scholarly journals Gene Expression Profiling of NFATc1-Knockdown in RAW 264.7 Cells: An Alternative Pathway for Macrophage Differentiation

Cells ◽  
2019 ◽  
Vol 8 (2) ◽  
pp. 131 ◽  
Author(s):  
Roberta Russo ◽  
Selene Mallia ◽  
Francesca Zito ◽  
Nadia Lampiasi
Keyword(s):  
Cell Differentiation ◽  
Expression Profiling ◽  
Development Process ◽  
Molecular Mechanisms ◽  
Alternative Pathway ◽  
Cell Types ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Transcriptional Program ◽  
Macrophage Differentiation

NFATc1, which is ubiquitous in many cell types, is the master regulator of osteoclastogenesis. However, the molecular mechanisms by which NFATc1 drives its transcriptional program to produce osteoclasts from macrophages (M) remains poorly understood. We performed quantitative PCR (QPCR) arrays and bioinformatic analyses to discover new direct and indirect NFATc1 targets. The results revealed that NFATc1 significantly modified the expression of 55 genes in untransfected cells and 31 genes after NFATc1-knockdown (≥2). Among them, we focused on 19 common genes that showed changes in the PCR arrays between the two groups of cells. Gene Ontology (GO) demonstrated that genes related to cell differentiation and the development process were significantly (p > 0.05) affected by NFATc1-knockdown. Among all the genes analyzed, we focused on GATA2, which was up-regulated in NFATc1-knockdown cells, while its expression was reduced after NFATc1 rescue. Thus, we suggest GATA2 as a new target of NFATc1. Ingenuity Pathway Analysis (IPA) identified up-regulated GATA2 and the STAT family members as principal nodes involved in cell differentiation. Mechanistically, we demonstrated that STAT6 was activated in parallel with GATA2 in NFATc1-knockdown cells. We suggest an alternative pathway for macrophage differentiation in the absence of NFATc1 due to the GATA2 transcription factor.

scholarly journals Lyophilized cell-free supernatants of Lactobacillus isolates exhibited antibiofilm, antioxidant, and reduces nitric oxide activity in lipopolysaccharide-stimulated RAW 264.7 cells

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e12586
Author(s):  
Phoomjai Sornsenee ◽  
Moragot Chatatikun ◽  
Watcharapong Mitsuwan ◽  
Kantapich Kongpol ◽  
Nateelak Kooltheat ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Molecular Mechanisms ◽  
Antioxidant Activities ◽  
Radical Scavenging ◽  
Pharmaceutical Products ◽  
Raw 264.7 Cells ◽  
Total Phenolic ◽  
Antibiofilm Activity ◽  
Fermented Foods ◽  
Raw 264.7

Background Probiotics can release bioactive substances known as postbiotics, which can inhibit pathogenic microorganisms, improve immunomodulation, reduce antioxidant production, and modulate the gut microbiota. Methods In this study, we evaluated the in vitro antimicrobial effects, antioxidant activity, and anti-inflammatory potential of 10 lyophilized cell-free supernatants (LCFS) of Lactobacillus isolates. LCFS was obtained via centrifugation and subsequent lyophilization of the supernatant collected from the culture medium ofeach isolate. The antibacterial and antibiofilm activities of the LCFS were determined using broth microdilution. The antioxidant potential was evaluated by measuring the total phenolic and flavonoid contents and 2,2-Diphennyl-1-picrylhydrazyl (DPPH) and 2,2’-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation (ABTS+) radical scavenging activities. Results All the isolates were able to inhibit the four tested pathogens. The isolates exhibited strong antibiofilm activity and eradicated the biofilms formed by Acinetobacter buamannii and Escherichia coli. All the prepared Lactobacillus LCFS contained phenols and flavonoids and exhibited antioxidant activities in the DPPH and ABTS+ radical scavenging assays. The MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay revealed that LCFS was not cytotoxic to RAW 264.7 cells. In addition, the ten Lactobacillus LCFS decreased the production of nitric oxide. Conclusions All the isolates have beneficial properties. This research sheds light on the role of postbiotics in functional fermented foods and pharmaceutical products. Further research to elucidate the precise molecular mechanisms of action of probiotics is warranted.

scholarly journals IκB Kinase-Dependent Chronic Activation of NF-κB Is Necessary for p21WAF1/Cip1 Inhibition of Differentiation-Induced Apoptosis of Monocytes

2001 ◽  
Vol 21 (6) ◽  
pp. 1930-1941 ◽  
Author(s):  
Kevin N. Pennington ◽  
Julie A. Taylor ◽  
Gary D. Bren ◽  
Carlos V. Paya
Keyword(s):  
Transcription Factor ◽  
Cell Differentiation ◽  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Dependent Manner ◽  
Macrophage Differentiation ◽  
Monocyte Differentiation ◽  
Cell Functions ◽  
Iκb Kinase ◽  
Induced Apoptosis

ABSTRACT The molecular mechanisms regulating monocyte differentiation to macrophages remain unknown. Although the transcription factor NF-κB participates in multiple cell functions, its role in cell differentiation is ill defined. Since differentiated macrophages, in contrast to cycling monocytes, contain significant levels of NF-κB in the nuclei, we questioned whether this transcription factor is involved in macrophage differentiation. Phorbol 12-myristate 13-acetate (PMA)-induced differentiation of the promonocytic cell line U937 leads to persistent NF-κB nuclear translocation. We demonstrate here that an increased and persistent IKK activity correlates with monocyte differentiation leading to persistent NF-κB activation secondary to increased IκBα degradation via the IκB signal response domain (SRD). Promonocytic cells stably overexpressing an IκBα transgene containing SRD mutations fail to activate NF-κB and subsequently fail to survive the PMA-induced macrophage differentiation program. The differentiation-induced apoptosis was found to be dependent on tumor necrosis factor alpha. The protective effect of NF-κB is mediated through p21WAF1/Cip1, since this protein was found to be regulated in an NF-κB-dependent manner and to confer survival features during macrophage differentiation. Therefore, NF-κB plays a key role in cell differentiation by conferring cell survival that in the case of macrophages is mediated through p21WAF1/Cip1.

scholarly journals A unique palindromic element mediates gamma interferon induction of mig gene expression.

1994 ◽  
Vol 14 (2) ◽  
pp. 914-922 ◽  
Author(s):  
P Wong ◽  
C W Severns ◽  
N B Guyer ◽  
T M Wright
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Regulatory Element ◽  
Regulation Of Gene Expression ◽  
Gamma Interferon ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Monocytic Cell ◽  
Platelet Factor ◽  
Ifn Gamma

To define the molecular mechanisms involved in the action of gamma interferon (IFN-gamma), we have analyzed the transcriptional regulation of the mig (monokine induced by gamma interferon) gene, a member of the platelet factor 4-interleukin-8 cytokine family that is expressed in murine macrophages specifically in response to IFN-gamma. Analysis of mig/CAT chimeric constructs transiently transfected into the RAW 264.7 mouse monocytic cell line revealed a unique IFN-gamma-responsive element (gamma RE-1). The sequence of this cis regulatory element defined by deletion analysis contains an imperfect inverted repeat extending 27 bp. Examination of mig/CAT constructs with mutations in gamma RE-1 revealed that the palindromic positions in the element were essential for activity. Consistent with its function as an enhancer, a single copy of gamma RE-1 conferred IFN-gamma inducibility to a heterologous (herpes simplex virus thymidine kinase) promoter. Exonuclease III protection assays demonstrated symmetrical protection of a mig promoter fragment centered about the gamma RE-1 palindromic sequence. Using the gel electrophoretic mobility shift assay, we identified a factor (gamma RF-1) present in nuclear extracts prepared from IFN-gamma-stimulated RAW 264.7 cells which binds to gamma RE-1. The activation of gamma RF-1 occurred rapidly (within 1 min) in response to IFN-gamma and was independent of protein synthesis. Similar to the expression of mig mRNA, the formation of gamma RF-1 was selectively induced by IFN-gamma and not IFN-alpha. The regulation of gene expression through gamma RF-1 and gamma RE-1 may explain the preferential activation of a subset of interferon-inducible genes by IFN-gamma.

scholarly journals Toxicogenomics in Drug Development

2003 ◽  
Vol 31 (5) ◽  
pp. 471-479 ◽  
Author(s):  
Nelson Guerreiro ◽  
Frank Staedtler ◽  
Olivier Grenet ◽  
Jeanne Kehren ◽  
Salah-Dine Chibout
Keyword(s):  
Gene Expression ◽  
Drug Development ◽  
Expression Profiling ◽  
Development Process ◽  
Molecular Mechanisms ◽  
Biomarker Discovery ◽  
Expression Profiles ◽  
Dna Polymorphisms ◽  
Drug Development Process ◽  
Predictive Toxicology

Toxicogenomics represents the merging of toxicology with technologies that have been developed, together with bioinformatics, to identify and quantify global gene expression changes. It represents a new paradigm in drug development and risk assessment, which promises to generate a wealth of information towards an increased understanding of the molecular mechanisms that lead to drug toxicity and efficacy, and of DNA polymorphisms responsible for individual susceptibility to toxicity. Gene expression profiling, through the use of DNA microarray and proteomic technologies will aid in establishing links between expression profiles, mode of action and traditional toxic endpoints. Such patterns of gene expression, or `molecular fingerprints' could be used as diagnostic or predictive markers of exposure, that is characteristic of a specific mechanism of induction of that toxic or efficacious effect. It is anticipated that toxicogenomics will be increasingly integrated into all phases of the drug development process particularly in mechanistic and predictive toxicology, and biomarker discovery. This review provides an overview of the expression profiling technologies applied in toxicogenomics, and discusses the promises as well as the future challenges of applying this discipline to the drug development process.

Molecular mechanisms of inhibitory activities of tanshinones on Lipopolysaccharide-lnduced nitric oxide generation in RAW 264.7 cells

2004 ◽  
Vol 27 (12) ◽  
pp. 1233-1237 ◽  
Author(s):  
Mong-Seok Choi ◽  
Dong-lm Cho ◽  
Hoo-Kyun Choi ◽  
Suhn Yong Im ◽  
Shi-Yong Ryu ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Molecular Mechanisms ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Inhibitory Activities

scholarly journals Characterization of βN-Octadecanoyl-5-hydroxytryptamide Anti-Inflammatory Effect

Molecules ◽  
2021 ◽  
Vol 26 (12) ◽  
pp. 3709
Author(s):  
Thais Biondino Sardella Giorno ◽  
Fernanda Alves Lima ◽  
Ana Laura Macedo Brand ◽  
Camila Martins de Oliveira ◽  
Claudia Moraes Rezende ◽  
...  
Keyword(s):  
Cell Types ◽  
In Vitro Assays ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Protein Extravasation ◽  
Anti Inflammatory ◽  
Different Cell Types

Background: N-octadecanoyl-5-hydroxytryptamide (C18-5HT) is an amide that can be obtained by the coupling of serotonin and octadecanoic acid. This study aims to characterize the in vivo and in vitro anti-inflammatory activity of C18-5HT. Methods: A subcutaneous air pouch model (SAP) was used. The exudates were collected from SAP after carrageenan injection to assess cell migration and inflammatory mediators production. RAW 264.7 cells were used for in vitro assays. Results: C18-5HT significantly inhibited leukocyte migration into the SAP as well as nitric oxide (NO) and cytokines production and protein extravasation. We also observed an reduction in some cytokines and an increase in IL-10 production. Assays conducted with RAW 264.7 cells indicated that C18-5HT inhibited NO and cytokine produced. Conclusions: Taken together, our data suggest that C18-5HT presents a significant effect in different cell types (leukocytes collected from exudate, mainly polumorphonuclear leukocytes and cell culture macrophages) and is a promising compound for further studies for the development of a new anti-inflammatory drug.

scholarly journals Anti-Inflammatory Effect of Rhapontici Radix Ethanol Extract via Inhibition of NF-κB and MAPK and Induction of HO-1 in Macrophages

2016 ◽  
Vol 2016 ◽  
pp. 1-13 ◽  
Author(s):  
Yun Hee Jeong ◽  
You-Chang Oh ◽  
Won-Kyung Cho ◽  
Nam-Hui Yim ◽  
Jin Yeul Ma
Keyword(s):  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Ethanol Extract ◽  
Inhibitory Effect ◽  
Peritoneal Macrophages ◽  
Biological Properties ◽  
Mapk Signaling ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Anti Inflammatory

Rhapontici Radix (RR) has been used in traditional medicine in East Asia and has been shown to have various beneficial effects. However, its biological properties or mechanism on inflammation-related diseases is unknown. The goal of this study was to determine the anti-inflammatory activity and underlying molecular mechanisms of Rhapontici Radix ethanol extract (RRE). The inhibitory effect of RRE on the production of NO, cytokines, inflammatory-related proteins, and mRNAs in LPS-stimulated macrophages was determined by the Griess assay, ELISA, Western blot analysis, and real-time RT-PCR, respectively. Our results indicate that treatment with RRE significantly inhibited the secretion of NO and inflammatory cytokines in RAW 264.7 cells and mouse peritoneal macrophages without cytotoxicity. We also found that RRE strongly suppressed the expression of iNOS and COX-2 and induced HO-1 expression. It also prevented nuclear translocation of NF-κB by inhibiting the phosphorylation and degradation of IκBα. Furthermore, the phosphorylation of MAPKs in LPS-stimulated RAW 264.7 cells was significantly inhibited by RRE. These findings suggest that RRE may operate as an effective anti-inflammatory agent by inhibiting the activation of NF-κB and MAPK signaling pathways and inducing HO-1 expression in macrophages. Our results suggest that RRE has potential value as candidate to inflammatory therapeutic phytomedicine.

scholarly journals A unique palindromic element mediates gamma interferon induction of mig gene expression

1994 ◽  
Vol 14 (2) ◽  
pp. 914-922
Author(s):  
P Wong ◽  
C W Severns ◽  
N B Guyer ◽  
T M Wright
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Regulatory Element ◽  
Regulation Of Gene Expression ◽  
Gamma Interferon ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Monocytic Cell ◽  
Platelet Factor ◽  
Ifn Gamma

To define the molecular mechanisms involved in the action of gamma interferon (IFN-gamma), we have analyzed the transcriptional regulation of the mig (monokine induced by gamma interferon) gene, a member of the platelet factor 4-interleukin-8 cytokine family that is expressed in murine macrophages specifically in response to IFN-gamma. Analysis of mig/CAT chimeric constructs transiently transfected into the RAW 264.7 mouse monocytic cell line revealed a unique IFN-gamma-responsive element (gamma RE-1). The sequence of this cis regulatory element defined by deletion analysis contains an imperfect inverted repeat extending 27 bp. Examination of mig/CAT constructs with mutations in gamma RE-1 revealed that the palindromic positions in the element were essential for activity. Consistent with its function as an enhancer, a single copy of gamma RE-1 conferred IFN-gamma inducibility to a heterologous (herpes simplex virus thymidine kinase) promoter. Exonuclease III protection assays demonstrated symmetrical protection of a mig promoter fragment centered about the gamma RE-1 palindromic sequence. Using the gel electrophoretic mobility shift assay, we identified a factor (gamma RF-1) present in nuclear extracts prepared from IFN-gamma-stimulated RAW 264.7 cells which binds to gamma RE-1. The activation of gamma RF-1 occurred rapidly (within 1 min) in response to IFN-gamma and was independent of protein synthesis. Similar to the expression of mig mRNA, the formation of gamma RF-1 was selectively induced by IFN-gamma and not IFN-alpha. The regulation of gene expression through gamma RF-1 and gamma RE-1 may explain the preferential activation of a subset of interferon-inducible genes by IFN-gamma.

scholarly journals Anti-Inflammatory Effects of Antarctic Lichen Umbilicaria antarctica Methanol Extract in Lipopolysaccharide-Stimulated RAW 264.7 Macrophage Cells and Zebrafish Model

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Ju-Mi Hong ◽  
Jung Eun Kim ◽  
Seul Ki Min ◽  
Kyung Hee Kim ◽  
Se Jong Han ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Protein Expression ◽  
Molecular Mechanisms ◽  
Inflammatory Responses ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Zebrafish Model ◽  
In Vivo Models ◽  
Mrna And Protein Expression ◽  
Anti Inflammatory

Umbilicaria antarctica (UA) is a member of the family Umbilicariaceae. To the best of our knowledge, no studies on its anti-inflammatory effects have been reported yet. In the present study, we examined its ability to suppress inflammatory responses and the molecular mechanisms underlying these abilities using lipopolysaccharide- (LPS-) stimulated RAW 264.7 cells and a zebrafish model of inflammation. We investigated the effects of UA on the production of nitric oxide (NO) and prostaglandin E2 (PGE2) in LPS-stimulated RAW 264.7 cells. To explore the anti-inflammatory mechanisms of UA, we measured the mRNA and protein expression of proinflammatory mediators in LPS-stimulated RAW 264.7 cells using quantitative RT-PCR and western blot analyses, respectively. UA significantly inhibited the production of NO, PGE2, interleukin- (IL-) 6, and tumor necrosis factor- (TNF-) α in the LPS-stimulated RAW 264.7 cells. It also suppressed the mRNA and protein expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and nuclear factor- (NF-) κB activation in LPS-stimulated RAW 264.7 cells and tail pin-cutting-induced zebrafish model. Collectively, these findings indicate that UA significantly inhibits LPS-stimulated inflammatory responses. These effects were considered to be strongly associated with the suppression of NF-κB activation. Overall, our results demonstrate that UA extract exerts strong anti-inflammatory activities in in vitro and in vivo models and suggest that UA may be an effective novel therapeutic agent for the treatment of inflammatory diseases.

scholarly journals Unraveling the complexity of thermogenic remodeling of white fat reveals potential antiobesity therapies

2021 ◽  
Vol 35 (21-22) ◽  
pp. 1395-1397
Author(s):  
Nabil Rabhi ◽  
Stephen R. Farmer
Keyword(s):  
Molecular Mechanisms ◽  
Cell Types ◽  
Cellular Interaction ◽  
Transcriptional Program ◽  
Previous Issue ◽  
Beige Adipocytes ◽  
Dietary Cues ◽  
Different Cell Types ◽  
White Fat ◽  
Complex Organ

Adipose tissue is a complex organ consisting of a mixture of mature adipocytes and stromal vascular cells. It displays a remarkable ability to adapt to environmental and dietary cues by changing its morphology and metabolic capacity. This plasticity is demonstrated by the emergence of interspersed thermogenic beige adipocytes within white depots in response to catecholamines secretion. Coordinated cellular interaction between different cell types within the tissue and a fine-tuned transcriptional program synergistically take place to promote beige remodeling. However, both cell–cell interactions and molecular mechanisms governing beige adipocyte appearance and maintenance are poorly understood. In this and the previous issue of Genes & Development, Shao and colleagues (pp. 1461–1474) and Shan and colleagues (pp. 1333–1338) advance our understanding of these issues and, in doing so, highlight potential therapeutic strategies to combat obesity-associated diseases.

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