bims-noxint 2022-04-10 papers

Issue of 2022‒04‒10
five papers selected by
Laia Caja Puigsubira
Uppsala University

Free Radic Res. 2022 Apr 04. 1-16

Hypoxia Induces Proliferation via NOX4-Mediated Oxidative Stress and TGF-β3 Signaling in Uterine Leiomyoma Cells.

Mariko Miyashita-Ishiwata, Malak El Sabeh, Lauren D Reschke, Sadia Afrin, Mostafa A Borahay.

Uterine leiomyomas, the most common tumors of the female reproductive system, are known to have a hypoxic microenvironment. However, the role of such environment in leiomyoma pathobiology remains unknown. The objective was to determine the effects of hypoxia on leiomyoma cells, and the mechanisms. We found that hypoxia induces proliferation and inhibits apoptosis in human leiomyoma cells. This pro-proliferative effect was accompanied by an increase in reactive oxygen species (ROS) generation and the expression of NADPH oxidase 4 (NOX4). The specific NOX4 inhibitor GLX351322 abrogated this hypoxia-induced ROS generation, cellular proliferation, and apoptosis inhibition. To further investigate the mechanism of NOX4-mediated proliferation, we treated leiomyoma cells grown in normoxia with media from leiomyoma cells cultured under hypoxia. This resulted in increased ROS generation and NOX4 expression, suggesting the hypoxia-induced effects are mediated by an autocrine mechanism. We worked to identify the nature of this autocrine factor. We found that the expression of TGF-β3 and its downstream signaling target pSmad3, are increased in hypoxic leiomyoma cells. To examine the hypothesis that TGF-β3 is, at least, a part of this autocrine mechanism, we treated hypoxic leiomyoma cells with the HIF-1α inhibitor KC7F2 which we discovered to ameliorate the hypoxia-induced TGF-β3 expression. Furthermore, pharmacologic inhibition with the TGF-β/Smad inhibitor SB431542 reduced hypoxia-induced NOX4 expression and ROS generation and attenuated cell proliferation. Thus, we have identified a novel mechanism by which hypoxia induces proliferation in leiomyoma cells. This finding adds to our understanding of leiomyoma pathobiology and can help in identifying new therapeutic targets.Supplemental data for this article can be accessed here.

Keywords: Leiomyoma; NOX4; ROS; TGF-β3; hypoxia; oxidative stress

DOI: https://doi.org/10.1080/10715762.2022.2061967

Cell Death Discov. 2022 Apr 08. 8(1): 177

Targeting NOX4 disrupts the resistance of papillary thyroid carcinoma to chemotherapeutic drugs and lenvatinib.

Ping Tang, Jianfeng Sheng, Xiujuan Peng, Renfei Zhang, Tao Xu, Jun Hu, Yuexi Kang, Baiyi Wu, Hao Dang.

Advanced differentiated thyroid cancer cells are subjected to extreme nutritional starvation which contributes to develop resistance to treatments; however, the underlying mechanism remains unclear. Cells were subjected to serum deprivation by culture in medium containing 0.5% fetal bovine serum. A CCK8 assay, cell death Detection ELISAPLUS kit, and PI staining were conducted to determine cell viability, cell apoptosis, and cell cycle, respectively. NADPH oxidase 4 (NOX4) knockdown-stable cell lines were generated by lentivirus-mediated shRNA knockdown in BCPAP cells and TPC-1 cells. Etoposide and doxorubicin, two chemotherapeutic drugs, as well as lenvatinib were utilized to determine the effect of NOX4 on drug resistance. Lenvatinib-resistant BCPAP cells (LRBCs) were established to confirm this effect. The underlining mechanisms of NOX4 under starvation were explored using western blot. Finally, GLX351322, an inhibitor targeting NOX4, was used to inhibit NOX4-derived ROS in vitro and detect its effect on drug resistance of tumor cells in vivo. NOX4 is overexpressed under serum deprivation in BCPAP or TPC-1 cells. NOX4 knockdown impairs cell viability, increases cell apoptosis, extends G1 phase during cell cycle and modulates the level of energy-associated metabolites in starved cells. When the starved cells or LRBCs are treated with chemotherapeutic drugs or Lenvatinib, NOX4 knockdown inhibits cell viability and aggravates cell apoptosis depending on NOX4-derived ROS production. Mechanistically, starvation activates TGFβ1/SMAD3 signal, which mediates NOX4 upregulation. The upregulated NOX4 then triggers ERKs and PI3K/AKT pathway to influence cell apoptosis. GLX351322, a NOX4-derived ROS inhibitor, has an inhibitory effect on cell growth in vitro and the growth of BCPAP-derived even LRBCs-derived xenografts in vivo. These findings highlight NOX4 and NOX4-derived ROS as a potential therapeutic target in resistance to PTC.

DOI: https://doi.org/10.1038/s41420-022-00994-7

Redox Biol. 2022 Mar 24. pii: S2213-2317(22)00068-4. [Epub ahead of print]52 102296

Dietary salt initiates redox signaling between endothelium and vascular smooth muscle through NADPH oxidase 4.

Kai Er Ying, Wenguang Feng, Wei-Zhong Ying, Xingsheng Li, Dongqi Xing, Yong Sun, Yabing Chen, Paul W Sanders.

Prevention of phenotype switching of vascular smooth muscle cells is an important determinant of normal vascular physiology. Hydrogen peroxide (H2O2) promotes osteogenic differentiation of vascular smooth muscle cells through expression of Runt related transcription factor 2 (Runx2). In this study, an increase in dietary NaCl increased endothelial H2O2 generation through NOX4, a NAD(P)H oxidase. The production of H2O2 was sufficient to increase Runx2, osteopontin and osteocalcin in adjacent vascular smooth muscle cells from control littermate mice but was inhibited in mice lacking endothelial Nox4. A vascular smooth muscle cell culture model confirmed the direct involvement of the activation of protein kinase B (Akt) with inactivation of FoxO1 and FoxO3a observed in the control mice on the high NaCl diet. The present study also showed a reduction of catalase activity in aortas during high NaCl intake. The findings demonstrated an interesting cell-cell communication in the vascular wall that was initiated with H2O2 production by endothelium and was regulated by dietary NaCl intake. A better understanding of how dietary salt intake alters vascular biology may improve treatment of vascular disease that involves activation of Runx2.

Keywords: Cell signaling; Dietary salt; Hydrogen peroxide; Runx2; Vascular smooth muscle

DOI: https://doi.org/10.1016/j.redox.2022.102296

J Inflamm Res. 2022 ;15 2121-2139

Apelin-13 Attenuates Lipopolysaccharide-Induced Inflammatory Responses and Acute Lung Injury by Regulating PFKFB3-Driven Glycolysis Induced by NOX4-Dependent ROS.

Yafei Yuan, Wei Wang, Yue Zhang, Qiaohui Hong, Wenhui Huang, Lijuan Li, Zhanzhan Xie, Yixin Chen, Xu Li, Ying Meng.

Purpose: Acute lung injury (ALI) is a life-threatening condition with limited therapeutic options. Macrophage inflammation plays a key role in the development of ALI. Abnormal glycolysis of macrophages contributes to the inflammatory response. However, the role of macrophage glycolysis in ALI still requires investigation. Apelin-13 has been shown to protect against ALI, whereas the underlying mechanisms remain unclear. In this study, we explored the effect of apelin-13 on lipopolysaccharide (LPS)-induced inflammation and ALI via regulation of glycolysis by modulating redox homeostasis in macrophages.Methods: Serums from 34 patients with sepsis and 13 healthy volunteers were analyzed. In vivo, the protective effect of apelin-13 against LPS-induced ALI was evaluated using a mouse model of LPS-induced ALI. In vitro, mouse bone marrow macrophages (BMDMs) were pretreated with the antioxidant, NADPH oxidase (NOX) 4 (NOX4) small-interfering RNA (siRNA), the 6-phosphofructo-2 -kinase/fructose- 2,6-biphosphatase 3 (PFKFB3) siRNA, or the PFKFB3 overexpression plasmid before exposure to LPS.
Results: Serum apelin-13 levels were significantly elevated in patients with sepsis and sepsis-associated acute respiratory distress syndrome (ARDS) (P<0.0001). In vivo, apelin-13 suppressed LPS-induced ALI and inflammatory cytokine production (P<0.05). Furthermore, apelin-13 reduced hydrogen peroxide (H2O2) content, NOX4 protein levels, and glycolysis. In vitro, LPS stimulation elevated NOX4 protein levels and reactive oxygen species (ROS) production (P<0.05). These changes resulted in the accumulation of glycolysis in BMDMs. Treatment with antioxidant or NOX4 siRNA inhibited LPS-induced glycolysis and inflammatory cytokine production (P<0.05). Moreover, in vitro experiments revealed that PFKFB3 regulates the release of pro-inflammatory cytokines by modulating glycolysis. In contrast, the action of apelin-13 opposed the effects of LPS.
Conclusion: In conclusion, apelin-13 protects against LPS-induced inflammatory responses and ALI by regulating PFKFB3-driven glycolysis induced by NOX4-dependent ROS.

Keywords: NADPH oxidase 4; PFKFB3; acute lung injury; apelin-13; glycolysis; inflammation; mice

DOI: https://doi.org/10.2147/JIR.S348850

Cell Signal. 2022 Apr 04. pii: S0898-6568(22)00091-2. [Epub ahead of print] 110330

NOX2-derived hydrogen peroxide impedes the AMPK/Akt-mTOR signaling pathway contributing to cell death in neuronal cells.

Ruijie Zhang, Chunxiao Liu, Liu Yang, Tong Ji, Nana Zhang, Xiaoqing Dong, Xin Chen, Jing Ma, Wei Gao, Shile Huang, Long Chen.

Oxidative stress is closely related to the pathogenesis of Parkinson's disease (PD), a typical neurodegenerative disease. NADPH oxidase 2 (NOX2) is involved in hydrogen peroxide (H2O2) generation. Recently, we have reported that treatment with H2O2 and PD toxins, including 6-hydroxydopamine (6-OHDA), 1-Methyl-4-phenylpyridin-1-ium (MPP+) and rotenone, induces neuronal apoptosis by inhibiting the mTOR pathway. Here, we show that treatment with 6-OHDA, MPP+ or rotenone induced H2O2 generation by upregulating the levels of NOX2 and its regulatory proteins (p22phox, p40phox, p47phox, p67phox, and Rac1), leading to apoptotic cell death in PC12 cells and primary neurons. Inhibition of NOX2 with apocynin or diphenyleneiodonium, or knockdown of NOX2 powerfully attenuated PD toxins-evoked NOX2 and H2O2, thereby hindering activation of AMPK, inhibition of Akt/mTOR, and induction of apoptosis in neuronal cells. Pretreatment with catalase, a H2O2-scavenging enzyme, blocked the effects of PD toxins on NOX2-dependent H2O2 production, AMPK/Akt/mTOR signaling and apoptosis in the cells. Similar effects were also seen in the cells pretreated with Mito-TEMPO, a mitochondria-selective superoxide scavenger, implying a mitochondrial H2O2-dependent mechanism involved. Further research revealed that ectopic expression of constitutively active Akt or dominant negative AMPKα, or inhibition of AMPK with compound C suppressed PD toxins-induced expression of NOX2 and its regulatory proteins, as well as consequential H2O2 production and apoptosis in the cells. Taken together, these results indicate that certain PD toxins can impede the AMPK/Akt-mTOR signaling pathway leading to neuronal apoptosis by eliciting NOX2-derived H2O2 production. Our findings suggest that neuronal loss in PD may be prevented by regulating the NOX2, AMPK/Akt-mTOR signaling and/or applying antioxidants to ameliorate oxidative stress.

Keywords: AMPK; Akt; Hydrogen peroxide; NADPH oxidase 2; Neuronal cells; mTOR

DOI: https://doi.org/10.1016/j.cellsig.2022.110330