bims-noxint 2022-06-19 papers

Issue of 2022–06–19
six papers selected by
Laia Caja Puigsubira, Uppsala University

Commun Biol. 2022 Jun 14. 5(1): 583

Nox4 expression in osteo-progenitors controls bone development in mice during early life.

Jin-Ran Chen, Oxana P Lazarenko, Michael L Blackburn, Jennifer F Chen, Christopher E Randolph, Jovanny Zabaleta, Katrin Schroder, Kim B Pedersen, Martin J J Ronis.

Tightly regulated and cell-specific NADPH-oxidases (Nox) represent one of the major sources of reactive oxygen species (ROS) signaling molecules that are involved in tissue development and stem cell self-renewal. We have characterized the role of Nox4 in osteo-progenitors during postnatal bone development. Nox4 expression in bone and ROS generation were increased during early osteoblast differentiation and bone development. Stromal osteoblastic cell self-renewal, proliferation and ROS production were significantly lower in samples from whole-body Nox4 knockout mice (Nox4-/-) and conditional knockout (CKO) mice with depletion of Nox4 in the limb bud mesenchyme compared with those from control mice (Nox4fl/fl), but they were reversed after 9 passages. In both sexes, bone volume, trabecular number and bone mineral density were significantly lower in 3-week old CKO and Nox4-/- mice compared with Nox4fl/fl controls. This was reflected in serum levels of bone formation markers alkaline phosphatase (ALP) and procollagen 1 intact N-terminal propeptide (P1NP). However, under-developed bone formation in 3-week old CKO and Nox4-/- mice quickly caught up to levels of control mice by 6-week of age, remained no different at 13-week of age, and was reversed in 32-week old male mice. Osteoclastogenesis showed no differences among groups, however, CTX1 reflecting osteoclast activity was significantly higher in 3-week old male CKO and Nox4-/- mice compared with control mice, and significantly lower in 32-week old Nox4-/- mice compared with control mice. These data suggest that Nox4 expression and ROS signaling in bone and osteoblastic cells coordinately play an important role in osteoblast differentiation, proliferation and maturation.

DOI: https://doi.org/10.1038/s42003-022-03544-0

PLoS One. 2022 ;17(6): e0269130

NADPH-oxidase 4 gene over-expression in peripheral blood lymphocytes of the schizophrenia patients.

Elizaveta S Ershova, Galina V Shmarina, Andrey V Martynov, Natalia V Zakharova, Roman V Veiko, Pavel E Umriukhin, George P Kostyuk, Sergey I Kutsev, Natalia N Veiko, Svetlana V Kostyuk.

INTRODUCTION: Increased systemic oxidative stress is common in schizophrenia (SZ) patients. NADPH-oxidase 4 (NOX4) is the cell oxidoreductase, catalyzing the hydrogen peroxide formation. Presumably, NOX4 is the main oxidative stress factor in a number of diseases such as cardiovascular diseases and cancer. We hypothesized that NOX4 may be involved in the oxidative stress development caused by the disease in the schizophrenic patients' peripheral blood lymphocytes (PBL).
MATERIALS AND METHODS: The SZ group included 100 patients (68 men and 32 women aged 28 ± 11 years). The control group included 60 volunteers (35 men and 25 women aged 25 ± 12 years). Flow cytometry analysis (FCA) was used for DNA damage markers (8-oxodG, ɣH2AX), pro- and antiapoptotic proteins (BAX1 and BCL2) and the master-regulator of anti-oxidant response NRF2 detection in the lymphocytes of the untreated SZ patients (N = 100) and the healthy control (HC, N = 60). FCA and RT-qPCR were used for NOX4 and RNANOX4 detection in the lymphocytes. RT-qPCR was used for mtDNA quantitation in peripheral blood mononuclear cells. Cell-free DNA concentration was determined in blood plasma fluorimetrically.
RESULTS: 8-oxodG, NOX4, and BCL2 levels in the PBL in the SZ group were higher than those in the HC group (p < 0.001). ɣH2AX protein level was increased in the subgroup with high 8-oxodG (p<0.02) levels and decreased in the subgroup with low 8-oxodG (p <0.0001) levels. A positive correlation was found between 8-oxodG, ɣH2AX and BAX1 levels in the SZ group (p <10-6). NOX4 level in lymphocytes did not depend on the DNA damage markers values and BAX1 and BCL2 proteins levels. In 15% of PBL of the HC group a small cellular subfraction was found (5-12% of the total lymphocyte pool) with high DNA damage level and elevated BAX1 protein level. The number of such cells was maximal in PBL samples with low NOX4 protein levels.
CONCLUSION: Significant NOX4 gene expression was found a in SZ patients' lymphocytes, but the corresponding protein is probably not a cause of the DNA damage.

DOI: https://doi.org/10.1371/journal.pone.0269130

Phytomedicine. 2022 May 29. pii: S0944-7113(22)00293-8. [Epub ahead of print]103 154214

Scutellarin acts on the AR-NOX axis to remediate oxidative stress injury in a mouse model of cerebral ischemia/reperfusion injury.

Minzhen Deng, Jingbo Sun, Lilin Peng, Yan Huang, Wen Jiang, Shuang Wu, Lihua Zhou, Sookja Kim Chung, Xiao Cheng.

BACKGROUND: Oxidative stress plays an important role in the pathology of ischemic stroke. Studies have confirmedthat scutellarin has antioxidant effects against ischemic injury, and we also reported that the involvement of Aldose reductase (AR) in oxidative stress and cerebral ischemic injury, in this study we furtherly explicit whether the antioxidant effect of scutellarin on cerebral ischemia injury is related to AR gene regulation and its specific mechanism.
METHODS: C57BL/6N mice (Wild-type, WT) and AR knockout (AR-/-) mice suffered from transient middle cerebral artery occlusion (tMCAO) injury (1 h occlusion followed by 3 days reperfusion), and scutellarin was administered from 2 h before surgery to 3 days after surgery. Subsequently, neurological function was assessed by the modified Longa score method, the histopathological morphology observed with 2,3,5-triphenyltetrazolium chloride (TTC) and hematoxylin-eosin (HE) staining. Enzyme-linked immunosorbent assay (Elisa) was used to detect the levels of ROS, 4-hydroxynonenal (4-HNE), 8-hydroxydeoxyguanosine (8-OHDG), Neurotrophin-3 (NT-3), poly ADP-ribose polymerase-1 (PARP1) and 3-nitrotyrosine (3-NT) in the ischemic penumbra regions. Quantitative proteomics profiling using quantitative nano-HPLC-MS/MS were performed to compare the protein expression difference between AR-/- and WT mice with or without tMCAO injury. The expression of AR, nicotinamide adenine dinucleotide phosphate oxidases (NOX1, NOX2 and NOX4) in the ipsilateral side of ischemic brain were detected by qRT-PCR, Western blot and immunofluorescence co-staining with NeuN.
RESULTS: Scutellarin treatment alleviated brain damage in tMCAO stroke model such as improved neurological function deficit, brain infarct area and neuronal injury and reduced the expression of oxidation-related products, moreover, also down-regulated tMCAO induced AR mRNA and protein expression. In addition, the therapeutic effect of scutellarin on the reduction of cerebral infarction area and neurological function deficits abolished in AR-/- mice under ischemia cerebral injury, which indicated that the effect of scutellarin treatment on tMCAO injury is through regulating AR gene. Proteomic analysis of AR-/- and WT mice indicated AR knockout would affect oxidation reaction even as NADPH related process and activity in mice under cerebral ischemia conditions. Moreover, NOX isoforms (NOX1, NOX2 and NOX4) mRNA and protein expression were significant decreased in neurons of penumbra region in AR-/- mice compared with that in WT mice at 3d after tMCAO injury, which indicated that AR should be the upstream protein regulating NOX after cerebral ischemia.
CONCLUSIONS: We first reported that AR directly regulates NOX subtypes (not only NOX2 but also NOX1 and NOX4) after cerebral ischaemic injury. Scutellarin specifically targets the AR-NOX axis and has antioxidant effects in mice with cerebral ischaemic injury, providing a theoretical basis and accurate molecular targets for the clinical application of scutellarin.

Keywords: Aldose reductase; Ischemia-reperfusion; NADPH oxidase; Quantitative proteomic analysis; Scutellarin

DOI: https://doi.org/10.1016/j.phymed.2022.154214

J Immunol Res. 2022 ;2022 3815853

IL6 Induces mtDNA Leakage to Affect the Immune Escape of Endometrial Carcinoma via cGAS-STING.

Xue Zeng, Xiaosong Li, Yundong Zhang, Chaoxia Cao, Qin Zhou.

Endometrial carcinoma (EC) is a commonly diagnosed gynecological malignancy. Interleukin-6 (IL6) plays a critical role in modulating the progression of several types of tumors, including EC. However, the specific mechanism of IL6 in regulating EC progression has not been clearly elucidated. In this study, we performed a series of functional experiments to explore the potential mechanisms involved in IL6 function in the progression of EC. Here, we found that IL6 increased reactive oxygen species (ROS) generation by enhancing the NADPH oxidase (NOX) level and induced mtDNA leakage in EC cells, which further caused the activation of the downstream cGAS-STING signaling and increased production of extracellular vesicle (EV) production from EC cells. Besides, the activation of cGAS-STING signaling enhanced the expression of type I IFN and its downstream molecule PD-L1 through the TBK1-IRF3 pathway. Importantly, a high level mtDNA and PD-L1 were present in EVs derived from IL6-induced EC cells; these vesicles were shown to be able to induce T cell apoptosis. Finally, anti-PD-L1 treatment in mice showed that blockade of PD-L1 significantly reversed tumor immune escape mediated by IL6-induced EVs. Together, we provide evidence that IL6 induced mtDNA leakage to regulate the immune escape of EC cells. Our findings may provide a novel clue for the development of therapeutic targets for EC.

DOI: https://doi.org/10.1155/2022/3815853

Cancer Lett. 2022 Jun 11. pii: S0304-3835(22)00267-1. [Epub ahead of print] 215783

Fangchinoline inhibits non-small cell lung cancer metastasis by reversing epithelial-mesenchymal transition and suppressing the cytosolic ROS-related Akt-mTOR signaling pathway.

Bonan Chen, Yue Song, Yujuan Zhan, Shikun Zhou, Junzi Ke, Weizhen Ao, Yigan Zhang, Qiqi Liang, Minhui He, Shuhui Li, Fuda Xie, Haonan Huang, Wai Nok Chan, Alvin H K Cheung, Brigette B Y Ma, Wei Kang, Ka Fai To, Jianyong Xiao.

Few drugs alleviate non-small cell lung cancer (NSCLC) metastasis effectively. Small molecular screening demonstrated that fangchinoline (Fan) reversed epithelial-mesenchymal transition (EMT) in NSCLC cells, inhibiting cell invasion and migration. RNA sequencing (RNA-seq) of Fan-treated NSCLC cells revealed that Fan potently quenched the NADP+ metabolic process. Molecular docking analysis revealed that Fan directly and specifically targeted NOX4. NOX4 was associated with poor prognosis in NSCLC in both The Cancer Genome Atlas (TCGA) and Hong Kong cohorts. In mitochondrial DNA-depleted ρ0 NSCLC cells, Fan decreased cytosolic reactive oxygen species (ROS) to inhibit the Akt-mTOR signaling pathway by directly promoting NOX4 degradation. In TCGA and Hong Kong cohorts, NOX4 upregulation acted as a driver event as it positively correlated with metastasis and oxidative stress. Single-cell RNA-seq indicated that NOX4 was overexpressed, especially in cancer cells, cancer stem cells, and endothelial cells. In mice, Fan significantly impeded subcutaneous xenograft formation and reduced metastatic nodule numbers in mouse lung and liver. Drug sensitivity testing demonstrated that Fan suppressed patient-derived organoid growth dose-dependently. Fan is a potent small molecule for alleviating NSCLC metastasis by directly targeting NOX4 and is a potential novel therapeutic agent.

Keywords: Cytosolic ROS; Fangchinoline; NOX4; Non-small cell lung cancer

DOI: https://doi.org/10.1016/j.canlet.2022.215783

Front Immunol. 2022 ;13 853848

MicroRNA-182-5p Attenuates Asthmatic Airway Inflammation by Targeting NOX4.

Zhiguang Wang, Yilan Song, Jingzhi Jiang, Yihua Piao, Li Li, Qiaoyun Bai, Chang Xu, Hanye Liu, Liangchang Li, Hongmei Piao, Guanghai Yan.

Bronchial asthma is characterized by chronic airway inflammation, airway hyperresponsiveness, and airway remodeling. MicroRNA (miRNA) has recently been implicated in the pathogenesis of asthma. However, the mechanisms of different miRNAs in asthma are complicated, and the mechanism of miRNA-182-5p in asthma is still unclear. Here, we aim to explore the mechanism of miRNA182-5p in asthma-related airway inflammation. Ovalbumin (OVA)-induced asthma model was established. MiRNA Microarray Analysis was performed to analyze the differentially expressed miRNAs in the asthma model. We found that the expression of miRNA-182-5p was significantly decreased in OVA-induced asthma. In vitro, IL-13 stimulation of BEAS-2B cells resulted in a significant up-regulation of NOX4 (nicotinamide adenine dinucleotide phosphate oxidase 4), accompanied by mitochondrial damage-induced apoptosis, NLRP3 (NOD-like receptor family pyrin domain-containing 3)/IL-1β activation, and reduced miRNA-182-5p. In contrast, overexpression of miRNA-182-5p significantly inhibited epithelial cell apoptosis and NLRP3/IL-1β activation. In addition, we found that miRNA-182-5p could bind to the 3' untranscripted region of NOX4 mRNA and inhibit epithelial cell inflammation by reducing oxidative stress and mitochondrial damage. In vivo, miRNA-182-5p agomir treatment significantly reduced the percentage of eosinophils in bronchoalveolar lavage fluid, and down-regulated Th2 inflammatory factors, including IL-4, IL-5, and OVA induced IL-13. Meanwhile, miRNA-182-5p agomir reduced the peribronchial inflammatory cell infiltration, goblet cell proliferation and collagen deposition. In summary, targeting miRNA-182-5p may provide a new strategy for the treatment of asthma.

Keywords: NOX4; Ovalbumin (OVA); airway inflammation; asthma; miR-182-5p

DOI: https://doi.org/10.3389/fimmu.2022.853848