NADPH oxidase inhibition rescues keratinocytes from elevated oxidative stress in a 2D atopic dermatitis and psoriasis model

Emmert, Hila; Fonfara, Melina; Rodríguez, Elke; Weidinger, Stephan

doi:10.1111/exd.14148

Cited by 25 publications

(17 citation statements)

References 51 publications

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“…Other NOX isoforms could also contribute to oxidative stress. In vitro studies have demonstrated that NOX4 is the main ROS source in human psoriatic fibroblasts and is essential for redox-mediated modulation of the metabolism in keratinocytes 90 , while inhibition of NOX1 in keratinocytes derived from the skin of atopic dermatitis patients significantly reduced ROS production 91 . Furthermore, NOX4 is highly expressed in endothelial cells and is involved in osteoarthritis, renal diseases and angiogenesis during ischemia, hypoxia and inflammation 92 - 94 .…”

Section: Discussionmentioning

confidence: 99%

Immunomodulatory role of reactive oxygen species and nitrogen species during T cell-driven neutrophil-enriched acute and chronic cutaneous delayed-type hypersensitivity reactions

et al. 2021

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Rationale: Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are important regulators of inflammation. The exact impact of ROS/RNS on cutaneous delayed-type hypersensitivity reaction (DTHR) is controversial. The aim of our study was to identify the dominant sources of ROS/RNS during acute and chronic trinitrochlorobenzene (TNCB)-induced cutaneous DTHR in mice with differently impaired ROS/RNS production. Methods: TNCB-sensitized wild-type, NADPH oxidase 2 (NOX2)- deficient (gp91 phox-/- ), myeloperoxidase-deficient (MPO -/- ), and inducible nitric oxide synthase-deficient (iNOS -/- ) mice were challenged with TNCB on the right ear once to elicit acute DTHR and repetitively up to five times to induce chronic DTHR. We measured ear swelling responses and noninvasively assessed ROS/RNS production in vivo by employing the chemiluminescence optical imaging (OI) probe L-012. Additionally, we conducted extensive ex vivo analyses of inflamed ears focusing on ROS/RNS production and the biochemical and morphological consequences. Results: The in vivo L-012 OI of acute and chronic DTHR revealed completely abrogated ROS/RNS production in the ears of gp91 phox-/- mice, up to 90 % decreased ROS/RNS production in the ears of MPO -/- mice and unaffected ROS/RNS production in the ears of iNOS -/- mice. The DHR flow cytometry analysis of leukocytes derived from the ears with acute DTHR confirmed our in vivo L-012 OI results. Nevertheless, we observed no significant differences in the ear swelling responses among all the experimental groups. The histopathological analysis of the ears of gp91 phox-/- mice with acute DTHRs revealed slightly enhanced inflammation. In contrast, we observed a moderately reduced inflammatory immune response in the ears of gp91 phox-/- mice with chronic DTHR, while the inflamed ears of MPO -/- mice exhibited the strongest inflammation. Analyses of lipid peroxidation, 8-hydroxy-2'deoxyguanosine levels, redox related metabolites and genomic expression of antioxidant proteins revealed similar oxidative stress in all experimental groups. Furthermore, inflamed ears of wild-type and gp91 phox-/- mice displayed neutrophil extracellular trap (NET) formation exclusively in acute but not chronic DTHR. Conclusions: MPO and NOX2 are the dominant sources of ROS/RNS in acute and chronic DTHR. Nevertheless, depletion of one primary source of ROS/RNS exhibited only marginal but conflicting impact on acute and chronic cutaneous DTHR. Thus, ROS/RNS are not a single entity, and each species has different properties at certain stages of the disease, resulting in different outcomes.

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Section: Discussionmentioning

confidence: 99%

Immunomodulatory role of reactive oxygen species and nitrogen species during T cell-driven neutrophil-enriched acute and chronic cutaneous delayed-type hypersensitivity reactions

et al. 2021

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“…Psoriasis is commonly treated by targeting TNFα signalling (Chima & Lebwohl, 2018), and Nox1 inhibition reduced inflammation and improved human keratinocyte survival in an experimental model of psoriasis (Emmert et al . 2020). The expression changes favouring 8D over 8C in psoriatic skin again suggest a compensatory response to downregulate pro‐inflammatory Nox‐dependent signalling.…”

Section: Discussionmentioning

confidence: 99%

Oxidant‐resistant LRRC8A/C anion channels support superoxide production by NADPH oxidase 1

Choi

Rohrbough

Nguyen

et al. 2021

The Journal of Physiology

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Key points LRRC8A‐containing anion channels associate with NADPH oxidase 1 (Nox1) and regulate superoxide production and tumour necrosis factor‐α (TNFα) signalling. Here we show that LRRC8C and 8D also co‐immunoprecipitate with Nox1 in vascular smooth muscle cells. LRRC8C knockdown inhibited TNFα‐induced O2•− production, receptor endocytosis, nuclear factor‐κB (NF‐κB) activation and proliferation while LRRC8D knockdown enhanced NF‐κB activation. Significant changes in LRRC8 isoform expression in human atherosclerosis and psoriasis suggest compensation for increased inflammation. The oxidant chloramine‐T (ChlorT, 1 mM) weakly (∼25%) inhibited LRRC8C currents but potently (∼80%) inhibited LRRC8D currents. Substitution of the extracellular loop (EL1, EL2) domains of 8D into 8C conferred significantly stronger (69%) ChlorT‐dependent inhibition. ChlorT exposure impaired subsequent current block by DCPIB, which occurs through interaction with EL1, further implicating external oxidation sites. LRRC8A/C channels most effectively sustain Nox1 activity at the plasma membrane. This may result from their ability to remain active in an oxidized microenvironment. Abstract Tumour necrosis factor‐α (TNFα) activates NADPH oxidase 1 (Nox1) in vascular smooth muscle cells (VSMCs), producing superoxide (O2•–) required for subsequent signalling. LRRC8 family proteins A–E comprise volume‐regulated anion channels (VRACs). The required subunit LRRC8A physically associates with Nox1, and VRAC activity is required for Nox activity and the inflammatory response to TNFα. VRAC currents are modulated by oxidants, suggesting that channel oxidant sensitivity and proximity to Nox1 may play a physiologically relevant role. In VSMCs, LRRC8C knockdown (siRNA) recapitulated the effects of siLRRC8A, inhibiting TNFα‐induced extracellular and endosomal O2•− production, receptor endocytosis, nuclear factor‐κB (NF‐κB) activation and proliferation. In contrast, siLRRC8D potentiated NF‐κB activation. Nox1 co‐immunoprecipitated with 8C and 8D, and colocalized with 8D at the plasma membrane and in vesicles. We compared VRAC currents mediated by homomeric and heteromeric LRRC8C and LRRC8D channels expressed in HEK293 cells. The oxidant chloramine T (ChlorT, 1 mM) weakly inhibited 8C, but potently inhibited 8D currents. ChlorT exposure also impaired subsequent current block by the VRAC blocker DCPIB, implicating external sites of oxidation. Substitution of the 8D extracellular loop domains (EL1, EL2) into 8C conferred significantly stronger ChlorT‐mediated inhibition of 8C currents. Our results suggest that LRRC8A/C channel activity can be effectively maintained in the oxidized microenvironment expected to result from Nox1 activation at the plasma membrane. Increased ratios of 8D:8C expression may potentially depress inflammatory responses to TNFα. LRRC8A/C channel downregulation represents a novel strategy to reduce TNFα‐induced inflammation.

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“…Psoriasis is a skin disease commonly treated by targeting TNFα signaling (Chima & Lebwohl, 2018). Inhibition of Nox1 reduced inflammation and improved human keratinocyte survival in an experimental model of psoriasis (Emmert et al ., 2020). Thus, the observed expression changes favoring LRRC8D over LRRC8C in psoriatic skin are again consistent with downregulation of Nox-dependent inflammation.…”

Section: Discussionmentioning

confidence: 99%

Oxidant-Resistant LRRC8A/C Anion Channels Support Superoxide Production by Nox1

Rohrbough

Choi

Nguyen

et al. 2021

Preprint

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Tumor necrosis factor-α (TNFα) activates NADPH Oxidase 1 (Nox1) in vascular smooth muscle cells (VSMCs), producing superoxide species required for subsequent signaling. LRRC8 family proteins A-E comprise volume-regulated anion channels (VRACs). The required subunit LRRC8A physically associates with Nox1, and VRAC activity is required for Nox activity and the inflammatory response to TNFα. LRRC8 channel currents are modulated by oxidants, suggesting that oxidant sensitivity and proximity to Nox1 may play a physiologically relevant role. In VSMCs, LRRC8C knockdown (siRNA) recapitulated the effects of siLRRC8A, inhibiting TNFα-induced extracellular and endosomal superoxide production, receptor endocytosis, NF-κB activation, and proliferation. In contrast, siLRRC8D potentiated NF-κB activation. Nox1 co-immunoprecipitated with 8C and 8D, and co-localized with 8D at the plasma membrane and in vesicles. We compared VRAC currents mediated by homomeric and heteromeric 8C and 8D channels expressed in HEK293 cells. The oxidant chloramine T (ChlorT, 1 mM) weakly inhibited LRRC8C, but potently inhibited 8D currents. ChlorT exposure also greatly reduced subsequent current block by DCPIB, implicating external sites of oxidation. Substitution of the extracellular loop domains (EL1, EL2) of 8D onto 8C conferred significantly stronger ChlorT-dependent inhibition. 8A/C channel activity is thus more effectively maintained in the oxidized microenvironment expected to result from Nox1 activation at the plasma membrane. Increased ratios of 8D:8C expression may potentially depress inflammatory responses to TNFα. LRRC8A/C channel downregulation represents a novel strategy to reduce TNFα-induced inflammation.

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NADPH oxidase inhibition rescues keratinocytes from elevated oxidative stress in a 2D atopic dermatitis and psoriasis model

Cited by 25 publications

References 51 publications

Immunomodulatory role of reactive oxygen species and nitrogen species during T cell-driven neutrophil-enriched acute and chronic cutaneous delayed-type hypersensitivity reactions

Immunomodulatory role of reactive oxygen species and nitrogen species during T cell-driven neutrophil-enriched acute and chronic cutaneous delayed-type hypersensitivity reactions

Oxidant‐resistant LRRC8A/C anion channels support superoxide production by NADPH oxidase 1

Oxidant-Resistant LRRC8A/C Anion Channels Support Superoxide Production by Nox1

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