Neutral sphingomyelinase 2 regulates inflammatory responses in monocytes/macrophages induced by TNF-α

ABSTRACTObesity is associated with elevated levels of TNF-α and proinflammatory CD11c monocytes /macrophages. TNF-α mediated dysregulation in the plasticity of monocytes/macrophages is concomitant with pathogenesis of several inflammatory diseases, including metabolic syndrome, but the underlying mechanisms are incompletely understood. Since neutral sphingomyelinase 2 (nSMase2; product of the sphingomyelin phosphodiesterase 3 gene, SMPD3) is a key enzyme for ceramide production involved in inflammation, we investigated whether nSMase2 contributed to the inflammatory changes in the monocytes/macrophages induced by TNF-α. In this study, we demonstrate that the disruption of nSMase activity in monocytes/macrophages either by chemical inhibitor GW4869 or small interfering RNA (siRNA) against SMPD3 results in defects in the TNF-α mediated expression of CD11c. Furthermore, blockage of nSMase in monocytes/macrophages inhibited the secretion of inflammatory mediators IL-1b and MCP-1. In contrast, inhibition of acid SMase (aSMase) activity did not attenuate CD11c expression or secretion of IL-1b and MCP-1. TNF-α-induced phosphorylation of JNK, p38 and NF-κB was also attenuated by the inhibition of nSMase2. Moreover, NF-kB/AP-1 activity was blocked by the inhibition of nSMase2. SMPD3 was elevated in PBMCs from obese individuals and positively corelated with TNF-α gene expression. These findings indicate that nSMase2 acts, at least in part, as a master switch in the TNF-α mediated inflammatory responses in monocytes/macrophages.

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Neutral sphingomyelinase 2 regulates inflammatory responses in monocytes/macrophages induced by TNF-α

Scientific Reports ◽

10.1038/s41598-020-73912-5 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Fatema Al-Rashed ◽

Zunair Ahmad ◽

Reeby Thomas ◽

Motasem Melhem ◽

Ashley J. Snider ◽

...

Keyword(s):

Small Interfering Rna ◽

Inflammatory Responses ◽

Inflammatory Diseases ◽

Neutral Sphingomyelinase ◽

Tnf Α ◽

Key Enzyme ◽

Underlying Mechanisms ◽

Interfering Rna ◽

Inflammatory Changes ◽

Ceramide Production

Abstract Obesity is associated with elevated levels of TNF-α and proinflammatory CD11c monocytes/macrophages. TNF-α mediated dysregulation in the plasticity of monocytes/macrophages is concomitant with pathogenesis of several inflammatory diseases, including metabolic syndrome, but the underlying mechanisms are incompletely understood. Since neutral sphingomyelinase-2 (nSMase2: SMPD3) is a key enzyme for ceramide production involved in inflammation, we investigated whether nSMase2 contributed to the inflammatory changes in the monocytes/macrophages induced by TNF-α. In this study, we demonstrate that the disruption of nSMase activity in monocytes/macrophages either by chemical inhibitor GW4869 or small interfering RNA (siRNA) against SMPD3 results in defects in the TNF-α mediated expression of CD11c. Furthermore, blockage of nSMase in monocytes/macrophages inhibited the secretion of inflammatory mediators IL-1β and MCP-1. In contrast, inhibition of acid SMase (aSMase) activity did not attenuate CD11c expression or secretion of IL-1β and MCP-1. TNF-α-induced phosphorylation of JNK, p38 and NF-κB was also attenuated by the inhibition of nSMase2. Moreover, NF-kB/AP-1 activity was blocked by the inhibition of nSMase2. SMPD3 was elevated in PBMCs from obese individuals and positively corelated with TNF-α gene expression. These findings indicate that nSMase2 acts, at least in part, as a master switch in the TNF-α mediated inflammatory responses in monocytes/macrophages.

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Ceramide Kinase Regulates TNF-α-induced Immune Responses in Human Monocytic Cells

10.21203/rs.3.rs-146950/v1 ◽

2021 ◽

Author(s):

Fatema Al-Rashed ◽

Zunair Ahmad ◽

Ashley J Snider ◽

Reeby Thomas ◽

Shihab Kochumon ◽

...

Keyword(s):

Inflammatory Responses ◽

Inflammatory Diseases ◽

Monocytic Cells ◽

Metabolic Inflammation ◽

Ceramide Kinase ◽

Hla Dr ◽

Tnf Α ◽

Underlying Mechanisms ◽

Interfering Rna ◽

Ceramide 1

Abstract Ceramide kinase (CERK) phosphorylates ceramide to produce ceramide-1-phosphate (C1P), which is involved in the development of metabolic inflammation. TNF-α modulates inflammatory responses in monocytes associated with various inflammatory disorders; however, the underlying mechanisms remain not fully understood. Here, we investigated the role of CERK in TNF-α-induced inflammatory responses in monocytes. Our results show that disruption of CERK activity in monocytes either by the chemical inhibitor NVP- 231 or by small interfering RNA (siRNA) results in the defective expression of inflammatory markers including CD11c, CD11b and HLA-DR in response to TNF-α. Our data show that TNF-α upregulates ceramide phosphorylation. Inhibition of CERK in monocytes significantly reduced the secretion of IL-1β and MCP-1. Similar results were observed in CERK deficient cells. Phosphorylation of JNK, p38 and NF-κB resulting from TNF-α stimulation was reduced by inhibition of CERK. Additionally, NF-κB/AP-1 activity was suppressed by the inhibition of CERK. Clinically, obese individuals had higher levels of CERK expression in PBMCs compared to lean individuals, which correlated with TNF-α levels. Taken together, these results suggest that CERK plays a key role in regulating inflammatory responses in human monocytes during TNF-α stimulation. CERK may be a relevant target for developing novel therapies for chronic inflammatory diseases.

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Ceramide kinase regulates TNF-α-induced immune responses in human monocytic cells

Scientific Reports ◽

10.1038/s41598-021-87795-7 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Fatema Al-Rashed ◽

Zunair Ahmad ◽

Ashley J. Snider ◽

Reeby Thomas ◽

Shihab Kochumon ◽

...

Keyword(s):

Inflammatory Responses ◽

Inflammatory Diseases ◽

Monocytic Cells ◽

Metabolic Inflammation ◽

Ceramide Kinase ◽

Hla Dr ◽

Tnf Α ◽

Underlying Mechanisms ◽

Interfering Rna ◽

Ceramide 1

AbstractCeramide kinase (CERK) phosphorylates ceramide to produce ceramide-1-phosphate (C1P), which is involved in the development of metabolic inflammation. TNF-α modulates inflammatory responses in monocytes associated with various inflammatory disorders; however, the underlying mechanisms remain not fully understood. Here, we investigated the role of CERK in TNF-α-induced inflammatory responses in monocytes. Our results show that disruption of CERK activity in monocytes, either by chemical inhibitor NVP-231 or by small interfering RNA (siRNA), results in the defective expression of inflammatory markers including CD11c, CD11b and HLA-DR in response to TNF-α. Our data show that TNF-α upregulates ceramide phosphorylation. Inhibition of CERK in monocytes significantly reduced the secretion of IL-1β and MCP-1. Similar results were observed in CERK-downregulated cells. TNF-α-induced phosphorylation of JNK, p38 and NF-κB was reduced by inhibition of CERK. Additionally, NF-κB/AP-1 activity was suppressed by the inhibition of CERK. Clinically, obese individuals had higher levels of CERK expression in PBMCs compared to lean individuals, which correlated with their TNF-α levels. Taken together, these results suggest that CERK plays a key role in regulating inflammatory responses in human monocytes during TNF-α stimulation. CERK may be a relevant target for developing novel therapies for chronic inflammatory diseases.

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A Steroidal Saponin RCE-4 Inhibits Lipopolysaccharide-Stimulated Inflammatory Responses via Blocking PI3K/Akt-Mediated Nf-κB Activation in RAW264.7 Cells

Applied Mechanics and Materials ◽

10.4028/www.scientific.net/amm.568-570.1901 ◽

2014 ◽

Vol 568-570 ◽

pp. 1901-1906 ◽

Cited By ~ 1

Author(s):

Cai Hong Bai ◽

Hai Bo He ◽

Fan Cheng ◽

Kun Zou ◽

Jun Zhi Wang ◽

...

Keyword(s):

Inflammatory Cytokines ◽

Molecular Basis ◽

Inflammatory Responses ◽

Antiinflammatory Activity ◽

Inflammatory Diseases ◽

Steroidal Saponin ◽

Promising Candidate ◽

Akt Phosphorylation ◽

Tnf Α ◽

Underlying Mechanisms

Steroidal saponin: (1β,3β,5β,25S)-spirostan-1,3-diol1-[α-L-rhamnopyranosyl-(1→2)-β-D-xylopyranoside] (RCE-4) is the most abundant and bioactive members in Reineckia carnea, has been reported to possess antiinflammatory activity, but the underlying mechanisms remain largely unknown. The present aim was to study expression of inflammatory cytokines, on the basis of this investigation, the possible mechanism of RCE-4 was elucidated. In the present study, we found that the concentrations of TNF-α, IL-1β and IL-6 released from LPS-stimulated RAW264.7cells significantly increased compared to control (P<0.01, respectively). After pretreatment with RCE-4, the TNF-α, IL-1β and IL-6 levels significantly decreased compared with the LPS group (P<0.05, P<0.01, respectively). Further studies indicated that RCE-4 significantly suppressed Akt phosphorylation and NF-қB activation, and with the dose of RCE-4 increasing; their improvement became more and more strong. Our results showed that RCE-4 inhibited LPS-stimiulated TNF-α, IL-1β, IL-6 productions through the blockage of PI3K/Akt-mediated NF-κB activation. Our findings might provide a molecular basis for the ability of RCE-4 serving as a promising candidate for treating various inflammatory diseases.

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Toll-Like Receptor 2 Recognition of the Microsporidia Encephalitozoon spp. Induces Nuclear Translocation of NF-κB and Subsequent Inflammatory Responses

Infection and Immunity ◽

10.1128/iai.00733-08 ◽

2008 ◽

Vol 76 (10) ◽

pp. 4737-4744 ◽

Cited By ~ 21

Author(s):

Jeffrey Fischer ◽

Colby Suire ◽

Hollie Hale-Donze

Keyword(s):

Nuclear Translocation ◽

Inflammatory Responses ◽

Host Recognition ◽

Direct Role ◽

Necrosis Factor Alpha ◽

Human Macrophages ◽

Encephalitozoon Intestinalis ◽

Intracellular Parasites ◽

Tnf Α ◽

Interfering Rna

ABSTRACT Microsporidia are obligate intracellular parasites that are ubiquitous in nature and have been recognized as causing an important emerging disease among immunocompromised individuals. Limited knowledge exists about the immune response against these organisms, and virtually nothing is known about the receptors involved in host recognition. Toll-like receptors (TLR) are pattern recognition receptors that bind to specific molecules found on pathogens and signal a variety of inflammatory responses. In this study, we show that both Encephalitozoon cuniculi and Encephalitozoon intestinalis are preferentially recognized by TLR2 and not by TLR4 in primary human macrophages. This is the first demonstration of host receptor recognition of any microsporidian species. TLR2 ligation is known to activate NF-κB, resulting in inflammatory cytokines, such as tumor necrosis factor alpha (TNF-α) and interleukin-8 (IL-8). We found that the infection of primary human macrophages leads to the nuclear translocation of NF-κB in as early as 1 h and the subsequent production of TNF-α and IL-8. To verify the direct role of TLR2 parasite recognition in the production of these cytokines, the receptor was knocked down in primary human macrophages using small interfering RNA. This knockdown resulted in decreases in both the nuclear translocation of NF-κB and the levels of TNF-α and IL-8 after challenge with spores. Taken together, these experiments directly link the initial inflammatory response induced by Encephalitozoon spp. to TLR2 stimulation in human macrophages.

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Dimethyl itaconate inhibits TNF-α induced NF-κB signaling pathway in human epithelial cells

10.21203/rs.2.17569/v2 ◽

2020 ◽

Author(s):

Yalei Zhang ◽

Xiaobing Deng ◽

Hao Liang ◽

Annan Guo ◽

Kenan Li ◽

...

Keyword(s):

Inflammatory Responses ◽

Inflammatory Diseases ◽

Cysteine Residue ◽

Luciferase Assay ◽

Dependent Manner ◽

Nuclear Entry ◽

Tnf Α ◽

Novel Strategy ◽

Dose Dependent ◽

Dimethyl Itaconate

Abstract Background: Dimethyl itaconate (DMI), a membrane-permeable derivative of itaconate, was found to moderate IL-17-IκBζ-induced skin pathology including psoriasis in mouse experiments . TNF-α induced NF-κB pathway, which controls a variety of immune and inflammatory responses, was also proven to play a crucial role as mediator in psoriasis. However, whether DMI interacts with the TNF-α induced NF-κB pathway remains unclear. Results: Here we show that DMI inhibits TNF-α induced NF-κB transcriptional activities in dose-dependent manner in several human cell lines using dual luciferase assay and blocks the NF-κB nuclear entry. Moreover, DMI potently inhibits IKKβ dependent phosphorylation and degradation of IκBα in TNF-α induced activation of NF-κB pathway. We also demonstrate that DMI covalently binds to cysteine residue in IKKβ, a key regulator in NF-κB pathway, to suppress IKKβ activation and inhibit the canonical NF-κB pathway. Conclusion Our study presents a new mechanism for DMI as an anti-inflammatory agent that may have therapeutic potentials in treating NF-κB related human inflammatory diseases. Our results also suggest that itaconate produced by endogenous IRG1 may regulate NF-κB at post translation modification level, and the IRG1-itaconate-NF-κB axis could be targeted as a novel strategy for the treatment of IRG1-NF-κB mediated diseases.

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Load Down-Regulates TNF-Alpha Induced Cartilage Degradation in Part Through NF-KB and P38 Pathways

ASME 2007 Summer Bioengineering Conference ◽

10.1115/sbc2007-176541 ◽

2007 ◽

Author(s):

Valerie M. Wolfe ◽

Seonghun Park ◽

Marjana Tomic ◽

Peter A. Torzilli ◽

C. T. Christopher Chen

Keyword(s):

Inflammatory Responses ◽

Inflammatory Diseases ◽

Interleukin 1 ◽

Tensile Load ◽

Cartilage Degradation ◽

Acute Injury ◽

Cyclic Compression ◽

Tnf Α ◽

Necrosis Factor ◽

Pro Inflammatory Cytokines

Pro-inflammatory cytokines, such as interleukin-1 (IL-1) and tumor necrosis factor (TNF), can induce cartilage degradation after acute injury or in inflammatory diseases [1,2,3,7]. The degradative events are coordinated through the elevation and activation of two classes of enzymes, namely matrix metalloproteinases (MMPs) and aggrecanases (ADAMTS-4 and −5) [1,6]. Prior studies suggested that pro-inflammatory responses induced by IL-1β can be inhibited by tensile load [2] and more recently by cyclic compression [8]. It is, however, not clear whether load affects other cytokines, such as TNF-α. TNF-α is known to bind its receptor (TNFR1) to cause a cascade that ends with degradation of an inhibitor, IκBα, and release of the transcription factor NF-κB [3]. The actions of TNF-α are also known to be affected by at least three NF-κB independent pathways including the p38, ERK, and JNK pathways [4]. The objective of this study was to determine whether cyclic compression could affect TNF-α induced cartilage degradation and to determine the roles of p38, ERK, and JNK pathways in TNF-induced cartilage degradation. We hypothesized that cyclic loading would inhibit the degradative effects caused by TNF-α.

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Suppression of PPAR-γ attenuates insulin-stimulated glucose uptake by affecting both GLUT1 and GLUT4 in 3T3-L1 adipocytes

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00695.2006 ◽

2007 ◽

Vol 293 (1) ◽

pp. E219-E227 ◽

Cited By ~ 73

Author(s):

Wei Liao ◽

M. T. Audrey Nguyen ◽

Takeshi Yoshizaki ◽

Svetlana Favelyukis ◽

David Patsouris ◽

...

Keyword(s):

Glucose Uptake ◽

Insulin Signaling ◽

Adipocyte Differentiation ◽

Inflammatory Responses ◽

Critical Role ◽

Peroxisome Proliferator ◽

Peroxisome Proliferator Activated Receptor ◽

Ppar Γ ◽

Tnf Α ◽

Interfering Rna

Peroxisome proliferator-activated receptor-γ (PPAR-γ) plays a critical role in regulating insulin sensitivity and glucose homeostasis. In this study, we identified highly efficient small interfering RNA (siRNA) sequences and used lentiviral short hairpin RNA and electroporation of siRNAs to deplete PPAR-γ from 3T3-L1 adipocytes to elucidate its role in adipogenesis and insulin signaling. We show that PPAR-γ knockdown prevented adipocyte differentiation but was not required for maintenance of the adipocyte differentiation state after the cells had undergone adipogenesis. We further demonstrate that PPAR-γ suppression reduced insulin-stimulated glucose uptake without affecting the early insulin signaling steps in the adipocytes. Using dual siRNA strategies, we show that this effect of PPAR-γ deletion was mediated by both GLUT4 and GLUT1. Interestingly, PPAR-γ-depleted cells displayed enhanced inflammatory responses to TNF-α stimulation, consistent with a chronic anti-inflammatory effect of endogenous PPAR-γ. In summary, 1) PPAR-γ is essential for the process of adipocyte differentiation but is less necessary for maintenance of the differentiated state, 2) PPAR-γ supports normal insulin-stimulated glucose transport, and 3) endogenous PPAR-γ may play a role in suppression of the inflammatory pathway in 3T3-L1 cells.

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Paroxetine suppresses reactive microglia-mediated but not lipopolysaccharide-induced inflammatory responses in primary astrocytes

10.21203/rs.2.17327/v1 ◽

2019 ◽

Author(s):

Xiong Zhang ◽

Lan-Bing Zhu ◽

Jia-Hui He ◽

Hong-Qiu Zhang ◽

Shu-Ya Ji ◽

...

Keyword(s):

Inflammatory Responses ◽

Therapeutic Potential ◽

Minor Amount ◽

Innate Immune Responses ◽

Reactive Microglia ◽

Tnf Α ◽

Primary Astrocytes ◽

Underlying Mechanisms ◽

The Impact

Abstract Background Astrocytes are the most abundant glial cells in a brain that mediate inflammatory responses and provide trophic support for neurons. We have previously disclosed that paroxetine, a common selective serotonin reuptake inhibitor, ameliorates LPS-induced microglia activation. However, it remains elusive of the role of paroxetine in astrocytic responses. Methods Isolated primary astrocytes were pretreated with paroxetine and stimulated with different stimuli, lipopolysaccharide (LPS) or microglia conditioned medium pre-activated with LPS (M/Lps). Inflammatory and neurotrophic responses, underlying mechanisms and the impact on neuronal survival were assessed. Results Paroxetine had no impact on LPS-stimulated iNOS, TNF-α and IL-1β expression, but inhibited M/Lps-induced TNF-α and IL-1β expression in primary astrocytes. Paroxetine suppressed M/Lps- but not LPS-induced activation of NF-κB and had no impact on activation of MAPKs and STAT3. Incubation with the resulted astrocyte conditioned media caused no change in viability of SH-SY5Y cells. BDNF and MANF mRNA expressions were upregulated by M/Lps and paroxetine, respectively. However, M/Lps- or LPS-induced extracellular releases of NO, TNF-α and/or BDNF in astrocytes were in minor amount compared to those by microglia. Conclusions Paroxetine ameliorates the reactive microglia-mediated inflammatory responses in astrocytes partially via inhibition of NF-κB pathway, but has no impact on LPS-stimulated astrocyte activation. While the secondary astrocytic responses are not robust compared to the innate immune responses of microglia, our results support a therapeutic potential of paroxetine against neuroinflammation-associated neurological disorders such as Parkinson’s disease.

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Shikonin ameliorates Experimental Autoimmune Encephalomyelitis (EAE) via Immuno-modulatory, anti-apototic, and anti-oxidative activity

10.21203/rs.3.rs-29517/v1 ◽

2020 ◽

Author(s):

Mehrdad Nasrollahzadeh Sabet ◽

Sajjad Biglari ◽

Emran Esmaeilzadeh

Keyword(s):

Multiple Sclerosis ◽

Experimental Autoimmune Encephalomyelitis ◽

Inflammatory Diseases ◽

Expression Level ◽

Autoimmune Encephalomyelitis ◽

Glutathione Peroxidase 1 ◽

Tnf Α ◽

Ifn Γ ◽

Underlying Mechanisms ◽

Experimental Autoimmune

Abstract Background Multiple sclerosis is a common auto-immuno-inflammatory diseases of the central nervous system in adults. There are several underlying mechanisms for pathogenesis of the disease, including inflammation, oligodendrocyte apoptosis, and oxidative stress. Methods We have investigated the mechanism of Shikonin action in C57BL/6 experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis. Results Our results revealed that EAE induction significantly increased the extent of demyelination in the corpus callosum tissues of the animals, while treatment of the mice with Shikonin, significantly decreased the extent of demyelination. Real-time PCR based analyzing the brain samples from the EAE mice, revealed a significant enhancement in the expression level of TNF-α , IFN-γ and Bax genes as well as a reduction in the expression level of TGF-β and Bcl2. Shikonin treatment significantly reduced the expression level of TNF-α , IFN-γ and Bax. On the other hand, the expression levels of TGF-β and Bcl2 as well as the Glutathione peroxidase-1 (GPX-1) enzyme were significantly increased following Shikonin treatment. Conclusion This study emphasizes the immune-modulatory, anti-apoptotic, and anti-oxitive effects of Shikonin, which may have an important healing influence on the severity of EAE.

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