Pam3CSK4 Induces MMP-9 Expression in Human Monocytic THP-1 Cells

Al-Rashed, Fatema; Kochumon, Shihab; Usmani, Safa; Sindhu, Sardar; Ahmad, Rasheed

doi:10.1159/000475298

Cited by 30 publications

(29 citation statements)

References 40 publications

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“…ERK 1/2and JNK can be activated by a variety of stimuli in various types of cells [26]. Our findings are consistent with our previous studies showing that activation of MAPKs pathways, except MAPK p38, is important for the production of MMP-9 [10, 14]. We also identified that inhibition of NF-kB pathways significantly suppresses the production of CCL4.…”

Section: Discussionsupporting

confidence: 92%

Palmitate Activates CCL4 Expression in Human Monocytic Cells via TLR4/MyD88 Dependent Activation of NF-κB/MAPK/ PI3K Signaling Systems

Kochumon

Wilson

Chandy

et al. 2018

Cell Physiol Biochem

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Background/Aims: Obesity is associated with adipose tissue inflammation which plays a key role in the development of insulin resistance and type 2 diabetes (T2D). Saturated free fatty acids (SFAs) levels are found to be elevated in obesity and T2D. Chemokines are known to have potent inflammatory functions in a wide range of biological processes linked to immunological disorders. Since CCL4 (Chemokine (C-C motif) ligand 4), also known as macrophage inflammatory protein-1β (MIP-1β), plays an important role in the migration of monocytes into the adipose tissue, we investigated the expression of CCL4 in monocytic cells/macrophages following activation with free fatty acid palmitate. Methods: Human monocytic cell line THP-1 and macrophages derived from THP-1 and primary monocytes were stimulated with palmitate and LPS (positive control). CCL4 expression and secretion were measured with real time RT-PCR and ELISA respectively. Signaling pathways were identified by using THP-1-XBlue^TM cells, THP-1-XBlue^TM-defMyD cells, anti-TLR4 mAb and TLR4 siRNA. Results: Palmitate induces CCL4 expression at both mRNA and protein levels in human monocytic cells. Palmitate-induced CCL4 production was markedly suppressed by neutralizing anti-TLR-4 antibody. Additionally, silencing of TLR4 by siRNA also significantly suppressed the palmitate-induced up-regulation of CCL4. MyD88-deficient cells did not express CCL4 in response to palmitate treatment. Inhibition of NF-kB and MAPK pathways suppressed the palmitate mediated induction of CCL4. Moreover, induction of CCL4 was blocked by PI3 Kinase inhibitors LY294002 and wortmannin. Conclusion: Collectively, our results show that palmitate induces CCL4 expression via activation of the TLR4-MyD88/NF-kB/MAPK/ PI3K signaling cascade. Thus, our findings suggest that the palmitate-induced CCL4 production might be an underlying mechanism of metabolic inflammation.

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Section: Discussionsupporting

confidence: 92%

Palmitate Activates CCL4 Expression in Human Monocytic Cells via TLR4/MyD88 Dependent Activation of NF-κB/MAPK/ PI3K Signaling Systems

Kochumon

Wilson

Chandy

et al. 2018

Cell Physiol Biochem

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“…Each reaction contained 50 ng cDNA that was amplified with Inventoried TaqMan Gene Expression Assay products (SMPD3: Assay ID:Hs00920354_m1; SMPD2:Assay ID: Hs00906924_g1; SMPD1:Assay ID: Hs03679346_g1; CD11c: Assay ID: Hs00174217_m1; GAPDH: Hs03929097_g1;). The threshold cycle (Ct) values were normalized to the house-keeping gene GAPDH, and the amounts of target mRNA relative to control were calculated with ΔΔCt-method 35,36 . Relative mRNA expression was expressed as fold expression over average of control gene expression.…”

Section: Methodsmentioning

confidence: 99%

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

Al-Rashed

Ahmad

Thomas

et al. 2020

Sci Rep

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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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“…Each reaction contained 500 ng of cDNA that was amplified with Inventoried TaqMan Gene Expression Assay products (CSF2: Hs00929873; ACSL1: Hs00960561; GAPDH: Hs03929097_g1). The threshold cycle (Ct) values were normalized to the housekeeping gene GAPDH, and the amounts of target mRNA relative to control were calculated with the ΔΔCt method [ 25 , 26 ]. Relative mRNA expression was expressed as fold expression over average of control gene expression.…”

Section: Methodsmentioning

confidence: 99%

“…MDA-MB-231 cells were treated with LPS and incubated for 30 min with lysis buffer (10X Lysis Buffer, Cell Signaling, Danvers, MA, USA). The protein lysates were prepared and resolved by 12% SDS-PAGE, as described earlier [ 26 ]. Cellular proteins were transferred to an Immuno-Blot polyvinylidene difluoride (PVDF) membrane (Bio-Rad Laboratories, Hercules, CA, USA) by electroblotting.…”

Section: Methodsmentioning

confidence: 99%

LPS Induces GM-CSF Production by Breast Cancer MDA-MB-231 Cells via Long-Chain Acyl-CoA Synthetase 1

et al. 2020

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Granulocyte–macrophage colony-stimulating factor (GM-CSF) is a monomeric glycoprotein that has been implicated in the tumor growth and progression of different types of cancer. GM-CSF is produced by various non-immune cells including MDA-MB-231 in response to various stimuli. However, the role of lipopolysaccharide (LPS) in the regulation of GM-CSF in MDA-MB-231 breast cancer cells so far remains unclear. Herein, we asked whether LPS could induce GM-CSF production in MDA-MB-231 cells, and if so, which signaling pathway was involved. MDA-MB-231 cells were treated with LPS or tumor necrosis factor alpha (TNF-α; positive control), and GM-CSF expression levels were determined by qRT-PCR, ELISA, and confocal microscopy. Phosphorylation of the mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-kB) signaling proteins were evaluated by flow cytometry. Our results show that LPS induces GM-CSF expression at both mRNA and protein levels in MDA-MBA-231 cells. Inhibition of acyl-CoA synthetase 1 (ACSL1) activity in the cells with triacsin C significantly reduces the secretion of GM-CSF. Furthermore, the inhibition of ACSL1 activity significantly blocks the LPS-mediated phosphorylation of p38 MAPK, MEK1/2, extracellular signal-regulated kinase (ERK)1/2, c-Jun NH2-terminal kinase (JNK), and nuclear factor-κB (NF-kB) in the cells. These findings provide the first evidence that LPS induces ACSL1-dependent GM-CSF gene expression in MDA-MB-231 breast cancer cells, which requires the activation of p38 MAPK, MEK1/2, ERK1/2, JNK, and NF-kB.

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Pam3CSK4 Induces MMP-9 Expression in Human Monocytic THP-1 Cells

Cited by 30 publications

References 40 publications

Palmitate Activates CCL4 Expression in Human Monocytic Cells via TLR4/MyD88 Dependent Activation of NF-κB/MAPK/ PI3K Signaling Systems

Palmitate Activates CCL4 Expression in Human Monocytic Cells via TLR4/MyD88 Dependent Activation of NF-κB/MAPK/ PI3K Signaling Systems

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

LPS Induces GM-CSF Production by Breast Cancer MDA-MB-231 Cells via Long-Chain Acyl-CoA Synthetase 1

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