bims-noxint 2022-08-21 papers

Issue of 2022–08–21
three papers selected by
Laia Caja Puigsubira, Uppsala University

Free Radic Biol Med. 2022 Aug 12. pii: S0891-5849(22)00516-0. [Epub ahead of print]190 158-168

Spatial, temporal, and cell-type-specific expression of NADPH Oxidase isoforms following seizure models in rats.

Aseel Saadi, Sereen Sandouka, Etty Grad, Prince Kumar Singh, Tawfeeq Shekh-Ahmad.

The NADPH Oxidase (NOX) enzymes are key producers of reactive oxygen species (ROS) and consist of seven different isoforms, distributed across the tissues and cell types. The increasing level of ROS induces oxidative stress playing a crucial role in neuronal death and the development of epilepsy. Recently, NOX2 was reported as a primary source of ROS production, activated by NMDA receptor, a crucial marker of epilepsy development. Here, we demonstrate spatial, temporal, and cellular expression of NOX2 and NOX4 complexes in in-vitro and in-vivo seizure models. We showed that the expression of NOX2 and NOX4 was increased in the initial 24 h following a brief seizure induced by pentylenetetrazol. Interestingly, while this elevated level returns to baseline 48 h following seizure in the cortex, in the hippocampus these levels remain elevated up to one week following the seizure. Moreover, we showed that 1- and 2- weeks following status epilepticus (SE), expression of NOX2 and NOX4 remains significantly elevated both in the cortex and the hippocampus. Furthermore, in in-vitro seizure model, NOX2 and NOX4 isoforms were overexpressed in neurons and astrocytes following seizures. These results suggest that NOX2 and NOX4 in the brain have a transient response to seizures, and these responses temporally vary depending on, seizure duration, brain region (cortex or hippocampus), and cell types.

Keywords: Epilepsy; Kainic acid; NADPH Oxidase; Pentylenetetrazol; Reactive oxygen species

DOI: https://doi.org/10.1016/j.freeradbiomed.2022.08.009

Kidney Res Clin Pract. 2022 Jul 19.

NADPH oxidase inhibitor development for diabetic nephropathy through water tank model.

Hye Eun Lee, Seunghwan Shim, Yongseok Choi, Yun Soo Bae.

Oxidative stress can cause generation of uncontrolled reactive oxygen species (ROS) and lead to cytotoxic damage to cells and tissues. Recently, it has been shown that transient ROS generation can serve as a secondary messenger in receptor-mediated cell signaling. Although excessive levels of ROS are harmful, moderated levels of ROS are essential for normal physiological function. Therefore, regulating cellular ROS levels should be an important concept for development of novel therapeutics for treating diseases. The overexpression and hyperactivation of NADPH oxidase (Nox) can induce high levels of ROS, which are strongly associated with diabetic nephropathy. This review discusses the theoretical basis for development of the Nox inhibitor as a regulator of ROS homeostasis to provide emerging therapeutic opportunities for diabetic nephropathy.

Keywords: Diabetic nephropathies; NADPH oxidases; Nox inhibitor; Oxidative stress; Reactive oxygen species

DOI: https://doi.org/10.23876/j.krcp.21.269

Front Cardiovasc Med. 2022 ;9 942736

Coupling of phagocytic NADPH oxidase activity and mitochondrial superoxide production.

Sergey I Dikalov, Anna E Dikalova, Igor A Kirilyuk.

Superoxide radical plays an important role in redox cell signaling and physiological processes; however, overproduction of superoxide or insufficient activity of antioxidants leads to oxidative stress and contributes to the development of pathological conditions such as endothelial dysfunction and hypertension. Meanwhile, the studies of superoxide in biological systems represent unique challenges associated with short lifetime of superoxide, insufficient reactivity of the superoxide probes, and lack of site-specific detection of superoxide. In this work we have developed 15N-and deuterium-enriched spin probe 15N-CAT1H for high sensitivity and site-specific detection of extracellular superoxide. We have tested simultaneous tracking of extracellular superoxide by 15N-CAT1H and intramitochondrial superoxide by conventional 14N-containing spin probe mitoTEMPO-H in immune cells isolated from spleen, splenocytes, under basal conditions or stimulated with inflammatory cytokines IL-17A and TNFα, NADPH oxidase activator PMA, or treated with inhibitors of mitochondrial complex I rotenone or complex III antimycin A. 15N-CAT1H provides two-fold increase in sensitivity and improves detection since EPR spectrum of 15N-CAT1 nitroxide does not overlap with biological radicals. Furthermore, concurrent use of cell impermeable 15N-CAT1H and mitochondria-targeted 14N-mitoTEMPO-H allows simultaneous detection of extracellular and mitochondrial superoxide. Analysis of IL-17A- and TNFα-induced superoxide showed parallel increase in 15N-CAT1 and 14N-mitoTEMPO signals suggesting coupling between phagocytic NADPH oxidase and mitochondria. The interplay between mitochondrial superoxide production and activity of phagocytic NADPH oxidase was further investigated in splenocytes isolated from Sham and angiotensin II infused C57Bl/6J and Nox2KO mice. Angiotensin II infusion in wild-type mice increased the extracellular basal splenocyte superoxide which was further enhanced by complex III inhibitor antimycin A, mitochondrial uncoupling agent CCCP and NADPH oxidase activator PMA. Nox2 depletion attenuated angiotensin II mediated stimulation and inhibited both extracellular and mitochondrial PMA-induced superoxide production. These data indicate that splenocytes isolated from hypertensive angiotensin II-infused mice are "primed" for enhanced superoxide production from both phagocytic NADPH oxidase and mitochondria. Our data demonstrate that novel 15N-CAT1H provides high sensitivity superoxide measurements and combination with mitoTEMPO-H allows independent and simultaneous detection of extracellular and mitochondrial superoxide. We suggest that this new approach can be used to study the site-specific superoxide production and analysis of important sources of oxidative stress in cardiovascular conditions.

Keywords: electron paramagnetic resonance (EPR); mitochondria; phagocytic NADPH oxidase; spin probe; superoxide

DOI: https://doi.org/10.3389/fcvm.2022.942736