Western blot. Trajectory analysis assembled the T-cell subsets into three differentiation paths with distinctive phenotypic and functional prog rams. CD8 effector T cells and γδ T cells showed three different differentiation trajectories, while CD8_MAI T cells and naive T cells primarily had two differentiation trajectories. Cell-cell interaction analysis revealed strong T/B cells and neutrophils activation in cABMR. Thus, the study offers new insight into pathogenesis and may have implications for the identification of novel therapeutic targets for cABMR.

Upregulation of T Cell Receptor Signaling Pathway Components in Gestational Diabetes Mellitus Patients: Joint Analysis of mRNA and circRNA Expression Profiles.

ObjectiveGestational diabetes mellitus (GDM) is one of the most common complications of pregnancy, and its pathogenesis is still unclear. Studies have shown that circular RNAs (circRNAs) can regulate blood glucose levels by targeting mRNAs, but the role of circRNAs in GDM is still unknown. Therefore, a joint microarray analysis of circRNAs and their target mRNAs in GDM patients and healthy pregnant women was carried out.MethodsIn this study, microarray analyses of mRNA and circRNA in 6 GDM patients and 6 healthy controls were conducted to identify the differentially expressed mRNA and circRNA in GDM patients, and some of the discovered mRNAs and circRNAs were further validated in additional 56 samples by quantitative realtime PCR (qRT-PCR) and droplet digital PCR (ddPCR).ResultsGene ontology and pathway analyses showed that the differentially expressed genes were significantly enriched in T cell immune-related pathways. Cross matching of the differentially expressed mRNAs and circRNAs in the top 10 KEGG pathways identified 4 genes (CBLB, ITPR3, NFKBIA, and ICAM1) and 4 corresponding circRNAs (circ-CBLB, circ-ITPR3, circ-NFKBIA, and circ-ICAM1), and these candidates were subsequently verified in larger samples. These differentially expressed circRNAs and their linear transcript mRNAs were all related to the T cell receptor signaling pathway, and PCR results confirmed the initial microarray results. Moreover, circRNA/miRNA/mRNA interactions and circRNA-binding proteins were predicted, and circ-CBLB, circ-ITPR3, and circ-ICAM1 may serve as GDM-related miRNA sponges and regulate the expression of CBLB, ITPR3, NFKBIA, and ICAM1 in cellular immune pathways.ConclusionUpregulation of T cell receptor signaling pathway components may represent the major pathological mechanism underlying GDM, thus providing a potential approach for the prevention and treatment of GDM.

Effects of air exposure on survival, histological structure, non-specific immunity and gene expression of red claw crayfish (Cherax quadricarinatus)

Air exposure exerts unavoidable environmental stress during the transport of crayfish. This study evaluated survival, histological structure, non-specific immunity and gene expression in red claw crayfish (Cherax quadricarinatus) under air exposure. Survival experiment showed that crayfish under air exposure started dying after 24 h, and survival rate decreased to 62.2% and 20.0% after 36 h and 48 h respectively. In the second experiment, crayfish divided into six groups with different air exposure time (0 h, 4 h, 8 h, 12 h, 24 h and 36 h). Crayfish maintained in the normal culture environment were used as control (0 h), whereas other five groups were placed in tanks without water. Histopathological study found that air exposure resulted in swelling and vacuolation of hepatocytes and enterocytes. Total antioxidant capacity (T-AOC), activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and alkaline phosphatase (AKP) increased at 4–8 h short-term air exposure, and then decreased. T-AOC in hepatopancreas was suppressed significantly at 36 h exposure. Malondialdehyde (MDA) levels in both hepatopancreas and hemolymph raised after 4 h and then reduced to the initial level. Expression of antioxidant genes (SOD, GPx and Trx 1), immune-related genes (C-LZM, CRU-2, and SERPIN3), anti-stress related genes (HSP70 and HSP90) were induced after short-term air exposure, and then down-regulated. Expression of SERP, HSP21 and NFKBIA maintained high levels during 36 h air exposure, indicating that these genes play vital roles in the defense mechanism against air exposure. In brief, our study demonstrates that air exposure causes pathological changes in hepatopancreas and intestines, and altered antioxidant and immune status in C. quadricarinatus. The suppressed antioxidant status, leading to tissue oxidative damage, is one of the main reason for death of crayfish under long-term air exposure.

Study on the effect and mechanism of NFKBIA on cervical cancer progress in vitro and in vivo.

NFKBIA is frequently encountered. However, its expression and relevance of the proliferation, invasion, and migration in human cervical cancer (CC) remain unclear. The role and novel mechanism of NFKBIA in CC progression were investigated in this study. We analyzed the expression of NFKBIA in CC and adjacent normal tissues and explored the proliferation, migration, and invasion of HeLa cells by treating with either wild-type NFKBIA plasmid or NFKBIA siRNA. Effect of NFKBIA on the epithelial-mesenchymal transition (EMT) and the β-catenin-mediated transcription of target genes were evaluated subsequently. NFKBIA expression was lower in CC tissues than that of adjacent tissues. An obvious dysregulation of NFKBIA overexpression was revealed in CC cell proliferation, invasion, and migration, which differed from the effect of knockdown NFKBIA. NFKBIA overexpression facilitated the expression of both phosphorylated β-catenin and E-cadherin protein. It inhibited the expression of vimentin, TWIST, as well as downstream targets of β-catenin including c-MYC, TCF-4 and MMP14. Conversely, NFKBIA silencing elevated the expression of c-MYC, TCF-4, and MMP14, and promoted the EMT in HeLa cells. Both endogenous and exogenous NFKBIA interacted with β-catenin. Moreover, β-catenin overexpression stemmed effects of NFKBIA on the proliferation, migration, and invasion of HeLa cells. By overexpressing NFKBIA in vivo, the volume and size of tumors were notably decreased, while no obvious alteration was found in mice body weight. By inhibiting β-catenin-mediated transcription, NFKBIA functioning as a tumor suppressor might be introduced as a novel anti-metastatic agent for the treatment of targeted CC.

Abstract 1024: Combination activity of acalabrutinib and capivasertib in diffuse large B-cell lymphoma

Abstract Relapsed/refractory DLBCL is an aggressive B-cell malignancy with limited treatment options. BTK inhibitors have demonstrated preclinical and clinical activity in DLBCL of the activated B-cell (ABC) subset, but responses are limited and not durable. Therefore, an acalabrutinib (BTK inhibitor) combination screen was conducted in a panel of 9 DLBCL cell lines to identify synergistic and active combinations. The in vitro combination of acalabrutinib plus capivasertib (AKT inhibitor) demonstrated significant combination benefit in the ABC-DLBCL cell lines TMD8 (Loewe synergy score 9.3) and OCI-LY10 (Loewe synergy score 3.2). Capivasertib and acalabrutinib monotherapy gave 0% and 85% tumor growth inhibition (TGI) in TMD8 and 5% and 79% in OCI-LY10 tumour xenograft models respectively. The combination gave tumour regressions of 99% in TMD8 and 72% in OCI-LY10. To explore a potential mechanism of action, RNAseq analysis was performed on the TMD8 cell line and TMD8 xenograft tumors treated with the monotherapies and combination. 24-hour monotherapy treatment of TMD8 cells with acalabrutinib altered the expression of 7390 genes while capivasertib altered 398 genes (absolute fold change ≥ 1.25 and adjusted p-value < 0.05), with enrichment for genes regulated by NFkB signalling detected. Pathway analysis following combination treatment revealed significant decrease in expression of the G2M checkpoint pathway genes compared to acalabrutinib alone (Adjusted p-value = 0.007) as well as decreased expression of CDK1, AURKA and MYC, and significant shifts in the TNFa signalling via NFkB pathway (Adjusted p-value = 0.02). The NFkB signalling node was examined in more detail in TMD8 tumor samples treated with the combination. Lower expression of NFKBIA, NFKBID, EGR2 and BCL2A1 was evident in the combination group, compared to control tumors (p<0.01 for all). Interestingly, the combination strongly increased the expression of MS4A2 (CD20) compared to control (Adjusted p-value = 0.002), capivasertib monotherapy (Adjusted p-value = 0.004) and acalabrutinib monotherapy (Adjusted p-value = 0.02). Therefore, we tested the addition of Rituxan to acalabrutinib + capivasertib combination in the TMD8 xenograft model. The triple combination produced durable complete regressions in 5/5 mice after cessation of treatment whereas tumors regrew after cessation of treatment of the acalabrutinib + capivasertib doublet. These data suggest that the combination of BTK and AKT inhibition may enhance anti-tumor activity in ABC DLBCL, with the addition of anti-CD20 giving more durable tumour control. Citation Format: Kathleen Burke, Justine Roderick-Richardson, Natasha Narang, Brandon Willis, Hannah Dry, Lillian Castriotta, Alan Rosen, Jay Mettetal, Simon Barry, Andrew Bloecher. Combination activity of acalabrutinib and capivasertib in diffuse large B-cell lymphoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1024.

NF-кB pathway genes expression in chicken erythrocytes infected with avian influenza virus subtype H9N2

ABSTRACT 1. Chicken erythrocytes in blood vessels are the most abundant circulating cells, which participate in the host’s immune responses. The transcription factor nuclear factor-kappa B (NF-κB) plays a vital role in the inflammatory response following viral infections. However, the expression of the NF-κB pathway, and other immune-related genes in chicken erythrocytes infected with low pathogenic avian influenza virus (LPAIV H9N2), has not been extensively studied. 2. The following study determined the interaction of LPAIV H9N2 with chicken erythrocytes using indirect immunofluorescence microscopy. This was followed by investigating myeloid differentiation primary response 88 (MyD88), C-C motif chemokine ligand 5 (CCL5), melanoma differentiation-associated protein 5 (MDA5), the inhibitor of nuclear factor-kappa B kinase subunit epsilon (IKBKE), NF-κB inhibitor alpha (NFKBIA), NF-κB inhibitor epsilon (NFKBIE), interferon-alpha (IFN-α), colony-stimulating factor 3 (CSF3) and tumour necrosis factor receptor-associated factor 6 (TRAF6) by mRNA expression using quantitative real-time PCR (qRT-PCR) at four different time intervals (0, 2, 6 and 10 h). 3. There was a significant interaction between erythrocytes and LPAIV H9N2 virus. Furthermore, the mRNA expression of the NF-κB pathway and other immune-related genes were significantly up-regulated at 2 h post-infection in infected chicken erythrocytes, except for TRAF6, which were significantly downregulated. While at 0 h post-infection, IFN-α and CSF3 were significantly upregulated, whereas NFKBIA was significantly downregulated. Further expression of MDA5, CCL5 and NFKBIA was upregulated, while TRAF6 was downregulated at 6 h post-infection. In infected erythrocytes, expression of MyD88, CCL5 and IKBKE was upregulated. However, IFN-α and TRAF6 were downregulated at 10 h post-infection. 4. These results give initial evidence that the NF-κB pathway, and other genes related to immunity, in chicken erythrocytes may contribute to LPAIV subtype H9N2 and induce host immune responses.

The relationship of CCL4, BCL2A1, and NFKBIA genes with premature aging in women of Yin deficiency constitution.

Traditional Chinese Medicine (TCM) defined constitution as a health statue or physical fitness that determines individual susceptibility to diseases. Yin deficiency constitution (YinDC) is a type of constitution closely related to aging. Previous studies found that the characteristic genes of YinDC are part of the inflammatory aging signaling pathways (e.g., NF-kappa B). Therefore, the aim of the study was to further reveal the dysregulation of genes associated with inflammatory aging in YinDC women. This study adopted the industrial standard of constitutional judgment, and screened YinDC (n=30) and Balanced constitution (BC) (n=30) from women between the ages of 35 to 49, a range categorized as the degenerating period by TCM. Five genes CCL4, BCL2A1, NFKBIA, TAK1, and IL-8 were analyzed by qRT-PCR. Logistical regression revealed the correlation between body constitution and the expression of the five genes: the expression of NFKBIA and CCL4 mRNA was significantly up-regulated, whereas the expression of BCL2A1 mRNA was significantly down-regulated in YinDC (P<0.05). Age or weight, when included in the model, did not affected the correlations. Increased mRNA expression of CCL4 and NFKBIA and decreased mRNA expression of BCL2A1 may be the molecular basis of premature aging of YinDC women. These results provide a mechanistic basis for early conditioning of YinDC, anti-aging, and the prevention of aging-related diseases.