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ROS/TNF-α Crosstalk Triggers the Expression of IL-8 and MCP-1 in Human Monocytic THP-1 Cells via the NF-κB and ERK1/2 Mediated Signaling.

ABSTRACT: IL-8/MCP-1 act as neutrophil/monocyte chemoattractants, respectively. Oxidative stress emerges as a key player in the pathophysiology of obesity. However, it remains unclear whether the TNF-α/oxidative stress interplay can trigger IL-8/MCP-1 expression and, if so, by which mechanism(s). IL-8/MCP-1 adipose expression was detected in lean, overweight, and obese individuals, 15 each, using immunohistochemistry. To detect the role of reactive oxygen species (ROS)/TNF-α synergy as a chemokine driver, THP-1 cells were stimulated with TNF-α, with/without H2O2 or hypoxia. Target gene expression was measured by qRT-PCR, proteins by flow cytometry/confocal microscopy, ROS by DCFH-DA assay, and signaling pathways by immunoblotting. IL-8/MCP-1 adipose expression was significantly higher in obese/overweight. Furthermore, IL-8/MCP-1 mRNA/protein was amplified in monocytic cells following stimulation with TNF-α in the presence of H2O2 or hypoxia (p ˂ 0.0001). Synergistic chemokine upregulation was related to the ROS levels, while pre-treatments with NAC suppressed this chemokine elevation (p ≤ 0.01). The ROS/TNF-α crosstalk involved upregulation of CHOP, ERN1, HIF1A, and NF-κB/ERK-1,2 mediated signaling. In conclusion, IL-8/MCP-1 adipose expression is elevated in obesity. Mechanistically, ROS/TNF-α crosstalk may drive expression of these chemokines in monocytic cells by inducing ER stress, HIF1A stabilization, and signaling via NF-κB/ERK-1,2. NAC had inhibitory effect on oxidative stress-driven IL-8/MCP-1 expression, which may have therapeutic significance regarding meta-inflammation.

SUBMITTER: Akhter N

PROVIDER: S-EPMC8508672 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

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ROS/TNF-α Crosstalk Triggers the Expression of IL-8 and MCP-1 in Human Monocytic THP-1 Cells via the NF-κB and ERK1/2 Mediated Signaling.

Akhter Nadeem N Wilson Ajit A Thomas Reeby R Al-Rashed Fatema F Kochumon Shihab S Al-Roub Areej A Arefanian Hossein H Al-Madhoun Ashraf A Al-Mulla Fahd F Ahmad Rasheed R Sindhu Sardar S

International journal of molecular sciences 20210929 19

IL-8/MCP-1 act as neutrophil/monocyte chemoattractants, respectively. Oxidative stress emerges as a key player in the pathophysiology of obesity. However, it remains unclear whether the TNF-α/oxidative stress interplay can trigger IL-8/MCP-1 expression and, if so, by which mechanism(s). IL-8/MCP-1 adipose expression was detected in lean, overweight, and obese individuals, 15 each, using immunohistochemistry. To detect the role of reactive oxygen species (ROS)/TNF-α synergy as a chemokine driver, ...[more]

PMID: 34638857

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Project description:Obesity and metabolic syndrome involve chronic low-grade inflammation called metabolic inflammation as well as metabolic derangements from increased endotoxin and free fatty acids. It is debated whether the endoplasmic reticulum (ER) stress in monocytic cells can contribute to amplify metabolic inflammation; if so, by which mechanism(s). To test this, metabolic stress was induced in THP-1 cells and primary human monocytes by treatments with lipopolysaccharide (LPS), palmitic acid (PA), or oleic acid (OA), in the presence or absence of the ER stressor thapsigargin (TG). Gene expression of tumor necrosis factor (TNF)-α and markers of ER/oxidative stress were determined by qRT-PCR, TNF-α protein by ELISA, reactive oxygen species (ROS) by DCFH-DA assay, hypoxia-inducible factor 1-alpha (HIF-1α), p38, extracellular signal-regulated kinase (ERK)-1,2, and nuclear factor kappa B (NF-κB) phosphorylation by immunoblotting, and insulin sensitivity by glucose-uptake assay. Regarding clinical analyses, adipose TNF-α was assessed using qRT-PCR/IHC and plasma TNF-α, high-sensitivity C-reactive protein (hs-CRP), malondialdehyde (MDA), and oxidized low-density lipoprotein (OX-LDL) via ELISA. We found that the cooperative interaction between metabolic and ER stresses promoted TNF-α, ROS, CCAAT-enhancer-binding protein homologous protein (CHOP), activating transcription factor 6 (ATF6), superoxide dismutase 2 (SOD2), and nuclear factor erythroid 2-related factor 2 (NRF2) expression (p ≤ 0.0183),. However, glucose uptake was not impaired. TNF-α amplification was dependent on HIF-1α stabilization and p38 MAPK/p65 NF-κB phosphorylation, while the MAPK/NF-κB pathway inhibitors and antioxidants/ROS scavengers such as curcumin, allopurinol, and apocynin attenuated the TNF-α production (p ≤ 0.05). Individuals with obesity displayed increased adipose TNF-α gene/protein expression as well as elevated plasma levels of TNF-α, CRP, MDA, and OX-LDL (p ≤ 0.05). Our findings support a metabolic-ER stress cooperativity model, favoring inflammation by triggering TNF-α production via the ROS/CHOP/HIF-1α and MAPK/NF-κB dependent mechanisms. This study also highlights the therapeutic potential of antioxidants in inflammatory conditions involving metabolic/ER stresses.

Project description:BackgroundAccelerated cellular senescence within the nucleus pulposus (NP) region is a common feature of disc degeneration. Our previous work indicated that TNF-α promoted NP cell senescence. Although the intervertebral disc has been reported to be an estrogen-sensitive tissue, it is unclear whether estrogen can inhibit premature senescence of NP cells.ObjectiveTo investigate whether 17beta-estradiol (E2) can attenuate TNF-α-induced premature senescence of NP cells and the potential mechanism behind this regulatory process.MethodsIsolated NP cells and intact intervertebral discs from healthy rats were cultured with or without TNF-α, E2 or their combination. The pan estrogen receptor (ER) antagonist ICI 182780 was used to investigate the role of ER. Direct and indirect indicators including cell proliferation, SA-β-Gal activity, telomerase activity, cell cycle, and the expression of matrix macromolecules (aggrecan and collagen II) and senescence markers (p16 and p53) were used to evaluate the premature senescence of NP cells. Additionally, intracellular reactive oxygen species (ROS) and NF-κB/p65 activity were also detected in the NP cell cultures.ResultsIn the NP cell cultures, E2 significantly increased cell proliferation potency, telomerase activity and the expression of matrix macromolecules but attenuated SA-β-Gal activity, senescence marker (p53 and p16) expression and G1 cycle arrest in TNF-α-treated NP cells. Furthermore, E2 inhibited ROS generation and phospho-NF-κB/p65 expression in the TNF-α-treated NP cells. However, the ER antagonist ICI 182780 abolished the effects of E2 on TNF-α-treated NP cells. In the disc organ cultures, E2 also significantly increased matrix synthesis, whereas it decreased senescence marker (p53 and p16) expression, which could be abolished by the ER antagonist ICI 182780.ConclusionThe interaction between E2 and ER can attenuate TNF-α-induced premature senescence of rat NP cells through interfering with the ROS/NF-κB pathway.

Dataset Information

ROS/TNF-α Crosstalk Triggers the Expression of IL-8 and MCP-1 in Human Monocytic THP-1 Cells via the NF-κB and ERK1/2 Mediated Signaling.

Publications

ROS/TNF-α Crosstalk Triggers the Expression of IL-8 and MCP-1 in Human Monocytic THP-1 Cells via the NF-κB and ERK1/2 Mediated Signaling.

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