TNF-± Induces TSLP Expression By Human Lung Fibroblasts In An AP-1 / JNK Dependent Manner

Platelet-activating factor (PAF) is a potent proinflammatory phospholipid mediator of the lung. In this study, we demonstrate that PAF receptor mRNA and protein is expressed by human lung fibroblasts. Interaction of PAF with its specific receptor resulted in increases of tyrosine phosphorylation of several intracellular proteins, indicating that the PAF-receptor might be functionally active. PAF-induced transcription of protooncogenes c-fos and c-jun as well as of interleukin (IL)-6 and IL-8 genes in human fibroblasts. Transcription of the interleukins was followed by secretion of the respective proteins. Moreover, PAF enhanced proliferation of fibroblasts in a concentration-dependent manner. Using signaling inhibitors, we demonstrate that PAF-induced transcription of the c-fos, IL-6, and IL-8 genes, as well as proliferation, require activation of pertussis toxin-sensitive G proteins, tyrosine kinases, and protein kinase C (PKC). In contrast, transcription of c-jun was blocked by pertussis toxin, but not by inhibitors for tyrosine kinases or PKC. These data suggest that PAF stimulates distinct signaling pathways in human lung fibroblasts. In addition, the activation of human fibroblasts by PAF leads to enhanced proliferation and to the expression of proinflammatory cytokines, which may contribute to the pathophysiological changes in pulmonary inflammation.

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Differential regulation of metalloproteinase production, proliferation and chemotaxis of human lung fibroblasts by PDGF, Interleukin-1β and TNF-α

Mediators of Inflammation ◽

10.1080/09629350020002895 ◽

2000 ◽

Vol 9 (3-4) ◽

pp. 155-160 ◽

Cited By ~ 64

Author(s):

Masahiro Sasaki ◽

Masayuki Kashima ◽

Takefumi Ito ◽

Akiko Watanabe ◽

Noriko Izumiyama ◽

...

Keyword(s):

Matrix Metalloproteinase ◽

Human Lung ◽

Lung Fibroblast ◽

Differential Sensitivity ◽

Lung Fibroblasts ◽

Type I ◽

Dependent Manner ◽

Human Lung Fibroblast ◽

Human Lung Fibroblasts ◽

Tnf Α

Fibroblast migration, proliferation, extracellular matrix protein synthesis and degradation,all of which play important roles in inflammation, are them selves induced by various growth factors and cytokines. Less is known about the interaction of these substances on lung fibroblast function in pulmonary fibrosis.The goal of this study was to investigate the effects of PDGF alone and in combination with IL–1β and TNF–α on the production of human lung fibroblast matrix metalloproteinases, proliferation, and the chemotactic response. The assay for MMPs activity against FITC labeled type I and IV collagen was based on the specificity of the enzyme cleavage of collagen. Caseinolytis and gelatinolytic activities of secreted proteinases were analyzed by zymography. Fibronectin in conditioned media was measured using human lung fibronectin enzyme immunoassay. Cell proliferation was measured by 3H-Thymidine incorporation assay. Cell culture supernatants were tested for PGE2 content by ELISA. Chemotactic activity was measured using the modified Boyden chamber.Matrix metalloproteinase assay indicated that IL–1β, TNF–α and PDGF induced intestitial collagenase (MMP-1) production. MMP assay also indicated that IL–1β and TNF–α had inhibitory effects on MMP-2,9(gelatinaseA,B) production. Casein zymography confirmed that IL–1β stimulated stromlysin (matrix metalloproteinase 3; MMP–3) and gelatin zymography demonstrated that TNF–α induced MMP–9 production in human lung fibroblast, whereas PDGF alone did not. PDGF in combination with IL–1β and TNF–α induced MMP–3 and MMP–9 activity, as demonstrated by zymography. PDGF stimulated lung fibroblast proliferation in a concentration-dependent manner, whereas IL–1β and TNF–α alone had no effect. In contrast, the proliferation of human lung fibroblasts by PDGF was inhibited in the presence of IL–1β and TNF–α, and this inhibition was not a consequence of any elevation of PGE2. PDGF stimulated fibroblast chemotaxis in a concentrationdependent manner, and this stimulation was augmented by combining PDGF with IL–1β and TNF–α.These findings suggested that PDGF differentially regulated MMPs production in combination with cytokines, and further that MMP assay and zymography had differential sensitivity for detecting MMPs. The presence of cytokines with PDGF appears to modulate the proliferation and chemotaxis of human lung fibroblasts.

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Amorphous Silica Nanoparticles Obtained by Laser Ablation Induce Inflammatory Response in Human Lung Fibroblasts

Materials ◽

10.3390/ma12071026 ◽

2019 ◽

Vol 12 (7) ◽

pp. 1026 ◽

Cited By ~ 4

Author(s):

Sorina Voicu ◽

Mihaela Balas ◽

Miruna Stan ◽

Bogdan Trică ◽

Andreea Serban ◽

...

Keyword(s):

Inflammatory Response ◽

Silica Nanoparticles ◽

Human Lung ◽

Lung Fibroblasts ◽

No Production ◽

Dependent Manner ◽

Amorphous Sio2 ◽

Human Lung Fibroblasts ◽

Transmission Electron ◽

Cox 2

Silica nanoparticles (SiO2 NPs) represent environmentally born nanomaterials that are used in multiple biomedical applications. Our aim was to study the amorphous SiO2 NP-induced inflammatory response in MRC-5 human lung fibroblasts up to 72 hours of exposure. The intracellular distribution of SiO2 NPs was measured by transmission electron microscopy (TEM). The lactate dehydrogenase (LDH) test was used for cellular viability evaluation. We have also investigated the lysosomes formation, protein expression of interleukins (IL-1β, IL-2, IL-6, IL-8, and IL-18), COX-2, Nrf2, TNF-α, and nitric oxide (NO) production. Our results showed that the level of lysosomes increased in time after exposure to the SiO2 NPs. The expressions of interleukins and COX-2 were upregulated, whereas the expressions and activities of MMP-2 and MMP-9 decreased in a time-dependent manner. Our findings demonstrated that the exposure of MRC-5 cells to 62.5 µg/mL of SiO2 NPs induced an inflammatory response.

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Prostacyclin analogs stimulate VEGF production from human lung fibroblasts in culture

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00129.2007 ◽

2008 ◽

Vol 294 (6) ◽

pp. L1226-L1232 ◽

Cited By ~ 26

Author(s):

Koichiro Kamio ◽

Tadashi Sato ◽

Xiangde Liu ◽

Hisatoshi Sugiura ◽

Shinsaku Togo ◽

...

Keyword(s):

Human Lung ◽

Mesenchymal Cells ◽

Lung Fibroblasts ◽

Intracellular Camp ◽

Alveolar Wall ◽

Mrna Levels ◽

Dependent Manner ◽

Collagen Gels ◽

Human Lung Fibroblasts ◽

Cell Functions

Prostacyclin is a short-lived metabolite of arachidonic acid that is produced by several cells in the lung and prominently by endothelial cells. It increases intracellular cAMP levels activating downstream signaling thus regulating vascular mesenchymal cell functions. The alveolar wall contains a rich capillary network as well as a population of mesenchymal cells, i.e., fibroblasts. The current study evaluated the hypothesis that prostacyclin may mediate signaling between endothelial and mesenchymal cells in the alveolar wall by assessing the ability of prostacyclin analogs to modulate fibroblast release of VEGF. To accomplish this study, human lung fibroblasts were cultured in routine culture on plastic support and in three-dimensional collagen gels with or without three prostacyclin analogs, carbaprostacyclin, iloprost, and beraprost, and the production of VEGF was evaluated by ELISA and quantitative real-time PCR. Iloprost and beraprost significantly stimulated VEGF mRNA levels and protein release in a concentration-dependent manner. These effects were blocked by the adenylate cyclase inhibitor SQ-22536 and by the protein kinase A (PKA) inhibitor KT-5720 and were reproduced by a direct PKA activator but not by an activator of exchange protein directly activated by cAMP (Epac), indicating that cAMP-activated PKA signaling mediated the effect. Since VEGF serves to maintain the pulmonary microvasculature, the current study suggests that prostacyclin is part of a bidirectional signaling network between the mesenchymal and vascular cells of the alveolar wall. Prostacyclin analogs, therefore, have the potential to modulate the maintenance of the pulmonary microcirculation by driving the production of VEGF from lung fibroblasts.

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Human lung fibroblasts release chemokinetic activity for monocytes constitutively

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1998.275.2.l223 ◽

1998 ◽

Vol 275 (2) ◽

pp. L223-L230 ◽

Cited By ~ 6

Author(s):

Sekiya Koyama ◽

Etsuro Sato ◽

Tsuyoshi Masubuchi ◽

Akemi Takamizawa ◽

Hiroshi Nomura ◽

...

Keyword(s):

Molecular Weight ◽

Molecular Sieve ◽

Transforming Growth Factor ◽

Human Lung ◽

Lung Fibroblasts ◽

Dependent Manner ◽

Lipoxygenase Inhibitors ◽

Human Lung Fibroblasts ◽

Gm Csf ◽

Molecular Sieve Column

We determined whether human lung fibroblasts (HLFs) might release mediators that are responsible for monocyte chemokinetic activity (MCA) constitutively. HLF supernatant fluids showed MCA in a time-dependent manner ( P < 0.001). Checkerboard analysis of 24- and 72-h supernatant fluids showed that the activity was chemokinetic. Partial characterization of 24- and 72-h supernatant fluids revealed that the mediators released after 24 h were predominantly composed of lipid-soluble activity, and MCA was blocked by lipoxygenase inhibitors. The mediators released after 72 h were predominantly trypsin sensitive and blocked by cycloheximide. Molecular-sieve column chromatography identified four peaks of MCA. A polyclonal antibody to monocyte chemoattractant protein-1 (MCP-1) inhibited MCA by 20% after 24 h and by 40% after 72 h. Granulocyte-macrophage colony-stimulating factor (GM-CSF) and transforming growth factor-β (TGF-β) antibodies attenuated MCA released after 72 h by 30 and 10%, respectively. These antibodies inhibited corresponding molecular-weight peaks separated by molecular-sieve column. The concentrations of MCP-1, GM-CSF, and TGF-β were 4,698 ± 242, 26.8 ± 3.8, and 550 ± 15 pg/ml, respectively. A leukotriene B4(LTB4)-receptor antagonist attenuated the total MCA and the lowest molecular weight peak of MCA. The concentrations of LTB4were 153.4 ± 12.4 (24 h) and 212 ± 16.6 (72 h) pg/ml. These findings suggest that HLFs may modulate the recruitment of monocytes into the lung by releasing MCP-1, GM-CSF, TGF-β, and LTB4constitutively.

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Pharmacologic inhibition of lactate production prevents myofibroblast differentiation

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00058.2015 ◽

2015 ◽

Vol 309 (11) ◽

pp. L1305-L1312 ◽

Cited By ~ 20

Author(s):

Robert Matthew Kottmann ◽

Emma Trawick ◽

Jennifer L. Judge ◽

Lindsay A. Wahl ◽

Amali P. Epa ◽

...

Keyword(s):

Lactic Acid ◽

Pulmonary Fibrosis ◽

Human Lung ◽

Lung Fibroblasts ◽

Myofibroblast Differentiation ◽

Dependent Manner ◽

Human Lung Fibroblasts ◽

Ldh Activity ◽

Dose Dependent ◽

Pharmacologic Inhibition

Myofibroblasts are one of the primary cell types responsible for the accumulation of extracellular matrix in fibrosing diseases, and targeting myofibroblast differentiation is an important therapeutic strategy for the treatment of pulmonary fibrosis. Transforming growth factor-β (TGF-β) has been shown to be an important inducer of myofibroblast differentiation. We previously demonstrated that lactate dehydrogenase and its metabolic product lactic acid are important mediators of myofibroblast differentiation, via acid-induced activation of latent TGF-β. Here we explore whether pharmacologic inhibition of LDH activity can prevent TGF-β-induced myofibroblast differentiation. Primary human lung fibroblasts from healthy patients and those with pulmonary fibrosis were treated with TGF-β and or gossypol, an LDH inhibitor. Protein and RNA were analyzed for markers of myofibroblast differentiation and extracellular matrix generation. Gossypol inhibited TGF-β-induced expression of the myofibroblast marker α-smooth muscle actin (α-SMA) in a dose-dependent manner in both healthy and fibrotic human lung fibroblasts. Gossypol also inhibited expression of collagen 1, collagen 3, and fibronectin. Gossypol inhibited LDH activity, the generation of extracellular lactic acid, and the rate of extracellular acidification in a dose-dependent manner. Furthermore, gossypol inhibited TGF-β bioactivity in a dose-dependent manner. Concurrent treatment with an LDH siRNA increased the ability of gossypol to inhibit TGF-β-induced myofibroblast differentiation. Gossypol inhibits TGF-β-induced myofibroblast differentiation through inhibition of LDH, inhibition of extracellular accumulation of lactic acid, and inhibition of TGF-β bioactivity. These data support the hypothesis that pharmacologic inhibition of LDH may play an important role in the treatment of pulmonary fibrosis.

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Immune mediators increase adherence of T-lymphocytes to human lung fibroblasts

AJP Cell Physiology ◽

10.1152/ajpcell.1989.256.2.c336 ◽

1989 ◽

Vol 256 (2) ◽

pp. C336-C340 ◽

Cited By ~ 6

Author(s):

F. Hampson ◽

M. Monick ◽

M. W. Peterson ◽

G. W. Hunninghake

Keyword(s):

T Lymphocytes ◽

Human Lung ◽

Interleukin 1 ◽

Interleukin 2 ◽

Pulmonary Sarcoidosis ◽

Lung Fibroblasts ◽

Dependent Manner ◽

Human Lung Fibroblasts ◽

Immune Mediators ◽

Necrosis Factor

The purpose of the present study was to determine whether immunological mediators that are known to be released in the lungs of patients with active sarcoidosis might increase the adherence of human T-lymphocytes to human lung fibroblasts. The studies demonstrate that interleukin 1 (IL-1), tumor necrosis factor (TNF), gamma-interferon (IFN), and interleukin 2 (IL-2) increase the adherence of T-lymphocytes to fibroblasts in a dose- and time-dependent manner. An effect of IL-1, TNF, and IFN was observed at concentrations less than 1 ng/ml; an effect of IL-2 was not observed unless greater than or equal to 60 ng/ml of the mediator was used. The effect of the mediators was primarily on the fibroblasts; however, a significant increase in adherence was also observed when the T-lymphocytes were preincubated with the mediators. These observations suggest that there may be an intimate relationship between the immune response and the fibrotic response in the lungs of patients with pulmonary sarcoidosis or in other disorders where tissue fibrosis is mediated by immunological processes.

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HGF synthesis in human lung fibroblasts is regulated by oncostatin M

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00166.2005 ◽

2006 ◽

Vol 290 (6) ◽

pp. L1097-L1103 ◽

Cited By ~ 14

Author(s):

Murielle Cohen ◽

Sylvain Marchand-Adam ◽

Véronique Lecon-Malas ◽

Joëlle Marchal-Somme ◽

Anne Boutten ◽

...

Keyword(s):

Wound Repair ◽

Human Lung ◽

De Novo ◽

Mapk Pathway ◽

Oncostatin M ◽

Lung Fibroblasts ◽

Prostaglandin E ◽

Dependent Manner ◽

Human Lung Fibroblasts ◽

P38 Mapk Inhibitor

Oncostatin M (OSM) is a IL-6 family cytokine locally produced in acute lung injury. Its profibrotic properties suggest a role in lung wound repair. Hepatocyte growth factor (HGF), produced by fibroblasts, is involved in pulmonary epithelial repair. We investigated the role of OSM in HGF synthesis by human lung fibroblasts. We showed that OSM upregulated HGF mRNA in MRC5 cells and in human lung fibroblasts, whereas IL-6 and leukemia inhibitory factor did not. OSM induced HGF secretion to a similar extent as IL-1β in both a time- and dose-dependent manner. HGF was released in its cleaved mature form, and its secretion was completely inhibited in the presence of cycloheximide, indicating a de novo protein synthesis. OSM in combination with prostaglandin E2, a powerful HGF inductor, led to an additive effect. OSM and indomethacin in combination further increased HGF secretion. This could be explained, at least in part, by a moderate upregulation of specific OSM receptor β mRNA expression through cyclooxygenase inhibition.These results demonstrate that OSM-induced HGF synthesis did not involve a PGE2pathway. OSM-induced HGF secretion was inhibited by PD-98059 (a specific pharmacological inhibitor of ERK1/2), SB-203580 (a p38 MAPK inhibitor), and SP-600125 (a JNK inhibitor) by 70, 82, and 100%, respectively, whereas basal HGF secretion was only inhibited by SP-600125 by 30%. Our results demonstrate a specific upregulation of HGF synthesis by OSM, most likely through a MAPK pathway, and support the suggestion that OSM may participate in lung repair through HGF production.

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Human lung fibroblasts produce proresolving peroxisome proliferator-activated receptor-γ ligands in a cyclooxygenase-2-dependent manner

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00272.2016 ◽

2016 ◽

Vol 311 (5) ◽

pp. L855-L867 ◽

Cited By ~ 14

Author(s):

Shannon H. Lacy ◽

Collynn F. Woeller ◽

Thomas H. Thatcher ◽

Krishna Rao Maddipati ◽

Kenneth V. Honn ◽

...

Keyword(s):

Human Lung ◽

Cyclooxygenase 2 ◽

Lung Fibroblasts ◽

Luciferase Reporter ◽

Chronic Obstructive ◽

Peroxisome Proliferator ◽

Dependent Manner ◽

Peroxisome Proliferator Activated Receptor ◽

Human Lung Fibroblasts ◽

Cox 2

Human lung fibroblasts (HLFs) act as innate immune sentinel cells that amplify the inflammatory response to injurious stimuli. Here, we use targeted lipidomics to explore the hypothesis that HLFs also play an active role in the resolution of inflammation. We detected cyclooxygenase-2 (COX-2)-dependent production of both proinflammatory and proresolving prostaglandins (PGs) in conditioned culture medium from HLFs treated with a proinflammatory stimulus, IL-1β. Among the proresolving PGs in the HLF lipidome were several known ligands for peroxisome proliferator-activated receptor-γ (PPARγ), a transcription factor whose activation in the lung yields potent anti-inflammatory, antifibrotic, and proresolving effects. Next, we used a cell-based luciferase reporter to confirm the ability of HLF supernatants to activate PPARγ, demonstrating, for the first time, that primary HLFs activated with proinflammatory IL-1β or cigarette smoke extract produce functional PPARγ ligands; this phenomenon is temporally regulated, COX-2- and lipocalin-type PGD synthase-dependent, and enhanced by arachidonic acid supplementation. Finally, we used luciferase reporter assays to show that several of the PGs in the lipidome of activated HLFs independently activate PPARγ and/or inhibit NFκB. These results indicate that HLFs, as immune sentinels, regulate both proinflammatory and proresolving responses to injurious stimuli. This novel endogenous resolution pathway represents a new therapeutic target for globally important inflammatory diseases such as chronic obstructive pulmonary disease.

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