bims-noxint 2022-01-23 papers

bims-noxint

Biomed News

on NADPH oxidases in tumorigenesis

Issue of 2022–01–23
six papers selected by
Laia Caja Puigsubira, Uppsala University

Anti-Inflammatory Effects of Rhamnetin on Bradykinin-Induced Matrix Metalloproteinase-9 Expression and Cell Migration in Rat Brain Astrocytes.
Endothelial NOX5 Expression Modulates Thermogenesis and Lipolysis in Mice Fed with a High-Fat Diet and 3T3-L1 Adipocytes through an Interleukin-6 Dependent Mechanism.
The TBX1/miR-193a-3p/TGF-β2 Axis Mediates CHD by Promoting Ferroptosis.
Connexin32 ameliorates epithelial-to-mesenchymal-transition in diabetic renal tubular via inhibiting NOX4 (original article).
Overexpression of NOX2 Exacerbates AngII-Mediated Cardiac Dysfunction and Metabolic Remodelling.
Forsythiaside A Regulates Activation of Hepatic Stellate Cells by Inhibiting NOX4-Dependent ROS.

Int J Mol Sci. 2022 Jan 06. pii: 609. [Epub ahead of print]23(2):

Anti-Inflammatory Effects of Rhamnetin on Bradykinin-Induced Matrix Metalloproteinase-9 Expression and Cell Migration in Rat Brain Astrocytes.

Chien-Chung Yang, Li-Der Hsiao, Ya-Fang Shih, Zih-Yao Yu, Chuen-Mao Yang.

  Bradykinin (BK) has been shown to induce matrix metalloproteinase (MMP)-9 expression and participate in neuroinflammation. The BK/MMP-9 axis can be a target for managing neuroinflammation. Our previous reports have indicated that reactive oxygen species (ROS)-mediated nuclear factor-kappaB (NF-κB) activity is involved in BK-induced MMP-9 expression in rat brain astrocytes (RBA-1). Rhamnetin (RNT), a flavonoid compound, possesses antioxidant and anti-inflammatory effects. Thus, we proposed RNT could attenuate BK-induced response in RBA-1. This study aims to approach mechanisms underlying RNT regulating BK-stimulated MMP-9 expression, especially ROS and NF-κB. We used pharmacological inhibitors and siRNAs to dissect molecular mechanisms. Western blotting and gelatin zymography were used to evaluate protein and MMP-9 expression. Real-time PCR was used for gene expression. Wound healing assay was applied for cell migration. 2',7'-dichlorodihydrofluorescein diacetate (H2DCF-DA) and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) were used for ROS generation and NOX activity, respectively. Promoter luciferase assay and chromatin immunoprecipitation (ChIP) assay were applied to detect gene transcription. Our results showed that RNT inhibits BK-induced MMP-9 protein and mRNA expression, promoter activity, and cell migration in RBA-1 cells. Besides, the levels of phospho-PKCδ, NOX activity, ROS, phospho-ERK1/2, phospho-p65, and NF-κB p65 binding to MMP-9 promoter were attenuated by RNT. In summary, RNT attenuates BK-enhanced MMP-9 upregulation through inhibiting PKCδ/NOX/ROS/ERK1/2-dependent NF-κB activity in RBA-1.

Keywords:  BK; MMP-9; NADPH oxidase; ROS; astrocytes; neuroinflammation; rhamnetin

DOI:  https://doi.org/10.3390/ijms23020609
Antioxidants (Basel). 2021 Dec 24. pii: 30. [Epub ahead of print]11(1):

Endothelial NOX5 Expression Modulates Thermogenesis and Lipolysis in Mice Fed with a High-Fat Diet and 3T3-L1 Adipocytes through an Interleukin-6 Dependent Mechanism.

Jorge G García, Carlos de Miguel, Fermín I Milagro, Guillermo Zalba, Eduardo Ansorena.

  Obesity is a global health issue associated with the development of metabolic syndrome, which correlates with insulin resistance, altered lipid homeostasis, and other pathologies. One of the mechanisms involved in the development of these pathologies is the increased production of reactive oxygen species (ROS). One of the main producers of ROS is the family of nicotinamide adenine dinucleotide phosphate (NADPH) oxidases, among which NOX5 is the most recently discovered member. The aim of the present work is to describe the effect of endothelial NOX5 expression on neighboring adipose tissue in obesity conditions by using two systems. An in vivo model based on NOX5 conditional knock-in mice fed with a high-fat diet and an in vitro model developed with 3T3-L1 adipocytes cultured with conditioned media of endothelial NOX5-expressing bEnd.3 cells, previously treated with glucose and palmitic acid. Endothelial NOX5 expression promoted the expression and activation of specific markers of thermogenesis and lipolysis in the mesenteric and epididymal fat of those mice fed with a high-fat diet. Additionally, the activation of these processes was derived from an increase in IL-6 production as a result of NOX5 activity. Accordingly, 3T3-L1 adipocytes treated with conditioned media of endothelial NOX5-expressing cells, presented higher expression of thermogenic and lipolytic genes. Moreover, endothelial NOX5-expressing bEnd.3 cells previously treated with glucose and palmitic acid also showed interleukin (IL-6) production. Finally, it seems that the increase in IL-6 stimulated the activation of markers of thermogenesis and lipolysis through phosphorylation of STAT3 and AMPK, respectively. In conclusion, in response to obesogenic conditions, endothelial NOX5 activity could promote thermogenesis and lipolysis in the adipose tissue by regulating IL-6 production.

Keywords:  IL-6; NADPH oxidase 5; lipolysis; obesity; thermogenesis

DOI:  https://doi.org/10.3390/antiox11010030
Oxid Med Cell Longev. 2022 ;2022 5130546

The TBX1/miR-193a-3p/TGF-β2 Axis Mediates CHD by Promoting Ferroptosis.

Li Zhong, Huiqin Yang, Binlu Zhu, Xueqi Zhao, Meijun Xie, Meiling Cao, Chang Liu, Danyang Zhao, Yuan Lyu, Weiguang Shang, Bo Wang, Ying Wu, Xiuju Sun, Guangrong Qiu, Weineng Fu, Hongkun Jiang.

Congenital heart disease (CHD) is the most common noninfectious cause of death during the neonatal stage. T-box transcription factor 1 (TBX1) is the main genetic determinant of 22q11.2 deletion syndrome (22q11.2DS), which is a common cause of CHD. Moreover, ferroptosis is a newly discovered kind of programmed cell death. In this study, the interaction among TBX1, miR-193a-3p, and TGF-β2 was tested using quantitative reverse transcription polymerase chain reaction (qRT-PCR), Western blotting, and dual-luciferase reporter assays. TBX1 silencing was found to promote TGF-β2 messenger ribonucleic acid (mRNA) and protein expression by downregulating the miR-193a-3p levels in H9c2 cells. In addition, the TBX1/miR-193a-3p/TGF-β2 axis was found to promote ferroptosis based on assessments of lipid reactive oxygen species (ROS) levels, Fe2+ concentrations, mitochondrial ROS levels, and malondialdehyde (MDA) contents; Cell Counting Kit-8 (CCK-8) assays and transmission electron microscopy; and Western blotting analysis of glutathione peroxidase 4 (GPX4), nuclear factor erythroid 2-related factor 2 (NRF2), heme oxygenase-1 (HO-1), NADPH oxidase 4 (NOX4), and acyl-CoA synthase long-chain family member 4 (ACSL4) protein expression. The protein expression of NRF2, GPX4, HO-1, NOX4, and ACSL4 and the level of MDA in human CHD specimens were also detected. In addition, TBX1 and miR-193a-3p expression was significantly downregulated and TGF-β2 levels were high in human embryonic CHD tissues, as indicated by the H9c2 cell experiments. In summary, the TBX1/miR-193a-3p/TGF-β2 axis mediates CHD by inducing ferroptosis in cardiomyocytes. TGF-β2 may be a target gene for CHD diagnosis and treatment in children.

DOI: https://doi.org/10.1155/2022/5130546
Pharmacol Res. 2022 Jan 17. pii: S1043-6618(22)00029-9. [Epub ahead of print] 106084

Connexin32 ameliorates epithelial-to-mesenchymal-transition in diabetic renal tubular via inhibiting NOX4 (original article).

Xiaohong Sun, Haiming Xiao, Shanshan Li, Rui Chen, Zeyuan Lin, Yan Yang, Zhiquan Chen, Li Deng, Heqing Huang.

  Renal tubulointerstitial fibrosis (RIF), characterized by epithelial-to-mesenchymal transition (EMT) of renal tubular epithelial cells (TECs), is the main cause of diabetic renal fibrosis. Oxidative stress plays a pivotal role in the development of diabetic RIF. Connexin32 (Cx32), prominently expressed in renal TECs, has emerged as an important player in the regulation of oxidative stress. However, the role of Cx32 in diabetic RIF has not been explored yet. Here, we showed that adenovirus-mediated Cx32 overexpression suppressed EMT to ameliorate RIF and renal function in STZ-induced diabetic mice, while knockout (KO) of Cx32 exacerbated RIF in diabetic mice. Moreover, overexpression of Cx32 inhibited EMT and the production of extra cellular matrix (ECM) in high glucose (HG) induced NRK-52E cells, whereas knockdown of Cx32 showed the opposite effects. Furthermore, we showed that NOX4, the main source of ROS in renal tubular, was down-regulated by Cx32. Mechanistically, Cx32 down-regulated the expression of PKC alpha in a carboxyl-terminal-dependent manner, thereby inhibiting the phosphorylation at Thr147 of p22phox triggered by PKC alpha, which ultimately repressed the formation of the p22phox-NOX4 complex to reduce the protein level of NOX4. Thus, we establish Cx32 as a novel target and confirm the protection mechanism in RIF.

Keywords:  CYBA/p22phox; Cx32; D-glucose (PubChem CID: 5793); DAPI (PubChem CID: 160166); Dihydroethidium (PubChem CID: 128682); EMT; Lucifer Yellow (PubChem CID: 20835957); NOX4; PKC alpha; Renal tubulointerstitial fibrosis; Streptozocin (PubChem CID: 29327)

DOI:  https://doi.org/10.1016/j.phrs.2022.106084
Antioxidants (Basel). 2022 Jan 10. pii: 143. [Epub ahead of print]11(1):

Overexpression of NOX2 Exacerbates AngII-Mediated Cardiac Dysfunction and Metabolic Remodelling.

Synne S Hansen, Tina M Pedersen, Julie Marin, Neoma T Boardman, Ajay M Shah, Ellen Aasum, Anne D Hafstad.

  The present study aimed to examine the effects of low doses of angiotensin II (AngII) on cardiac function, myocardial substrate utilization, energetics, and mitochondrial function in C57Bl/6J mice and in a transgenic mouse model with cardiomyocyte specific upregulation of NOX2 (csNOX2 TG). Mice were treated with saline (sham), 50 or 400 ng/kg/min of AngII (AngII50 and AngII400) for two weeks. In vivo blood pressure and cardiac function were measured using plethysmography and echocardiography, respectively. Ex vivo cardiac function, mechanical efficiency, and myocardial substrate utilization were assessed in isolated perfused working hearts, and mitochondrial function was measured in left ventricular homogenates. AngII50 caused reduced mechanical efficiency despite having no effect on cardiac hypertrophy, function, or substrate utilization. AngII400 slightly increased systemic blood pressure and induced cardiac hypertrophy with no effect on cardiac function, efficiency, or substrate utilization. In csNOX2 TG mice, AngII400 induced cardiac hypertrophy and in vivo cardiac dysfunction. This was associated with a switch towards increased myocardial glucose oxidation and impaired mitochondrial oxygen consumption rates. Low doses of AngII may transiently impair cardiac efficiency, preceding the development of hypertrophy induced at higher doses. NOX2 overexpression exacerbates the AngII -induced pathology, with cardiac dysfunction and myocardial metabolic remodelling.

Keywords:  NOX2; angiotensin II; cardiac disease; cardiac efficiency; cardiac hypertrophy; hypertension

DOI:  https://doi.org/10.3390/antiox11010143
Oxid Med Cell Longev. 2022 ;2022 9938392

Forsythiaside A Regulates Activation of Hepatic Stellate Cells by Inhibiting NOX4-Dependent ROS.

Mengting Zhou, Xingtao Zhao, Li Liao, Ying Deng, Meichen Liu, Jing Wang, Xinyan Xue, Yunxia Li.

Hepatic stellate cells (HSCs) activation is an important step in the process of hepatic fibrosis. NOX4 and reactive oxygen species expressed in HSCs play an important role in liver fibrosis. Forsythiaside A (FA), a phenylethanoid glycoside extracted and isolated from Forsythiae Fructus, has significant antioxidant activities. However, it is not clear whether FA can play a role in inhibiting the HSCs activation through regulating NOX4/ROS pathway. Therefore, our purpose is to explore the effect and mechanism of FA on HSCs activation to alleviate liver fibrosis. LX2 cells were activated by TGF-β1 in vitro. MTT assay and Wound Healing assay were used to investigate the effect of FA on TGF-β1-induced LX2 cell proliferation and migration. Elisa kit was used to measure the expression of MMP-1 and TIMP-1. Western blot and RT-qPCR were used to investigate the expression of fibrosis-related COLI, α-SMA, MMP-1 and TIMP-1, and inflammation-related TNF-α, IL-6 and IL-1β. The hydroxyproline content was characterized using a biochemical kit. The mechanism of FA to inhibit HSCs activation and apoptosis was detected by DCF-DA probe, RT-qPCR, western blot and flow cytometry. NOX4 siRNA was used to futher verify the effect of FA on NOX4/ROS pathway. The results showed that FA inhibited the proliferation and migration of LX2 cells and adjusted the expression of MMP-1, TIMP-1, COLI, α-SMA, TNF-α, IL-6 and IL-1β as well as promoted collagen metabolism to show potential in anti-hepatic fibrosis. Mechanically, FA down-regulated NOX4/ROS signaling pathway to improve oxidation imbalances, and subsequently inhibited PI3K/Akt pathway to suppress proliferation. FA also promoted the apoptosis of LX2 cells by Bax/Bcl2 pathway. Furthermore, the effects of FA on TGF-β1-induced increased ROS levels and α-SMA and COLI expression were weaken by silencing NOX4. In conclusion, FA had potential in anti-hepatic fibrosis at least in part by remolding of extracellular matrix and improving oxidation imbalances to inhibit the activation of HSCs and promote HSCs apoptosis.

DOI: https://doi.org/10.1155/2022/9938392