scholarly journals Nrf2 Alleviates TNF-α-Induced Oxidative Stress Damage in Type II Alveolar Epithelial Cells by Down-Regulating the Expression of NOX1

2020 ◽  
Author(s):  
Weijing Wu ◽  
Jiamin Zhang ◽  
Xihua Lian ◽  
Xiaoping Lin ◽  
Xiaoshan Su ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Binding Site ◽  
Luciferase Activity ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Type Ii ◽  
Alveolar Epithelial ◽  
Tnf Α ◽  
Stress Damage

Abstract Objective: To study the roles of Nrf2 in acute lung injury (ALI) pathogenesis by investigating the effects of Nrf2 on regulating oxidative stress damage in TNF-α-induced type II alveolar epithelial cells (T2AECs).Methods: T2AECs were transfected with Nrf2 siRNA and overexpression vectors for six hours before being induced by TNF-α for 24 hours. Subsequently, levels of interleukins (IL-6 and IL-8), reactive oxygen species (ROS), malondialdehyde (MDA), total antioxidation capability (T-AOC), Nrf2, NOX1 and NF-kB were measured. Additionally, potential Nrf2 binding site in NOX1 promoter was predicted by AliBaba2.1 and two recombinant vectors, namely “pGL3-NOX1-1500” and “pGL3-NOX1-1489, were constructed by inserting the sequence of NOX1 promoter in full-length and that in the absence of Nrf2 binding site to pGL3 basic vector. T2AECs were transfected with these vectors prior to TNF-α induction and the luciferase activity was measured.Results: Levels of IL-6, IL-8, ROS and MDA were increased (P＜0.05) while T-AOC was decreased in TNF-α-induced A549 cells after the transfection of Nrf2 siRNA vector (P＜0.05). In contrast, concentrations of IL-6, IL-8, ROS and MDA were decreased (P＜0.05) whereas T-AOC was increased after the transfection of Nrf2 overexpression vector (P＜0.05). NOX1 promoter possesses one Nrf2 binding site. Cells transfected by “pGL3-NOX1-1500” vector had the highest luciferase activity, followed by cells transfected by “pGL3-NOX1-1489” vector and the control cells (P＜0.05).Conclusion: Nrf2 modulates NOX1 expression via binding to its promoter, by which against TNF-α-induced oxidative stress damage in T2AECs. Thus, Nrf2 might be a therapeutic target for ALI.

scholarly journals Nuclear Factor-kappaB Regulates the Transcription of NADPH Oxidase 1 in Human Alveolar Epithelial Cells

2020 ◽  
Author(s):  
Weijing Wu ◽  
Li Li ◽  
Xiaoshan Su ◽  
ZHIXING ZHU ◽  
Xiaoping Lin ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Ros Generation ◽  
Alveolar Epithelial ◽  
Tnf Α ◽  
Potential Regulatory Role ◽  
Human Alveolar Epithelial Cells

Abstract Objective Acute lung injury (ALI) is characterized by inflammation and oxidative stress. Nuclear factor-kappaB (NF-κB) mediates the expression of various inflammation-related genes, including the NADPH oxidase family. This study aimed to identify the potential regulatory role of NF-𝜅B on NADPH oxidases in TNF-α-induced oxidative stress in human alveolar epithelial cells. Methods Type II alveolar epithelial cell-derived A549 cells were treated with TNF-α for 24 hours to establish ALI cell models. RT-PCR, western blot, DCFH-DA ROS assay, Alibaba 2.1 online analysis, electrophoretic mobility shift assays and luciferase reporter analysis were employed to identify the potential regulatory role of NF-𝜅B on NADPH oxidases in TNF-α-induced oxidative stress in human alveolar epithelial cells. Results The expression of NF-κB/p65 was notably upregulated in TNF-α-stimulated A549 cells. NF-κB knockdown by siRNA significantly inhibited the TNF-α-induced ROS generation. Moreover, NF-𝜅B/p65 siRNA could inhibite the activation of NOX1, NOX2 and NOX4 mRNA and protein expression in TNF-α-stimulated A549 cells. The next study demonstrated that NF-𝜅B activated the transcription of NOX1 by binding to the -261 to -252 bp (NOX1/κB2, TAAAAATCCC) region of NOX1 promoter in TNF-α-stimulated A549 cells. Conclusion Our data demonstrated that NF-κB can aggravate TNF-α-induced ALI by regulating the activation of ROS generation and the expression of NOX1, NOX2 and NOX4. Moreover, NF-𝜅B could promote the NOX1 transcriptional activity via binding its promoter in TNF-α-stimulated A549 cells.

scholarly journals Nuclear factor-kappaB regulates the transcription of NADPH oxidase 1 in human alveolar epithelial cells

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Weijing Wu ◽  
Li Li ◽  
Xiaoshan Su ◽  
Zhixing Zhu ◽  
Xiaoping Lin ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Nuclear Factor ◽  
Alveolar Epithelial ◽  
Tnf Α ◽  
Potential Regulatory Role ◽  
Human Alveolar Epithelial Cells

Abstract Objective Acute lung injury (ALI) is characterized by inflammation and oxidative stress. Nuclear factor-kappaB (NF-κB) mediates the expression of various inflammation-related genes, including the NADPH oxidase family. This study aimed to identify the potential regulatory role of NF-κB on NADPH oxidases in tumor necrosis factor-α (TNF-α)-induced oxidative stress in human alveolar epithelial cells. Methods A549 cells were treated with TNF-α for 24 h to establish ALI cell models. RT-PCR, western blot, assessment of oxidative stress, Alibaba 2.1 online analysis, electrophoretic mobility shift assays and luciferase reporter analysis were employed to identify the potential regulatory role of NF-κB on NADPH oxidases in TNF-α-induced oxidative stress in human alveolar epithelial cells. Results The expression of NF-κB/p65 was notably upregulated in TNF-α-stimulated A549 cells. NF-κB knockdown by siRNA significantly inhibited the TNF-α-induced oxidative stress. Moreover, NF-κB/p65 siRNA could inhibit the activation of NOX1, NOX2 and NOX4 mRNA and protein expression in TNF-α-stimulated A549 cells. The next study demonstrated that NF-κB activated the transcription of NOX1 by binding to the -261 to -252 bp (NOX1/κB2, TAAAAATCCC) region of NOX1 promoter in TNF-α-stimulated A549 cells. Conclusion Our data demonstrated that NF-κB can aggravate TNF-α-induced ALI by regulating the oxidative stress response and the expression of NOX1, NOX2 and NOX4. Moreover, NF-κB could promote the NOX1 transcriptional activity via binding its promoter in TNF-α-stimulated A549 cells.

Expression of the tyrosine phosphatase LAR-PTP2 is developmentally regulated in lung epithelia

1994 ◽  
Vol 267 (3) ◽  
pp. L263-L270 ◽  
Author(s):  
D. Rotin ◽  
B. J. Goldstein ◽  
C. A. Fladd
Keyword(s):  
Epithelial Cells ◽  
Lung Development ◽  
Tyrosine Phosphatase ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii ◽  
Adult Type ◽  
Alveolar Epithelial

The role of tyrosine kinases in regulating cell proliferation, differentiation, and development has been well documented. In contrast, little is known about the role of protein tyrosine phosphatases (PTPs) in mammalian development. To identify PTPs that may be involved in lung development, we have isolated (by polymerase chain reaction) from rat fetal alveolar epithelial cells a cDNA fragment which was identified as the recently cloned tyrosine phosphatase LAR-PTP2. Analysis of tissue expression of LAR-PTP2 identified a approximately 7.5-kb message in the lung, which is also expressed weakly in brain, and an alternatively spliced approximately 6.0-kb message (LAR-PTP2B) expressed in brain. In the fetal lung, LAR-PTP2 was preferentially expressed in lung epithelial (but not fibroblast) cells grown briefly in primary culture, and its expression was tightly regulated during lung development, peaking at 20 days of gestational age (term = 22 days), when mature alveolar type II epithelium first appears. Accordingly, immunoblot analysis revealed high expression of endogenous LAR-PTP2 protein in alveolar epithelial cells from 21-day gestation fetuses. LAR-PTP2 was also expressed in lungs of newborn rats, but transcripts (and protein) were barely detectable in adult lungs and in the nonproliferating adult alveolar type II cells. Interestingly, expression was restored in the transformed adult type II-like A549 cells. These results suggest that LAR-PTP2 may play a role in the proliferation and/or differentiation of epithelial cells during lung development.

Parathyroid hormone-related protein, an autocrine regulatory factor in alveolar epithelial cells

1996 ◽  
Vol 270 (3) ◽  
pp. L353-L361 ◽  
Author(s):  
R. H. Hastings ◽  
D. Summers-Torres ◽  
T. C. Cheung ◽  
L. S. Ditmer ◽  
E. M. Petrin ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Primary Cultures ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Type Ii Cells ◽  
Type Ii ◽  
Alveolar Epithelial ◽  
Parathyroid Hormone Related Protein ◽  
Pthrp Receptor

Alveolar epithelial cells in vivo, primary cultures of adult rat type II cells, and human A549 alveolar carcinoma cells express parathyroid hormone-related protein (PTHrP). Here we demonstrated that type II cells and A549 cells also express the PTHrP receptor and that they exhibit differentiation-related responses to the amino-terminal PTHrP fragment, PTHrP-(1-34). PTHrP receptor expression in A549 cells was shown by detection of a 0.3-kb reverse transcriptase polymerase chain reaction product formed by primers specific for PTHrP receptor. In situ hybridization studies localized the site of production of PTHrP and PTHrP receptor mRNA in rat lung cells with morphology and location typical of type II cells. Primary cultures of such type II cells also expressed PTHrP receptor mRNA. Incubation with PTHrP-(1-34) stimulated disaturated phosphatidylcholine (DSPC) synthesis in A549 cells and increased the release of newly synthesized DSPC by cultured type II cells and A549 cells. In addition, PTHrP-(1-34) increased the number of lamellar bodies per type II cell and increased their expression of alkaline phosphatase in a dose-dependent manner. Thus PTHrP-(1-34) promoted a differentiated type II cell phenotype. Since cultured type II cells, alveolar epithelial cells in vivo, and A549 cells express PTHrP and the PTHrP receptor, PTHrP-(1-34) may be an autocrine regulatory factor in type II cells and lung cancer cells.

scholarly journals YY1 mediates TGF-β1-induced EMT and pro-fibrogenesis in alveolar epithelial cells

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Chuyi Zhang ◽  
Xiaoping Zhu ◽  
Yifei Hua ◽  
Qian Zhao ◽  
Kaijing Wang ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Pulmonary Fibrosis ◽  
Epithelial Mesenchymal Transition ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Lung Epithelial Cells ◽  
Lung Damage ◽  
Type Ii ◽  
Mesenchymal Transition ◽  
Alveolar Epithelial

Abstract Pulmonary fibrosis is a chronic, progressive lung disease associated with lung damage and scarring. The pathological mechanism causing pulmonary fibrosis remains unknown. Emerging evidence suggests prominent roles of epithelial–mesenchymal transition (EMT) of alveolar epithelial cells (AECs) in myofibroblast formation and progressive pulmonary fibrosis. Our previous work has demonstrated the regulation of YY1 in idiopathic pulmonary fibrosis and pathogenesis of fibroid lung. However, the specific function of YY1 in AECs during the pathogenesis of pulmonary fibrosis is yet to be determined. Herein, we found the higher level of YY1 in primary fibroblasts than that in primary epithelial cells from the lung of mouse. A549 and BEAS-2B cells, serving as models for type II alveolar pulmonary epithelium in vitro, were used to determine the function of YY1 during EMT of AECs. TGF-β-induced activation of the pro-fibrotic program was applied to determine the role YY1 may play in pro-fibrogenesis of type II alveolar epithelial cells. Upregulation of YY1 was associated with EMT and pro-fibrotic phenotype induced by TGF-β treatment. Targeted knockdown of YY1 abrogated the EMT induction by TGF-β treatment. Enforced expression of YY1 can partly mimic the TGF-β-induced pro-fibrotic change in either A549 cell line or primary alveolar epithelial cells, indicating the induction of YY1 expression may mediate the TGF-β-induced EMT and pro-fibrosis. In addition, the translocation of NF-κB p65 from the cytoplasm to the nucleus was demonstrated in A549 cells after TGF-β treatment and/or YY1 overexpression, suggesting that NF-κB-YY1 signaling pathway regulates pulmonary fibrotic progression in lung epithelial cells. These findings will shed light on the better understanding of mechanisms regulating pro-fibrogenesis in AECs and pathogenesis of lung fibrosis.

scholarly journals Effect of ulinastatin on paraquat-induced-oxidative stress in human type II alveolar epithelial cells

2013 ◽  
Vol 4 (2) ◽  
pp. 133 ◽  
Author(s):  
Xiao-xiao Meng ◽  
Rui-lan Wang ◽  
Shan Gao ◽  
Hui Xie ◽  
Jiu-ting Tan ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Alveolar Epithelial Cells ◽  
Type Ii ◽  
Alveolar Epithelial ◽  
Human Type

GM-CSF provides autocrine protection for murine alveolar epithelial cells from oxidant-induced mitochondrial injury

2012 ◽  
Vol 302 (3) ◽  
pp. L343-L351 ◽  
Author(s):  
Anne Sturrock ◽  
Elfateh Seedahmed ◽  
Mustafa Mir-Kasimov ◽  
Jonathan Boltax ◽  
Michael L. McManus ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Lung Injury ◽  
Alveolar Epithelial Cells ◽  
Type Ii ◽  
Mitochondrial Injury ◽  
Akt Activation ◽  
Alveolar Epithelial ◽  
Gm Csf

Exposure of mice to hyperoxia induces alveolar epithelial cell (AEC) injury, acute lung injury and death. Overexpression of granulocyte-macrophage colony-stimulating factor (GM-CSF) in the lung protects against these effects, although the mechanisms are not yet clear. Hyperoxia induces cellular injury via effects on mitochondrial integrity, associated with induction of proapoptotic members of the Bcl-2 family. We hypothesized that GM-CSF protects AEC through effects on mitochondrial integrity. MLE-12 cells (a murine type II cell line) and primary murine type II AEC were subjected to oxidative stress by exposure to 80% oxygen and by exposure to H2O2. Exposure to H2O2 induced cytochrome c release and decreased mitochondrial reductase activity in MLE-12 cells. Incubation with GM-CSF significantly attenuated these effects. Protection induced by GM-CSF was associated with Akt activation. GM-CSF treatment also resulted in increased expression of the antiapoptotic Bcl-2 family member, Mcl-1. Primary murine AEC were significantly more tolerant of oxidative stress than MLE-12 cells. In contrast to MLE-12 cells, primary AEC expressed significant GM-CSF at baseline and demonstrated constitutive activation of Akt and increased baseline expression of Mcl-1. Treatment with exogenous GM-CSF further increased Akt activation and Mcl-1 expression in primary AEC. Conversely, suppression of AEC GM-CSF expression by use of GM-CSF-specific small interfering RNA resulted in decreased tolerance of oxidative stress, Furthermore, silencing of Mcl-1 prevented GM-CSF-induced protection. We conclude that GM-CSF protects alveolar epithelial cells against oxidative stress-induced mitochondrial injury via the Akt pathway and its downstream components, including Mcl-1. Epithelial cell-derived GM-CSF may contribute to intrinsic defense mechanisms limiting lung injury.

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