bims-noxint 2021-07-11 papers

Issue of 2021–07–11
four papers selected by
Laia Caja Puigsubira, Uppsala University

Redox Biol. 2021 Jun 18. pii: S2213-2317(21)00209-3. [Epub ahead of print]45 102050

The hydrogen-peroxide producing NADPH oxidase 4 does not limit neointima development after vascular injury in mice.

Giulia K Buchmann, Christoph Schürmann, Manuela Spaeth, Wesley Abplanalp, Lukas Tombor, David John, Timothy Warwick, Flávia Rezende, Andreas Weigert, Ajay M Shah, Martin-Leo Hansmann, Norbert Weissmann, Stefanie Dimmeler, Katrin Schröder, Ralf P Brandes.

OBJECTIVE: The NADPH oxidase Nox4 is an important source of H2O2. Nox4-derived H2O2 limits vascular inflammation and promotes smooth muscle differentiation. On this basis, the role of Nox4 for restenosis development was determined in the mouse carotid artery injury model.
METHODS AND RESULTS: Genetic deletion of Nox4 by a tamoxifen-activated Cre-Lox-system did not impact on neointima formation in the carotid artery wire injury model. To understand this unexpected finding, time-resolved single-cell RNA-sequencing (scRNAseq) from injured carotid arteries of control mice and massive-analysis-of-cDNA-ends (MACE)-RNAseq from the neointima harvested by laser capture microdissection of control and Nox4 knockout mice was performed. This revealed that resting smooth muscle cells (SMCs) and fibroblasts exhibit high Nox4 expression, but that the proliferating de-differentiated SMCs, which give rise to the neointima, have low Nox4 expression. In line with this, the first weeks after injury, gene expression was unchanged between the carotid artery neointimas of control and Nox4 knockout mice.
CONCLUSION: Upon vascular injury, Nox4 expression is transiently lost in the cells which comprise the neointima. NADPH oxidase 4 therefore does not interfere with restenosis development after wire-induced vascular injury.

Keywords: Carotid injury; Inflammation; NADPH oxidase; Nox4; Reactive oxygen species; Restenosis; Single-cell RNA sequencing

DOI: https://doi.org/10.1016/j.redox.2021.102050

Sci Rep. 2021 Jul 08. 11(1): 14097

Regulation of Rac1 transcription by histone and DNA methylation in diabetic retinopathy.

Renu A Kowluru, Rakesh Radhakrishnan, Ghulam Mohammad.

Cytosolic ROS, generated by NADPH oxidase 2 (Nox2) in diabetes, damage retinal mitochondria, which leads to the development of retinopathy. A small molecular weight G-protein essential for Nox2 activation, Rac1, is also transcriptionally activated via active DNA methylation-hydroxymethylation. DNA methylation is a dynamic process, and can also be regulated by histone modifications; diabetes alters retinal histone methylation machinery. Our aim is to investigate the role of histone methylation (H3K9me3) of Rac1 promoter in dynamic DNA methylation- transcriptional activation. Using human retinal endothelial cells in 20 mM D-glucose, H3K9me3 at Rac1 promoter was quantified by chromatin-Immunoprecipitation technique. Crosstalk between H3K9me3 and DNA methylation was examined in cells transfected with siRNA of histone trimethyl-transferase, Suv39H1, or Dnmt1, exposed to high glucose. Key parameters were confirmed in retinal microvessels from streptozotocin-induced diabetic mice, with intravitreally administered Suv39H1-siRNA or Dnmt1-siRNA. Compared to cells in normal glucose, high glucose increased H3K9me3 and Suv39H1 binding at Rac1 promoter, and Suv39H1-siRNA prevented glucose-induced increase 5 hydroxy methyl cytosine (5hmC) and Rac1 mRNA. Similarly, in diabetic mice, Suv39H1-siRNA attenuated increase in 5hmC and Rac1 mRNA. Thus, H3K9me3 at Rac1 promoter assists in active DNA methylation-hydroxymethylation, activating Rac1 transcription. Regulation of Suv39H1-H3K9 trimethylation could prevent further epigenetic modifications, and prevent diabetic retinopathy.

DOI: https://doi.org/10.1038/s41598-021-93420-4

Neuropsychiatr Dis Treat. 2021 ;17 2089-2103

MicroRNA-126a-5p Exerts Neuroprotective Effects on Ischemic Stroke via Targeting NADPH Oxidase 2.

Yu Tan, Feng Zhou, Dejiang Yang, Xiaowei Zhang, Meihong Zeng, Lei Wan.

Background: Ischemic stroke is a destructive cerebrovascular disorder related to oxidative stress; NOX2 is a major source for ROS production; and miR-126a-5p is involved in several diseases, such as abdominal aortic aneurysm. We investigated the role of miR-126a-5p in regulating NOX2 in ischemic stroke.
Methods: MiR-126a-5p and NOX2 were examined in the brains of rats subjected to cerebral ischemia/reperfusion (I/R) by RT-PCR and Western blot. MiR-126a-5p agomir was delivered to examine the effects of miR-126a-5p on I/R injury. The neurological deficit, infarct volume, and brain water content were evaluated. NOX activity, ROS production, and MDA and SOD levels were detected to assess oxidative stress. H&E staining was used to examine cell state. Apoptosis was evaluated by TUNEL, caspase-3 activity, and cleaved-caspase-3 protein level. The relationship between miR-126a-5p and NOX2 was analyzed by bioinformatics and luciferase reporter assay. MiR-126a-5p mimic, miR-126a-5p inhibitor, or pcDNA-NOX2 were transfected in SH-SY5Y cells to further assess the effects of miR-126a-5p on OGD/R-induced cells injury.
Results: NOX2 was upregulated and miR-126a-5p was down-regulated in the brains of I/R rats. MiR-126a-5p agomir obviously reduced the neurological deficit, infarct volume, brain water content, oxidative stress, and apoptosis in I/R rats. MiR-126a-5p targeted NOX2 directly and regulated NOX2 negatively. Moreover, miR-126a-5p mimic elevated cell viability and inhibited oxidative stress and apoptosis in OGD/R-treated SH-SY5Y cells, while miR-126a-5p inhibitor had the opposite effects. NOX2 overexpression antagonized the protective effects of miR-126a-5p mimic on OGD/R-induced cell injury.
Conclusion: MiR-126a-5p is a novel potential target for ischemic stroke therapy due to its protection against cerebral I/R injury via directly targeting NOX2.

Keywords: NOX2; cerebral ischemia/reperfusion; miR-126a-5p; oxidative stress

DOI: https://doi.org/10.2147/NDT.S293611

Redox Biol. 2021 Jun 18. pii: S2213-2317(21)00210-X. [Epub ahead of print]45 102051

Nox2 impairs VEGF-A-induced angiogenesis in placenta via mitochondrial ROS-STAT3 pathway.

Chengjun Hu, Zifang Wu, Zihao Huang, Xiangyu Hao, Shuqi Wang, Jinping Deng, Yulong Yin, Chengquan Tan.

Aberrant placental angiogenesis is associated with fetal intrauterine growth restriction (IUGR), but the mechanism underlying abnormal placental angiogenesis remains largely unknown. Here, lower vessel density and higher expression of NADPH oxidases 2 (Nox2) were observed in the placentae for low birth weight (LBW) fetuses versus normal birth weight (NBW) fetuses, with a negative correlation between Nox2 and placental vessel density. Moreover, it was revealed for the first time that Nox2 deficiency facilitates angiogenesis in vitro and in vivo, and vascular endothelial growth factor-A (VEGF-A) has an essential role in Nox2-controlled inhibition of angiogenesis in porcine vascular endothelial cells (PVECs). Mechanistically, Nox2 inhibited phospho-signal transducer and activator of transcription 3 (p-STAT3) in the nucleus by inducing the production of mitochondrial reactive oxygen species (ROS). Dual-luciferase assay confirmed that knockdown of Nox2 reduces the expression of VEGF-A in an STAT3 dependent manner. Our results indicate that Nox2 is a potential target for therapy by increasing VEGF-A expression to promote angiogenesis and serves as a prognostic indicator for fetus with IUGR.

Keywords: Angiogenesis; IUGR; Nox2; Placenta; STAT3; VEGF-A

DOI: https://doi.org/10.1016/j.redox.2021.102051