Transcription Factor NF-kappaB Research Articles

IL1R2 is a Novel Prognostic Biomarker for Lung Adenocarcinoma.

The aim of this study is to figure out the role of IL1R2 in LUAD (lung adenocarcinoma). IL1R2, a special member of IL-1 receptor family, binds to IL-1 and plays an important role in inhibiting IL-1 pathway, which seems to be involved in tumorigenesis. Emerging studies demonstrated higher IL1R2 expression levels in several malignancies. In the present study, we assessed the expression of IL1R2 in LUAD tissues with immunohistochemistry and explored various databases to determine whether it could be a potential prognostic biomarker and therapeutic target. The expression level of IL1R2 in lung adenocarcinoma was analyzed by Immunohistochemistry and UALCAN database. The correlation between IL1R2 expression and the patient prognosis was identified by Kaplan-Meier plotter. The correlation of IL1R2 expression with immune infiltrates was clarified by TIMER database. The protein-protein interaction network and gene functional enrichment analysis were constructed and performed by STRING and Metascape database. Immunohistochemistry showed that the expression of IL1R2 was higher in tumor tissues of LUAD patients and that patients with lower IL1R2 level have a better prognosis than their counterparts. We validated our findings in several online databases and found that IL1R2 gene was also positively correlated with B cells and neutrophils and biomarkers of CD8+T cells and exhausted T cells. PPI network and gene enrichment analyses showed that expression of IL1R2 was also associated with complex functionspecific networks involving IL-1 signal, NF-KappaB transcription factors. According to these findings, we demonstrated that IL1R2 was involved in the progression and prognosis of LUAD and the underlying mechanism needs further investigation.

Abstract 1073: Longitudinal Imaging Of NF-kappaB In Murine Pulmonary Embolism

Objective: Pulmonary embolism (PE) is a common condition that urgently needs novel diagnostic and therapeutic strategies. Here, we investigated PE-induced inflammation via longitudinal imaging of transcription factor NF-kappaB (NFkB) activation signals in murine PE. Understanding the temporal activation of inflammatory mediators, such as NFkB, in PE may lead to future medical interventions or better patient stratification. Methods: NFkB activation was assessed using male, 12-16w old, NFkB-GFP-luciferase (NGL) reporter mice (n=20 PE animals). Stasis venous thrombosis (VT) was induced in the femoral veins of donor mice on day 0. On day 1, the thrombus was resected and intravenously injected into recipient mice to create the PE model. PE animals underwent bioluminescence imaging on days -1, 1, 4, and 7. Regions-of-interest (ROI) areas were kept constant and placed over areas of PE as verified in other modalities. Lung and PE morphometrics were imaged by fluorescence-mediated tomography and computed tomography using a fibrin-specific molecular imaging agent, FTP11 (i.v. 150nmol/kg). Cardiac function pre- and post-PE was assessed by ultrasound and pulmonary arterial pressure measurements. Mouse cohorts were sacrificed 1, 4, and 7 days following stasis-VT embolization and underwent histological analysis of the PE. Results: The mean volume of the femoral stasis-VT was 1.21±0.25 mm 3 . The bioluminescence signal plateau was found at 4-8 minutes post-intravenous injection of D-luciferin in PE animals. The six-minute post-injection image was utilized in comparing longitudinal images. Bioluminescence from the activation of NFkB in the PE ROI showed a maximal signal in the day 1 PE with a 55.0% (334 to 151RU) decrease in signal from day 1 to day 7(p<0.05). Histological and protein assays validated the activation and localization of NFkB. In PE animals, immunohistochemistry of infiltrating CD45-positive cells showed increased cell numbers from day 1 to day 7 (p<0.05). Since PE is confided spatially to a lung lobe, contralateral PE-negative lobes also showed increased infiltrating cells compared to sham animals. Conclusions: Quantitative serial imaging pre- and post-PE demonstrates an inflammatory timeline in PE-induced NFkB activation.

Prognostic and immunological role of adaptor related protein complex 3 subunit mu2 in colon cancer

The expression levels and prognostic role of AP3M2 in colorectal adenocarcinoma (CRAC) have yet to be fully unveiled. Our study comprehensively investigated the clinical significance of AP3M2 in colorectal cancer through an extensive bioinformatics data mining process (TCGA, GEO, GEPIA, Timer, Ualcan, ROCPLOT, and David), followed by experimental validation. We found AP3M2 is a cancer gene, which can be used to distinguish between colorectal cancer and colorectal adenomas, liver metastasis, lung metastasis, colorectal polyp. Higher AP3M2 expression levels were associated with longer overall survival in colon adenocarcinoma. AP3M2 might be the primary biomarker for oxaliplatin in colon cancer and an acquired resistance biomarker for oxaliplatin and 5-fu. AP3M2 was positively associated with CD274, CTLA4. AP3M2 might be associated with T-cell, NF-kappaB transcription factor activity, and response to hypoxia. AP3M2 could predict chemotherapy effectiveness and prognosis for colon cancer patients. AP3M2 might inhibit tumor growth via influencing tumor-infiltrating immune cells in the context of Tumor microenvironment. AP3M2 plays as an oncogene in CRAC and is suggested as a new potential biotarget for therapy.

Single-Cell Sequencing Reveals a Close Correlation between the Number of MDSCs and the Gene Expression Levels of GPX1 and S100A8/S100A9 in Patients with Myelofibrosis

Myelofibrosis is a type of myeloproliferative disorder, and research indicates that the immunosuppressive tumor microenvironment plays a significant role in the pathogenesis of myelofibrosis, with myeloid-derived suppressor cells (MDSCs) being the primary inhibitory cells involved in shaping the tumor immune microenvironment. In this study, we used flow cytometry to detect the number of MDSCs in the bone marrow of myelofibrosis patients and revealed the relationship between MDSCs and gene expression levels through single-cell transcriptome sequencing. We utilized flow cytometry to identify the number of Lin-HLA-DR-CD33+ labeled MDSCs in the bone marrow of 56 myelofibrosis patients and 20 healthy individuals. Additionally, we performed single-cell transcriptome sequencing on samples from two myelofibrosis patients, one showing a favorable response to Lurcitanib treatment and the other showing an ineffective response. The results of flow cytometry revealed a significant increase in the percentage of MDSCs in the bone marrow of myelofibrosis patients (15.16±6.25%) compared to healthy individuals (1.72±0.88%) (p<0.0001) (Fig a, b). Among the two patients subjected to single-cell sequencing, the percentage of MDSCs in their samples was 19.3% and 8.9% (Fig c). Following treatment with different concentrations of decitabine, the MDSCs cell count progressively decreased with increasing treatment time and concentration (Fig d).Subsequently, we identified the major cell types in the two samples through single-cell sequencing, which included T cells, Neutrophils, Monocytes, Erythroblasts, B cells, Platelets, Myelocytes, Hematopoietic Stem Cells（HSCs）, and progenitor Myelocytes (Fig e). The analysis of marker gene expression for each major cell type showed good specificity (Fig f). By comparing the proportions of different cell types between the two samples, it was observed that the high MDSC group had a decrease in the proportions of B cells, T cells, Monocytes, and Platelets, while an increase in the proportions of Neutrophils, Erythroblasts, HSC, Myelocytes, and progenitor Myelocytes (Fig g).Furthermore, we analyzed the highly differentially expressed genes in different cell types and identified significantly enriched pathways. The results showed significant downregulation of the GPX1 gene in monocytes of the high MDSC group, whereas the S100A8/S100A9 genes were significantly upregulated in HSC (Fig h). Enrichment analysis revealed significant enrichment of pathways related to reactive oxygen species metabolism, immune-inflammatory response, etc., in monocytes of the high MDSCs group, and significant enrichment of pathways related to NF-kappaB transcription factor activity regulation, Jak-STATs signaling, mTOR signaling in HSC. Research has shown that the number of MDSCs is elevated in patients with myelofibrosis . Lurcitanib exhibits poor therapeutic efficacy in myelofibrosis patients with high MDSCs counts, whereas decitabine demonstrates more favorable treatment outcomes. Single-cell sequencing results revealed significant downregulation of the GPX1 gene in monocytes of myelofibrosis patients with high MDSCs counts. GPX1 gene encodes glutathione peroxidase 1, which plays a role in reducing oxidative stress and inflammation, and has anti-fibrotic effects.Conversely, in HSC of myelofibrosis patients with high MDSCs counts, the S100A8/S100A9 gene expression is significantly upregulated. This gene can induce neutrophil chemotaxis and adhesion, serving as a danger-associated molecular pattern (DAMP) molecule and stimulating innate immune cells through pattern recognition receptors (e.g., TLR4 and AGER), subsequently activating MAP kinase and NF-kappa-B signaling pathways, leading to inflammation and fibrosis.Thus, it can be inferred that the number of MDSCs in myelofibrosis patients is negatively correlated with GPX1 gene expression and positively correlated with S100A8/S100A9 gene expression. Acknowledgement: This research was funded by the Key R&D Program of Zhejiang, No. 2022C03137; Public Technology Application Research Program of Zhejiang, China, No. LGF21H080003; Zhejiang Medical Association Clinical Medical Research special fund project, No. 2022ZYC-D09. Correspondence to: Dr Jian Huang, Department of Hematology, The First Affiliated Hospital of Zhejiang University School of Medicine. househuang@zju.edu.cn

TMT-Based Proteomics Analysis of Senescent Nucleus Pulposus from Patients with Intervertebral Disc Degeneration.

Lower back pain, a leading cause of disability worldwide, is associated with intervertebral disc degeneration (IDD) in approximately 40% of cases. Although nucleus pulposus (NP) cell senescence is a major contributor to IDD, the underlying mechanisms remain unclear. We collected NP samples from IDD patients who had undergone spinal surgery. Healthy and senescent NP tissues (n = 3) were screened using the Pfirrmann grading system combined with immunohistochemistry, as well as hematoxylin and eosin, Safranin O, Alcian blue, and Masson staining. Differentially expressed proteins (DEPs) were identified using quantitative TMT-based proteomics technology. Bioinformatics analyses included gene ontology (GO) annotation, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and protein-protein interaction (PPI) analyses. In addition, immunofluorescence was used to verify protein expression. In total, 301 DEPs were identified in senescent NP tissues, including 92 upregulated and 209 downregulated proteins. In GO, DEPs were primarily associated with NF-kappaB transcription factor, extracellular regions, cellular protein metabolic processes, and post-translational protein modification. The enriched KEGG pathways included TGF-β, Wnt, RAP1, interleukin-17, extracellular matrix-receptor adhesion, and PI3K/Akt signaling pathways. PPI analysis demonstrated interactions between multiple proteins. Finally, immunofluorescence verified the expressions of MMP3, LUM, TIMP1, and CDC42 in senescent NP cells. Our study provides valuable insights into the mechanisms underlying senescent NP tissues in IDD patients. DEPs provide a basis for further investigation of the effects of senescent factors on IDD.

The influence of the commensal skin bacterium Staphylococcus epidermidis on the epidermal barrier and inflammation: Implications for atopic dermatitis.

The skin microbiota is a crucial component in maintaining cutaneous barrier function. Staphylococcus epidermidis is considered as a beneficial commensal member of the cutaneous microbiota promoting skin health. However, S. epidermidis is also frequently detectable in the skin of patients with the inflammatory skin disease atopic dermatitis (AD) and some studies reported a significantly higher presence of S. epidermidis in severe AD as compared to mild AD. Therefore, this study aimed to analyse the impact of S. epidermidis on the expression of cutaneous inflammatory mediators and skin barrier molecules. Various S. epidermidis skin-derived isolates activated the proinflammatory transcription factor NF-kappaB and induced expression of AD-associated proinflammatory cytokines in human primary keratinocytes and 3D skin equivalents. Skin barrier molecules such as filaggrin were downregulated by S. epidermidis. In general, AD-derived S. epidermidis strains elicited a higher response than strains derived from the skin of healthy individuals. Taken together, our results provide further evidence that the abundance of S. epidermidis in AD may trigger the inflammatory scenario associated with this disease.

MicroRNAs Promote the Progression of Sepsis-Induced Cardiomyopathy and Neurovascular Dysfunction Through Upregulation of NF-kappaB Signaling Pathway-Associated HDAC7/ACTN4.

IntroductionThe objective of this study was to determine the NF-kappaB pathway, hub genes, and transcription factors (TFs) in monocytes implicated in the progression of neurovascular-related sepsis-induced cardiomyopathy (SIC) as well as potential miRNAs with regulatory functions.Methods: Sepsis-induced cardiomyopathy—and heart failure (HF)-related differentially expressed genes (DEGs) between SIC and HF groups were identified separately by differential analysis. In addition, DEGs and differentially expressed miRNAs (DEmiRNAs) in monocytes between sepsis and the HC group were identified. Then, common DEGs in SIC, HF, and monocyte groups were identified by intersection analysis. Based on the functional pathways enriched by these DEGs, genes related to the NF-kB-inducing kinase (NIK)/NF-kappaB signaling pathway were selected for further intersection analysis to obtain hub genes. These common DEGs, together with sepsis-related DEmiRNAs, were used to construct a molecular interplay network and to identify core TFs in the network.Results: A total of 153 upregulated genes and 25 downregulated genes were obtained from SIC-, HF-, and monocyte-related DEGs. Functional pathway analysis revealed that the upregulated genes were enriched in NF-κB signaling pathway. A total of eight genes associated with NF-κB signaling pathway were then further identified from the 178 DEGs. In combination with sepsis-related DEmiRNAs, HDAC7/ACTN4 was identified as a key transcriptional regulatory pair in the progression of SIC and in monocyte regulation. hsa-miR-23a-3p, hsa-miR-3175, and hsa-miR-23b-3p can regulate the progression of SIC through the regulation of HDAC7/ACTN4. Finally, gene set enrichment analysis (GSEA) suggested that HDAC7/ACTN4 may be associated with apoptosis in addition to the inflammatory response.Conclusion: hsa-miR-23a-3p, hsa-miR-3175, and hsa-miR-23b-3p are involved in SIC progression by regulating NF-κB signaling signaling pathway-related HDAC7/ACTN4 in monocytes and cardiac tissue cells. These mechanisms may contribute to sepsis-induced neurovascular damage.

Abstract P5-10-03: Mutant p53 and ERK1/2 MAPK cooperate with the production of TNBC inflammatory secretome

Abstract TP53 is the most frequently mutated gene in most types of human cancer, including breast cancer. The triple-negative breast cancer (TNBC) subtype in particular displays TP53 mutation in approximately 80% of patients. Unlike other breast cancer subtypes (e.g., ER/PR-positive or HER2-positive), TNBC patients currently lack an approved highly effective targeted therapy. Notably, most TNBC acquires TP53 mutations, which, in addition to the loss of canonical p53 functions, can result in gain-of-function, activating different cellular mechanisms involved in tumor phenotypes such as proliferation, metastasis, invasiveness, and angiogenesis. Here we evaluate the role of mutant p53 in cancer phenotypes of TNBC cell lines, in particular their inflammatory profile. As expected, loss of p53 protein by small interfering RNA depletion did not show a major impact on the cell viability of MDA-MB231 and Hs578t cells in MTT assay. However, depleting mutant p53 knockdown made MDA-MB231 cells more susceptible to treatment with methyl-methane sulfonate, a genotoxic alkylating agent. Interestingly, cell invasion as measured by the transwell assay demonstrated that depletion of mutant p53 depletion decreased the invasiveness potential of MDA-MB231 and Hs578t cells which was substantiated with decreased migration of MDA-MB231 cells in a scratch assay over 24 h. RNA sequencing of MDA-MB231 and Hs578t cells revealed that mutant p53 knockdown decreased the expression of several constitutively expressed pro-inflammatory cytokines such as IL8, IL6, CXCL2, and CXCL3, but not genes associated with survival to alkylating agents (NRF2 and Endoplasmic reticulum stress markers) or genes typically regulated by wild-type p53 when compared to control-silenced and MMS-treated cells as indicated by Pathway Enrichment Analysis using the Enrich R and DAVID tools. These results were confirmed by ELISA quantification of IL8, IL6, and CXCL2 in MDA-MB231 and Hs578t transfected with two sequences of siRNA targeting mutant p53. On the other hand, RNA sequencing revealed some constitutively expressed genes known to be involved in breast cancer cells malignancy, such as PTGS2 (COX-2 enzyme gene) and MMP1, which were not affected by p53 knockdown. We found that MMP1, and PGE2 (the product of PTGS2/COX-2 enzyme), are upregulated by the ERK1/2 MAPK signaling pathway as determined in MDA-MB231 cells treated with the MEK1/2 inhibitor UO126 and sorafenib. UO126 and sorafenib also decreased IL8 and IL6 production. Furthermore, combined mutant p53 knockdown with MEK/ERK1/2 pathway inhibition caused a more pronounced IL8 and IL6 inhibition when compared to either p53 knockdown or UO126/sorafenib alone, whereas MMP1 and PGE2 levels were only reduced by MEK/ERK1/2 inhibitor treatments. Interestingly, neither mutant p53 knockdown impacted ERK1/2 phosphorylation status nor did UO126/sorafenib alter mutant p53 immunocontent. Reporter assays showed that mutant p53 promotes NFkappaB reporter activation, and MEK/ERK1/2 controls both NFkappaB and AP-1 transcription factors, both associated with the expression of the secretome components evaluated herein. Functional cell assays showed that concomitant inhibition of mutant p53 and MEK/ERK1/2 pathways reduce cell proliferation, invasion, and migration, indicating that mutant p53 protein gain of function cooperates with ERK1/2 MAPK signaling pathway to promote secretome production and malignant phenotypes in TNBC cell models. Citation Format: Raquel Nascimento das Neves, Aparna Gorthi, Alexander James Roy Bishop, Alfeu Zanotto Filho. Mutant p53 and ERK1/2 MAPK cooperate with the production of TNBC inflammatory secretome [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-10-03.

Inflammatory activation of surface molecule shedding by upregulation of the pseudoprotease iRhom2 in colon epithelial cells

The metalloproteinase ADAM17 contributes to inflammatory and proliferative responses by shedding of cell-surface molecules. By this ADAM17 is implicated in inflammation, regeneration, and permeability regulation of epithelial cells in the colon. ADAM17 maturation and surface expression requires the adapter proteins iRhom1 or iRhom2. Here we report that expression of iRhom2 but not iRhom1 is upregulated in intestinal tissue of mice with acute colitis. Our analysis of public databases indicates elevated iRhom2 expression in mucosal tissue and epithelial cells from patients with inflammatory bowel disease (IBD). Consistently, expression of iRhom2 but not iRhom1 is upregulated in colon or intestinal epithelial cell lines after co-stimulation with tumor necrosis factor (TNF) and interferon gamma (IFNgamma). This upregulation can be reduced by inhibition of Janus kinases or transcription factors NF-kappaB or AP-1. Upregulation of iRhom2 can be mimicked by iRhom2 overexpression and is associated with enhanced maturation and surface expression of ADAM17 which then results in increased cleavage of transforming growth factor (TGF) alpha and junctional adhesion molecule (JAM)-A. Finally, the induction of these responses is suppressed by inhibition of iRhom2 transcription. Thus, inflammatory induction of iRhom2 may contribute to upregulated ADAM17-dependent mediator and adhesion molecule release in IBD. The development of iRhom2-dependent inhibitors may allow selective targeting of inflammatory ADAM17 activities.

205 The skin commensal Staphylococcus epidermidis induces inflammatory mediators in keratinocytes: Implications for atopic dermatitis

The chronic inflammatory skin disease atopic dermatitis (AD) is characterized by a dysbiosis of the microbiota. This is reflected by a decreased microbial diversity and the abundance of specific potential pathogenic strains such as Staphylococcus (S.) aureus. Staphylococcus (S.) epidermidis is an abundant member of the cutaneous microbiota and is considered generally as an apathogenic commensal that exhibits beneficial effects in AD, e.g. by controlling the growth of S. aureus. However, S. epidermidis is also frequently present in skin lesions of AD patients and a correlation between abundance of S. epidermidis and severity of AD has been reported. We therefore hypothesized that S. epidermidis may exhibit proinflammatory activities. To evaluate this hypothesis we stimulated cultured human keratinocytes and 3D skin models with different S. epidermidis isolates derived either from healthy skin or from lesional skin of AD patients. Real-time PCR and ELISA analyses revealed a significant induced expression of the AD-relevant cytokine thymic stromal lymphopoietin (TSLP) in keratinocytes and 3D skin after stimulation with S. epidermidis strains. S. epidermidis strains derived from lesional AD skin induced a higher TSLP expression than strains derived from healthy skin. Similar results were also obtained with the pro-inflammatory cytokines IL-17c and TNF-alpha. Both, S. epidermidis strains derived from healthy skin and strains derived from AD skin activated the inflammation-associated transcription factor NF-kappaB. Moreover, expression of the barrier-associated protein filaggrin was downregulated by S. epidermidis. Together, the observed S. epidermidis-mediated induction of proinflammatory mediators and downregulation of filaggrin in keratinocytes and 3D skin models may explain why AD patients carrying a high number of S. epidermidis are associated with a more severe inflamed disease state.

Network pharmacology integrated with experimental validation reveals the regulatory mechanism of plastrum testudinis in treating senile osteoporosis

Ethnopharmacological relevancePlastrum testudinis (PT) has been used in traditional Chinese medicine to treat bone diseases such as senile osteoporosis (SOP) for thousands of years. However, the underlying mechanisms remain largely unknown. Aim of the studyThis study aims to investigate the possible molecular mechanism of PT in the treatment of SOP using an integrated strategy of network pharmacology and experimental validation. Materials and methodsThe compounds of PT and its targets were identified through the BATMAN-TCM database. The SOP-related targets were retrieved from the GeneCards database. Protein-protein interaction information was obtained by inputting the intersection targets into the STRING database. Cytoscape software was used to construct a protein-protein interaction network and a PT-compound-target-SOP network. Using Cytoscape and R software, we conducted GO function and KEGG pathway enrichment analyses. We also conducted in vivo and in vitro experiments to verify the network pharmacology findings. ResultsIn total, 6 active compounds and 342 targets of PT were screened, of which 57 common targets were related to SOP. The GO biological process enrichment analysis identified 880 entries, mainly relating to the regulation of hormone response, the cell apoptotic process, the apoptotic signaling pathway, NF-kappaB transcription factor activity, fatty acid transportation, osteoclast differentiation, macrophage activation, and inflammatory response. The KEGG pathway enrichment analysis identified 52 entries, including 14 related signaling pathways, which mainly involved the TNF, MAPK, IL-17, AGE-RAGE, estrogen, relaxin, and other signaling pathways. Our in vivo experiments confirmed that PT alleviates SOP, while the in vitro experiments demonstrated that PT exerts a suppressive effect on osteoclast differentiation and bone resorption in a concentration-dependent manner. Furthermore, we observed that PT downregulates the expression of osteoclast-specific genes, including C-FOS, TNF, and BDNF, in the MAPK signaling pathway. ConclusionThrough network pharmacology and experimental validation, this study is the first to report that PT downregulates the expression of osteoclast-specific genes, including C-FOS, TNF, and BDNF, in the MAPK signaling pathway, thus exerting a suppressive effect on osteoclast differentiation and bone resorption, which may be the molecular mechanism for PT treatment of SOP.

Circular RNA CircNOLC1, Upregulated by NF-KappaB, Promotes the Progression of Prostate Cancer via miR-647/PAQR4 Axis.

BackgroundCircRNAs recently have shown critical roles in tumor biology. However, their roles in prostate cancer (PCa) remains largely unclear.MethodsCircRNA microarrays were performed in immortal prostate cell line RWPE1 and PCa cell lines as DU145, PC3, LNCaP, C4-2, and 22RV1. Combined with upregulated circRNAs in PCa tissues, circNOLC1 expression was validated in PCa cells and tissues via qRT-PCR and FISH. Sanger sequencing, actinomycin D, gDNA, and cDNA, RNase R assays were used to assess the circular characteristics of circNOLC1. CCK-8, colony formation, transwell migration assays, and mice xenograft models were conducted to evaluate the functions of PCa cells after circNOLC1 knockdown and overexpression. RNA pulldown, luciferase reporter assay, FISH (fluorescence in situ hybridization), and CHIP were utilized to illustrate the further mechanisms of circNOLC1.ResultsOur research indicated that circNOLC1 was overexpressed in PCa cells and tissues, and circNOLC1 was more stable than linear NOLC1 mRNA. CircNOLC1 promoted PCa cells proliferation and migration in vitro and vivo. Additionally, we found that circNOLC1 could upregulate PAQR4 expression by sponging miR-647, leading to the activation of PI3K/Akt pathway. Moreover, NF-kappaB was identified to bind to the NOLC1 promoter sites and upregulated both NOLC1 and circNOLC1 expression.ConclusionCircNOLC1, elevated by transcription factor NF-kappaB, promotes PCa progression via a miR-647/PAQR4 axis, and circNOLC1 is a potential biomarker and target for PCa treatment.

Expression of p52, a non-canonical NF-kappaB transcription factor, is associated with poor ovarian cancer prognosis

BackgroundThe canonical and non-canonical nuclear factor-kappaB (NF-κB) signaling pathways have key roles in cancer, but studies have previously evaluated only the association of canonical transcription factors and ovarian cancer survival. Although a number of in vitro and in vivo studies have demonstrated mechanisms by which non-canonical NF-κB signaling potentially contributes to ovarian cancer progression, a prognostic association has yet to be shown in the clinical context.MethodsWe assayed p65 and p52 (major components of the canonical and non-canonical NF-κB pathways) by immunohistochemistry in epithelial ovarian tumor samples; nuclear and cytoplasmic staining were semi-quantified by H-scores and dichotomized at median values. Associations of p65 and p52 with progression-free survival (PFS) and overall survival (OS) were quantified by Hazard Ratios (HR) from proportional-hazards regression.ResultsAmong 196 cases, median p52 and p65 H-scores were higher in high-grade serous cancers. Multivariable regression models indicated that higher p52 was associated with higher hazards of disease progression (cytoplasmic HR: 1.54; nuclear HR: 1.67) and death (cytoplasmic HR: 1.53; nuclear HR: 1.49), while higher nuclear p65 was associated with only a higher hazard of disease progression (HR: 1.40) in unadjusted models. When cytoplasmic and nuclear staining were combined, p52 remained significantly associated with increased hazards of disease progression (HR: 1.91, p = 0.004) and death (HR: 1.70, p = 0.021), even after adjustment for p65 and in analyses among only high-grade serous tumors.ConclusionsThis is the first study to demonstrate that p52, a major component of non-canonical NF-κB signaling, may be an independent prognostic factor for epithelial ovarian cancer, particularly high-grade serous ovarian cancer. Approaches to inhibit non-canonical NF-κB signaling should be explored as novel ovarian cancer therapies are needed.

Identification of Hub genes associated with infection of three lung cell lines by SARS-CoV-2 with integrated bioinformatics analysis.

Identification of Hub genes associated with infection of three lung cell lines by SARS-CoV-2 with integrated bioinformatics analysis.

Coronavirus Disease 2019 and Hypertension: The Role of Angiotensin-Converting Enzyme 2 and the Renin-Angiotensin System.

Hypertension emerged from early reports as a potential risk factor for worse outcomes for persons with coronavirus disease 2019 (COVID-19). Among the putative links between hypertension and COVID-19 is a key counter-regulatory component of the renin-angiotensin system (RAS): angiotensin-converting enzyme 2 (ACE2). ACE2 facilitates entry of severe acute respiratory syndrome coronavirus 2, the virus responsible for COVID-19, into host cells. Because RAS inhibitors have been suggested to increase ACE2 expression, health-care providers and patients have grappled with the decision of whether to discontinue these medications during the COVID-19 pandemic. However, experimental models of analogous viral pneumonias suggest RAS inhibitors may exert protective effects against acute lung injury. We review how RAS and ACE2 biology may affect outcomes in COVID-19 through pulmonary and other systemic effects. In addition, we briefly detail the data for and against continuation of RAS inhibitors in persons with COVID-19 and summarize the current consensus recommendations from select specialty organizations.

Interleukin-1b-Induced IRAK1 Ubiquitination is Required for TH-GM-CSF Cell Differentiation in T Cell-Mediated Inflammation

Accumulating evidence suggests granulocyte macrophage-colony stimulating factor (GM-CSF) can function as an inflammatory mediator, but whether GM-CSF-producing CD4+ T cells (TH-GM-CSF) are a distinct T helper cell subset is lacking. Herein we demonstrate that interleukin (IL)-1b exclusively drives differentiation of naive CD4+ T cells into TH-GM-CSF cells via inducing ubiquitination of IL-1 receptor-associated kinase 1 (IRAK1) and subsequent activation of the transcription factor NF-kappaB (NF-κB), independent of RAR-related orphan receptor gamma (RORt) required for TH17 differentiation. In vivo, TH-GM-CSF cells are present in murine Citrobacter Rodentium infections and mediate colitis following adoptive transfer of CD4+ T cells into Rag1-/- mice via GM-CSF-induced macrophage activation. The TH-GM-CSF cell phenotype is stable and distinct from the TH17 genetic program, but IL-1b can convert pre-formed TH17 cells into TH-GM-CSF cells, thereby accounting for previously reported associations between IL-17 and GM-CSF. Together, our results newly identify IL-1b/NFκB-dependent TH-GM-CSF cells as a unique T helper cell subset and highlight the importance of CD4+ T cell-derived GM-CSF induced macrophage activation as a previously undescribed T cell effector mechanism. Funding Statement: Huabao Xiong was supported by NIH funding RO1AI104688. Peter. S Heeger was supported by RO1AI132405 and RO1AI071185. Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: The animal study protocols were approved by the Institutional Animal Care and Use Committees of Mount Sinai School of Medicine.

Abstract 1573: Non-canonical NF-kappaB signaling in the tumor microenvironment in relation to ovarian cancer survival

Abstract Introduction: Despite advances in treatment, ovarian cancer associated mortality has not substantially improved over the last 50 years. Understanding mechanisms that underlie disease progression is crucial to the development of new treatments for ovarian cancer. Mediated by a family of transcription factors, the canonical and non-canonical nuclear factor-kappaB signaling pathways have known roles in cell proliferation, as well as immune, inflammatory, and neoplastic processes. Non-canonical NF-kappaB signaling requires processing of a cytoplasmic p100/RelB dimer to a mature p52/RelB complex, capable of nuclear translocation to activate transcription of target genes. Prior studies on canonical NF-kappaB expression and ovarian cancer survival have been inconsistent, but no studies to date have evaluated the role of non-canonical NF-kappaB transcription factors. Methods: We conducted immunohistochemical staining for p52, a component of the non-canonical NF-kappaB signaling pathway, on a clinically annotated tissue microarray of ovarian tumors collected from 1994-2004 at the Vanderbilt University Medical Center. Nuclear and cytoplasmic p52 staining in tumors was semi-quantified by H-score and expression was dichotomized at median values. Associations with progression-free survival (PFS) and overall survival (OS) were quantified by hazard ratios (HR) and 95% confidence intervals (CI) using Cox proportional-hazards regression. Results: Among 194 ovarian cancer cases, p52 expression was predominantly cytoplasmic, with a relatively smaller proportion of tumor cells showing nuclear staining (median H-scores: 135.6 vs. 1.0). Cytoplasmic and nuclear p52 H-scores were significantly higher among high grade, advanced stage, and serous tumors (all Wilcoxon rank sum p<0.01). Kaplan-Meier survival analysis showed that cases with high nuclear p52 expression had significantly shorter time to disease progression (5-year PFS high vs. low: 72% vs. 17%, log-rank p<0.001). In regression models that included adjustment for age, stage, grade, and histologic subtype, high nuclear p52 was associated with a more than two-fold shorter PFS (HR 2.30, 95% CI 1.44-4.67). Significance of this finding was unaltered by additional adjustment for race, debulking, and platinum sensitivity (HR 2.26, 95% CI 1.15-4.45). In contrast, significant associations between high nuclear p52 and worse OS, and all associations for cytoplasmic p52 staining were attenuated after adjustment for clinical covariates. Conclusions: This is the first study to demonstrate that nuclear p52, representing active non-canonical NF-kappaB signaling, may be an independent prognostic factor for epithelial ovarian cancer. Thus, interventions that inhibit non-canonical NF-kappaB signaling should be explored as novel therapies to limit ovarian cancer progression. Citation Format: Demetra Hufnagel, Andrew J. Wilson, Jamie Saxon, Timothy Blackwell, Dineo Khabele, Marta A. Crispens, Alicia Beeghly-Fadiel, Fiona Yull. Non-canonical NF-kappaB signaling in the tumor microenvironment in relation to ovarian cancer survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1573.

Interleukin-1β-induced IRAK1 ubiquitination is required for TH-GM-CSF cell differentiation in T cell-mediated inflammation

Accumulating evidence suggests granulocyte macrophage-colony stimulating factor (GM-CSF) can function as an inflammatory mediator, but whether GM-CSF-producing CD4+ T cells (TH-GM-CSF) are a distinct T helper cell subset is lacking. Herein we demonstrate that interleukin (IL)-1β exclusively drives differentiation of naïve CD4+ T cells into TH-GM-CSF cells via inducing ubiquitination of IL-1 receptor-associated kinase 1 (IRAK1) and subsequent activation of the transcription factor NF-kappaB (NF-κB), independent of RAR-related orphan receptor gamma (RORγt) required for TH17 differentiation. In vivo, TH-GM-CSF cells are present in murine Citrobacter Rodentium infections and mediate colitis following adoptive transfer of CD4+ T cells into Rag1−/− mice via GM-CSF-induced macrophage activation. The TH-GM-CSF cell phenotype is stable and distinct from the TH17 genetic program, but IL-1β can convert pre-formed TH17 cells into TH-GM-CSF cells, thereby accounting for previously reported associations between IL-17 and GM-CSF. Together, our results newly identify IL-1β/NF-κB-dependent TH-GM-CSF cells as a unique T helper cell subset and highlight the importance of CD4+ T cell-derived GM-CSF induced macrophage activation as a previously undescribed T cell effector mechanism.

Building the drug-GO function network to screen significant candidate drugs for myasthenia gravis.

Myasthenia gravis (MG) is an autoimmune disease. In recent years, considerable evidence has indicated that Gene Ontology (GO) functions, especially GO-biological processes, have important effects on the mechanisms and treatments of different diseases. However, the roles of GO functions in the pathogenesis and treatment of MG have not been well studied. This study aimed to uncover the potential important roles of risk-related GO functions and to screen significant candidate drugs related to GO functions for MG. Based on MG risk genes, 238 risk GO functions and 42 drugs were identified. Through constructing a GO function network, we discovered that positive regulation of NF-kappaB transcription factor activity (GO:0051092) may be one of the most important GO functions in the mechanism of MG. Furthermore, we built a drug-GO function network to help evaluate the latent relationship between drugs and GO functions. According to the drug-GO function network, 5 candidate drugs showing promise for treating MG were identified. Indeed, 2 out of 5 candidate drugs have been investigated to treat MG. Through functional enrichment analysis, we found that the mechanisms between 5 candidate drugs and associated GO functions may involve two vital pathways, specifically hsa05332 (graft-versus-host disease) and hsa04940 (type I diabetes mellitus). More interestingly, most of the processes in these two pathways were consistent. Our study will not only reveal a new perspective on the mechanisms and novel treatment strategies of MG, but also will provide strong support for research on GO functions.

Expression profile and bioinformatics analysis of COMMD10 in BALB/C mice and human.

COMMD10, a member of COMMD protein, has been proved to target p65 NF-kappaB (nuclear factor-kappaB) subunit and reduce its nuclear translocation, thereby leading to the inactivation of NF-kappaB pathway and suppression of colorectal cancer invasion and metastasis. The aim of this study is to explore its expression pattern and tissue distribution in human normal tissues and other tumor tissues and to investigate the relevant mechanism. We firstly provided the expression profile and histological distribution of COMMD10 in various BALB/c mice tissues and identified the biological distribution of COMMD10 in different kinds of human normal and tumor tissues. We verified the expression profile of COMMD10 using TCGA database. The interacting genes of COMMD10 were predicted by using STRING using. Finally, we performed database, and the microRNAs targeting COMMD10 were predicted using miRDB, miRWalk, TargetScan and microRNA. GO and KEGG pathway analyses were performed to predict the biological function of COMMD10 and its interacting genes. mRNA expression of COMMD10 showed the highest level in the lung and spleen, and the lowest level in the heart and brain. Immunohistochemistry detection revealed that COMMD10 was expressed in different tissues with different degrees and was was located mainly in the cytoplasm. Subsequently, we showed that COMMD10 displayed various degrees of expression in different human normal tissues that mainly located in cytoplasm, while COMMD10 of liver cells resided in both nucleus and cytoplasm. All the tumor tissues except breast small cell carcinoma, breast phyllodes tumor, lung adenocarcinoma, thymoma, cervical cancer and bladder urothelial carcinoma showed that COMMD10 was positive staining in cytoplasm. Kaplan-Meier plotter indicated that renal clear cell carcinoma patients with increased expression level of COMMD10 exhibited longer survival. STRING database revealed that COMMD10 had 41 interacting genes, and data from 4 different databases indicated that hsa-miR-590-3p may be the potential regulator of COMMD10. GO analysis demonstrated that COMMD10 and its interacting genes were mainly enriched in Cullin-RING ubiquitin ligase complexes, binding and transport of copper ions, the transport and steady-state maintenance of copper ions, transcription, translation and transport of proteins, and negatively regulate the activity of NF-kappaB transcription factors. KEGG pathway showed that COMMD10 and its interacting genes were mainly involved in renal cell carcinoma, HIF-1 signaling pathways, ubiquitination-mediated proteolysis, endocytosis and mineral absorption. COMMD10 may play a tumor suppressive role in renal clear cell carcinoma through the miR-590-3p-COMMD10-Cul2-RBX1-NF-κB/HIF/NRF2 pathway and regulate the chemotherapy resistance of various tumor cells to cisplatin.