scholarly journals Compressive mechanical stress enhances susceptibility to interleukin-1 by increasing interleukin-1 receptor expression in 3D-cultured ATDC5 cells

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Yuki Takeda ◽  
Yasuo Niki ◽  
Yusuke Fukuhara ◽  
Yoshitsugu Fukuda ◽  
Kazuhiko Udagawa ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Mechanical Stress ◽  
Transient Receptor Potential ◽  
Interleukin 1 ◽  
Compressive Loading ◽  
Reactive Oxygen ◽  
Receptor Expression ◽  
Transient Receptor Potential Vanilloid ◽  
Mrna Levels ◽  
Oxygen Species

Abstract Background Mechanical overload applied on the articular cartilage may play an important role in the pathogenesis of osteoarthritis. However, the mechanism of chondrocyte mechanotransduction is not fully understood. The purpose of this study was to assess the effects of compressive mechanical stress on interleukin-1 receptor (IL-1R) and matrix-degrading enzyme expression by three-dimensional (3D) cultured ATDC5 cells. In addition, the implications of transient receptor potential vanilloid 4 (TRPV4) channel regulation in promoting effects of compressive mechanical loading were elucidated. Methods ATDC5 cells were cultured in alginate beads with the growth medium containing insulin-transferrin-selenium and BMP-2 for 6 days. The cultured cell pellet was seeded in collagen scaffolds to produce 3D-cultured constructs. Cyclic compressive loading was applied on the 3D-cultured constructs at 0.5 Hz for 3 h. The mRNA expressions of a disintegrin and metalloproteinases with thrombospondin motifs 4 (ADAMTS4) and IL-1R were determined with or without compressive loading, and effects of TRPV4 agonist/antagonist on mRNA expressions were examined. Immunoreactivities of reactive oxygen species (ROS), TRPV4 and IL-1R were assessed in 3D-cultured ATDC5 cells. Results In 3D-cultured ATDC5 cells, ROS was induced by cyclic compressive loading stress. The mRNA expression levels of ADAMTS4 and IL-1R were increased by cyclic compressive loading, which was mostly prevented by pyrollidine dithiocarbamate. Small amounts of IL-1β upregulated ADAMTS4 and IL-1R mRNA expressions only when combined with compressive loading. TRPV4 agonist suppressed ADAMTS4 and IL-1R mRNA levels induced by the compressive loading, whereas TRPV4 antagonist enhanced these levels. Immunoreactivities to TRPV4 and IL-1R significantly increased in constructs with cyclic compressive loading. Conclusion Cyclic compressive loading induced mRNA expressions of ADAMTS4 and IL-1R through reactive oxygen species. TRPV4 regulated these mRNA expressions, but excessive compressive loading may impair TRPV4 regulation. These findings suggested that TRPV4 regulates the expression level of IL-1R and subsequent IL-1 signaling induced by cyclic compressive loading and participates in cartilage homeostasis.

scholarly journals Macrophage uptake of oxidized and acetylated low-density lipoproteins and generation of reactive oxygen species are regulated by linear stiffness of the growth surface

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0260756
Author(s):  
Erika J. Gruber ◽  
Ali Y. Aygun ◽  
Cynthia A. Leifer
Keyword(s):  
Reactive Oxygen Species ◽  
Protein Expression ◽  
Transient Receptor Potential ◽  
Reactive Oxygen ◽  
Low Density Lipoproteins ◽  
Growth Surface ◽  
Transient Receptor Potential Vanilloid ◽  
Low Density ◽  
Oxygen Species ◽  
Lipid Uptake

Macrophages are key players in the development of atherosclerosis: they scavenge lipid, transform into foam cells, and produce proinflammatory mediators. At the same time, the arterial wall undergoes profound changes in its mechanical properties. We recently showed that macrophage morphology and proinflammatory potential are regulated by the linear stiffness of the growth surface. Here we asked whether linear stiffness also regulates lipid uptake by macrophages. We cultured murine bone marrow-derived macrophages (BMMs) on polyacrylamide gels modeling stiffness of healthy (1kPa) and diseased (10-150kPa) blood vessels. In unprimed BMMs, increased linear stiffness increased uptake of oxidized (oxLDL) and acetylated (acLDL) low density lipoproteins and generation of reactive oxygen species, but did not alter phagocytosis of bacteria or silica particles. Macrophages adapted to stiff growth surfaces had increased mRNA and protein expression of two key lipoprotein receptors: CD36 and scavenger receptor b1. Regulation of the lipoprotein receptor, lectin-like receptor for ox-LDL, was more complex: mRNA expression decreased but surface protein expression increased with increased stiffness. Focal adhesion kinase was required for maximal uptake of oxLDL, but not of acLDL. Uptake of oxLDL and acLDL was independent of rho-associated coiled coil kinase. Through pharmacologic inhibition and genetic deletion, we found that transient receptor potential vanilloid 4 (TRPV4), a mechanosensitive ion channel, plays an inhibitory role in the uptake of acLDL, but not oxLDL. Together, these results implicate mechanical signaling in the uptake of acLDL and oxLDL, opening up the possibility of new pharmacologic targets to modulate lipid uptake by macrophages in vivo.

scholarly journals Reactive Oxygen Species Downregulate Transient Receptor Potential Melastatin 6 Expression Mediated by the Elevation of miR-24-3p in Renal Tubular Epithelial Cells

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1893
Author(s):  
Chieko Hirota ◽  
Yui Takashina ◽  
Yuta Yoshino ◽  
Hajime Hasegawa ◽  
Ema Okamoto ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Transient Receptor Potential ◽  
Reactive Oxygen ◽  
Receptor Potential ◽  
Fluorescence Measurement ◽  
Oxygen Species ◽  
Green Fluorescence ◽  
Transient Receptor Potential Melastatin ◽  
Transient Receptor ◽  
Renal Tubular

Background: A low level of serum magnesium ion (Mg2+) is associated with type 2 diabetes mellitus (T2D). However, the molecular mechanism of Mg2+ deficiency has not been fully clarified. The current study sought to assesses the effect of reactive oxygen species on the expression of Mg2+ channels and miRNA. Methods: The expression of Mg2+ channels and miRNA were examined by real-time polymerase chain reaction. Intracellular Mg2+ concentration was measured by Magnesium Green fluorescence measurement. Results: The mRNA level of transient receptor potential melastatin 6 (TRPM6), which functions as Mg2+ influx channel in the distal convoluted tubule (DCT) of the kidney, was decreased by glycated albumin (GA), but not by insulin in rat renal tubule-derived NRK-52E cells. The mRNA levels of TRPM7, a homologue of TRPM6, and CNNM2, a Mg2+ efflux transporter located at the basolateral membrane of DCT, were changed by neither GA nor insulin. The generation of reactive oxygen species (ROS) was increased by GA. Hydrogen peroxide (H2O2) dose-dependently decreased TRPM6 mRNA, but it inversely increased the reporter activity of TRPM6. H2O2 accelerated the degradation of TRPM6 mRNA in actinomycin D assay without affecting TRPM7 and CNNM2 mRNA expressions. Nine miRNAs were considered as candidates for the regulator of stability of TRPM6 mRNA. Among them, miR-24-3p expression was increased by H2O2. The H2O2-induced reduction of TRPM6 mRNA was rescued by miR-24-3p siRNA. Magnesium Green fluorescence measurement showed that Mg2+ influx is suppressed by H2O2, which was rescued by an antioxidant and miR-24-3p siRNA. Conclusions: We suggest that GA decreases TRPM6 expression mediated by the elevation of ROS and miR-24-3p in renal tubular epithelial cells of T2D.

scholarly journals Doxorubicin-Induced Fetal Mesangial Cell Death Occurs Independently of TRPC6 Channel Upregulation but Involves Mitochondrial Generation of Reactive Oxygen Species

2021 ◽  
Vol 22 (14) ◽  
pp. 7589
Author(s):  
Anberitha T. Matthews ◽  
Hitesh Soni ◽  
Katherine E. Robinson-Freeman ◽  
Theresa A. John ◽  
Randal K. Buddington ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Cell Death ◽  
Transient Receptor Potential ◽  
Mesangial Cell ◽  
Reactive Oxygen ◽  
Receptor Potential ◽  
Oxygen Species ◽  
Glomerular Mesangial Cell ◽  
Trpc6 Channel ◽  
Fetal Pig

Doxorubicin (DOX), a category D pregnancy drug, is a chemotherapeutic agent that has been shown in animal studies to induce fetal toxicity, including renal abnormalities. Upregulation of the transient receptor potential cation (TRPC) 6 channel is involved in DOX-induced podocyte apoptosis. We have previously reported that TRPC6-mediated Ca2+ signaling promotes neonatal glomerular mesangial cell (GMC) death. However, it is unknown whether DOX alters mesangial TRPC expression or viability in the fetus. In this study, cell growth was tracked in control and DOX-treated primary GMCs derived from fetal pigs. Live-cell imaging demonstrated that exposure to DOX inhibited the proliferation of fetal pig GMCs and induced cell death. DOX did not alter the TRPC3 expression levels. By contrast, TRPC6 protein expression in the cells was markedly reduced by DOX. DOX treatment also attenuated the TRPC6-mediated intracellular Ca2+ elevation. DOX stimulated mitochondrial reactive oxygen species (mtROS) generation and mitophagy by the GMCs. The DOX-induced mtROS generation and apoptosis were reversed by the mitochondria-targeted antioxidant mitoquinone. These data suggest that DOX-induced fetal pig GMC apoptosis is independent of TRPC6 channel upregulation but requires mtROS production. The mtROS-dependent GMC death may contribute to DOX-induced fetal nephrotoxicity when administered prenatally.

Reactive oxygen species modulate HIF-1 mediated PAI-1 expression: involvement of the GTPase Rac1

2003 ◽  
Vol 89 (05) ◽  
pp. 926-935 ◽  
Author(s):  
Utta Berchner-Pfannschmidt ◽  
Christoph Wotzlaw ◽  
Robbert Cool ◽  
Joachim Fandrey ◽  
Helmut Acker ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Reactive Oxygen ◽  
Plasminogen Activator Inhibitor ◽  
Dominant Negative ◽  
Mrna Levels ◽  
Ros Production ◽  
Oxygen Species ◽  
Plasminogen Activator Inhibitor 1 ◽  
Pai 1

SummaryThe hypoxia-inducible transcription factor HIF-1 mediates upregulation of plasminogen activator inhibitor-1 (PAI-1) expression under hypoxia. Reactive oxygen species (ROS) have also been implicated in PAI-1 gene expression. However, the role of ROS in HIF-1-mediated regulation of PAI-1 is not clear. We therefore investigated the role of the GTPase Rac1 which modulates ROS production in the pathway leading to HIF-1 and PAI-1 induction.Overexpression of constitutively activated (RacG12V) or dominant-negative (RacT17N) Rac1 increased or decreased, respectively, ROS production. In RacG12V-expressing cells, PAI-1 mRNA levels as well as HIF-1α nuclear presence were reduced under normoxia and hypoxia whereas expression of RacT17N resulted in opposite effects. Treatment with the antioxidant pyrrolidinedithiocarbamate or coexpression of the redox factor-1 restored HIF-1 and PAI-1 promoter activity in RacG12V-cells. In contrast, NFκB activation was enhanced in RacG12V-cells, but abolished by RacT17N. Thus, these findings suggest a mechanism explaining modified fibrinolysis and tissue remodeling in an oxidized environment.

scholarly journals Recent advances in oxygen sensing and signal transduction in hypoxic pulmonary vasoconstriction

2017 ◽  
Vol 123 (6) ◽  
pp. 1647-1656 ◽  
Author(s):  
Ievgen Strielkov ◽  
Oleg Pak ◽  
Natasha Sommer ◽  
Norbert Weissmann
Keyword(s):  
Reactive Oxygen Species ◽  
Transient Receptor Potential ◽  
Oxygen Sensing ◽  
Hypoxic Pulmonary Vasoconstriction ◽  
Reactive Oxygen ◽  
Receptor Potential ◽  
Complex Iii ◽  
Oxygen Species ◽  
Pulmonary Vasoconstriction ◽  
Transient Receptor

Hypoxic pulmonary vasoconstriction (HPV) is a physiological reaction, which adapts lung perfusion to regional ventilation and optimizes gas exchange. Impaired HPV may cause systemic hypoxemia, while generalized HPV contributes to the development of pulmonary hypertension. The triggering mechanisms underlying HPV are still not fully elucidated. Several hypotheses are currently under debate, including a possible decrease as well as an increase in reactive oxygen species as a triggering event. Recent findings suggest an increase in the production of reactive oxygen species in pulmonary artery smooth muscle cells by complex III of the mitochondrial electron transport chain and occurrence of oxygen sensing at complex IV. Other essential components are voltage-dependent potassium and possibly L-type, transient receptor potential channel 6, and transient receptor potential vanilloid 4 channels. The release of arachidonic acid metabolites appears also to be involved in HPV regulation. Further investigation of the HPV mechanisms will facilitate the development of novel therapeutic strategies for the treatment of HPV-related disorders.

scholarly journals Decoy Receptor 3 Promotes Preosteoclast Cell Death via Reactive Oxygen Species-Induced Fas Ligand Expression and the IL-1α/IL-1 Receptor Antagonist Pathway

2020 ◽  
Vol 2020 ◽  
pp. 1-11
Author(s):  
Yi-Jen Peng ◽  
Ching-Tsung Peng ◽  
Yi-Hsuan Lin ◽  
Gu-Jiun Lin ◽  
Shing-Hwa Huang ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Bone Resorption ◽  
Fas Ligand ◽  
Interleukin 1 ◽  
Reactive Oxygen ◽  
Precursor Cells ◽  
Osteoclast Precursor ◽  
Oxygen Species ◽  
Decoy Receptor ◽  
Decoy Receptor 3

Purpose. Interleukin-1α (IL-1α) is a potent cytokine that plays a role in inflammatory arthritis and bone loss. Decoy receptor 3 (DCR3) is an immune modulator of monocytes and macrophages. The aim of this study was to investigate the mechanism of DCR3 in IL-1α-induced osteoclastogenesis. Methods. We treated murine macrophages with DCR3 during receptor activator of nuclear factor kappa Β ligand- (RANKL-) plus IL-1α-induced osteoclastogenesis to monitor osteoclast formation by tartrate-resistant acid phosphatase (TRAP) staining. Osteoclast activity was assessed using a pit formation assay. The mechanisms of inhibition were studied by biochemical analyses, including RT-PCR, immunofluorescent staining, flow cytometry, an apoptosis assay, immunoblotting, and ELISA. Results. DCR3 suppresses IL-1α-induced osteoclastogenesis in both primary murine bone marrow-derived macrophages (BMM) and RAW264.7 cells as it inhibits bone resorption. DCR3 induces RANKL-treated osteoclast precursor cells to express IL-1α, secretory IL-1ra (sIL-1ra), intracellular IL-1ra (icIL-1ra), reactive oxygen species (ROS), and Fas ligand and to activate IL-1α-induced interleukin-1 receptor-associated kinase 4 (IRAK4). The suppression of DCR3 during RANKL- or IL-1α-induced osteoclastogenesis may be due to the abundant secretion of IL-1ra, accumulation of ROS, and expression of Fas ligand in apoptotic osteoclast precursor cells. Conclusions. We concluded that there is an inhibitory effect of DCR3 on osteoclastogenesis via ROS accumulation and ROS-induced Fas ligand, IL-1α, and IL-1ra expression. Our results suggested that the upregulation of DCR3 in preosteoclasts might be a therapeutic target in inflammatory IL-1α-induced bone resorption.

scholarly journals SPARC Levels Modulate the Capacity of Mitomycin to Inhibit the Proliferation of Human Tenon’s Capsule Fibroblasts

2020 ◽  
Vol 2020 ◽  
pp. 1-7
Author(s):  
Yuanyuan Guo ◽  
Shouxiang Ni ◽  
Weiyan Zhou ◽  
Jiangping Hou ◽  
Jiaquan Shen
Keyword(s):  
Reactive Oxygen Species ◽  
Cell Density ◽  
Collagen Gel ◽  
Reactive Oxygen ◽  
Control Group ◽  
Mrna Levels ◽  
Oxygen Species ◽  
Antifibrotic Effect ◽  
Cell Densities

Purpose. To evaluate the role of SPARC in the antiproliferation effect of MMC on human Tenon’s fibroblasts (HTF). Method. Sixteen PACG patients aged 59 ± 10 years (31–72 years), including 6 males and 10 females, were recruited. Tenon tissue was harvested during filtering surgery. Cell density was evaluated after MMC application with different concentrations and application times, by which the optimized MMC application modality was determined. MMC, si-SPARC, or SPARC protein was used when needed to evaluate the cell densities under different conditions, by which the role of SPARC in MMC-mediated antifibrotic process was identified. Results. Considering that the cell densities, as well as SPARC expression on mRNA and protein levels, are relatively stable when the MMC concentration is higher than 0.02% and exposure time longer than 90 s, we chose the MMC application pattern with 0.02% and 90 s as an optimized pattern for the downstream work. Compared to control, the si-SPARC and MMC downregulated the SPARC protein by 91% (P<0.01) and 65% (P<0.01) and mRNA by 96% (P<0.01) and 64% (P<0.01), respectively. MMC decreases the cell densities by 53.50% compared to control. si-SPARC + MMC dramatically deceased the cell density no matter compared to the control group (P<0.01) or MMC group (P<0.01); correspondingly, the relative collagen gel area in the MMC + si-SPARC group was higher than that in the MMC group or si-SPARC group (P<0.05). The reactive oxygen species expression in the MMC + si-SPARC group is higher than that in the MMC group (P<0.05). Conclusion. This study demonstrates that in HTF, (1) MMC downregulates the expression of SPARC in protein and mRNA levels; (2) SPARC depletion has synergistic effect on the antifibrotic effect of MMC; and (3) reactive oxygen species are the possible mediator in the antifibrotic effect of MMC and si-SPARC.

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