Prevent ROS Production Research Articles

Tetramethylpyrazine attenuates renal tubular epithelial cell ferroptosis in contrast-induced nephropathy by inhibiting transferrin receptor and intracellular reactive oxygen species.

Contrast-induced nephropathy (CIN) is a leading cause of hospital-acquired acute kidney injury (AKI). Recently, ferroptosis was reported to be crucial for AKI pathogenesis. Our previous studies indicated antioxidant tetramethylpyrazine (TMP) prevent CIN in vivo. However, whether ferroptosis is involved in TMP nephroprotective mechanism against CIN is unclear. In the present study, we investigated the role of renal tubular epithelial cell ferroptosis in TMP reno-protective effect against CIN and the molecular mechanisms by which TMP regulates ferroptosis. Classical contrast-medium, Iohexol, was used to construct CIN models in rats and HK-2 cells. Results showed that tubular cell injury was accompanied by ferroptosis both in vivo and in vitro, including the typical features of ferroptosis, Fe2+ accumulation, lipid peroxidation and decreased glutathione peroxidase 4 (GPX4). Ferroptosis inhibition by classic inhibitors Fer-1 and DFO promoted cell viability and reduced intracellular ROS production. Additionally, TMP significantly inhibited renal dysfunction, reduced AKI biomarkers, prevented ROS production, inhibited renal Fe2+ accumulation and increased GPX4 expression. Expressions of various proteins associated with iron ion metabolism, including transferrin receptor (TFRC), divalent metal transporter 1, iron-responsive element binding protein 2, ferritin heavy chain 1, ferroportin 1, and heat shock factor binding protein 1, were examined using mechanistic analyses. Among these, TFRC changes were the most significant after TMP pretreatment. Results of siRNA knockdown and plasmid overexpression of TFRC indicated that TFRC is essential for TMP to alleviate ferroptosis and reduce LDH release, Fe2+ accumulation and intracellular ROS. Our findings provide crucial insights about the potential of TMP in treating AKI associated with ferroptosis.

Mitochondrial dysfunction and its association with age-related disorders.

Aging is a complex process that features a functional decline in many organelles. Various factors influence the aging process, such as chromosomal abnormalities, epigenetic changes, telomere shortening, oxidative stress, and mitochondrial dysfunction. Mitochondrial dysfunction significantly impacts aging because mitochondria regulate cellular energy, oxidative balance, and calcium levels. Mitochondrial integrity is maintained by mitophagy, which helps maintain cellular homeostasis, prevents ROS production, and protects against mtDNA damage. However, increased calcium uptake and oxidative stress can disrupt mitochondrial membrane potential and permeability, leading to the apoptotic cascade. This disruption causes increased production of free radicals, leading to oxidative modification and accumulation of mitochondrial DNA mutations, which contribute to cellular dysfunction and aging. Mitochondrial dysfunction, resulting from structural and functional changes, is linked to age-related degenerative diseases. This review focuses on mitochondrial dysfunction, its implications in aging and age-related disorders, and potential anti-aging strategies through targeting mitochondrial dysfunction.

Rhinacanthin-C but Not -D Extracted from Rhinacanthus nasutus (L.) Kurz Offers Neuroprotection via ERK, CHOP, and LC3B Pathways.

Neurodegenerative diseases present an increasing problem as the world’s population ages; thus, the discovery of new drugs that prevent diseases such as Alzheimer’s, and Parkinson’s diseases are vital. In this study, Rhinacanthin-C and -D were isolated from Rhinacanthus nasustus, using ethyl acetate, followed by chromatography to isolate Rhinacanthin-C and -D. Both compounds were confirmed using NMR and ultra-performance-LCMS. Using glutamate toxicity in HT-22 cells, we measured cell viability and apoptosis, ROS build-up, and investigated signaling pathways. We show that Rhinacanthin-C and 2-hydroxy-1,4-naphthoquinone have neuroprotective effects against glutamate-induced apoptosis in HT-22 cells. Furthermore, we see that Rhinacanthin-C resulted in autophagy inhibition and increased ER stress. In contrast, low concentrations of Rhinacanthin-C and 2-hydroxy-1,4-naphthoquinone prevented ER stress and CHOP expression. All concentrations of Rhinacanthin-C prevented ROS production and ERK1/2 phosphorylation. We conclude that, while autophagy is present in HT-22 cells subjected to glutamate toxicity, its inhibition is not necessary for cryoprotection.

Carajurin Induces Apoptosis in Leishmania amazonensis Promastigotes through Reactive Oxygen Species Production and Mitochondrial Dysfunction.

Carajurin is the main constituent of Arrabidaea chica species with reported anti-Leishmania activity. However, its mechanism of action has not been described. This study investigated the mechanisms of action of carajurin against promastigote forms of Leishmania amazonensis. Carajurin was effective against promastigotes with IC50 of 7.96 ± 1.23 μg.mL−1 (26.4 µM), and the cytotoxic concentration for peritoneal macrophages was 258.2 ± 1.20 μg.mL−1 (856.9 µM) after 24 h of treatment. Ultrastructural evaluation highlighted pronounced swelling of the kinetoplast with loss of electron-density in L. amazonensis promastigotes induced by carajurin treatment. It was observed that carajurin leads to a decrease in the mitochondrial membrane potential (p = 0.0286), an increase in reactive oxygen species production (p = 0.0286), and cell death by late apoptosis (p = 0.0095) in parasites. Pretreatment with the antioxidant NAC prevented ROS production and significantly reduced carajurin-induced cell death. The electrochemical and density functional theory (DFT) data contributed to support the molecular mechanism of action of carajurin associated with the ROS generation, for which it is possible to observe a correlation between the LUMO energy and the electroactivity of carajurin in the presence of molecular oxygen. All these results suggest that carajurin targets the mitochondria in L. amazonensis. In addition, when assessed for its drug-likeness, carajurin follows Lipinski’’s rule of five, and the Ghose, Veber, Egan, and Muegge criteria.

Smooth muscle Rac1 contributes to pulmonary hypertension

Pulmonary arterial hypertension (PAH) is a multifactorial and chronic disease characterized by an increase in pulmonary artery (PA) resistance leading to right ventricle (RV) failure. Endothelial dysfunction and alteration NO/cGMP signaling in PA play a major role in PAH. We described the involvement of Rac1 in the control of systemic blood pressure through its involvement in NO-mediated relaxation of arterial smooth muscle cell (SMC). The aim of this study was to decipher the role of SMC Rac1 in PAH. Rac1 activation, Rac-GTP, is determined by immunofluorescence in lungs biopsies from control and PAH patients. PAH is induced by exposure of wild-type (WT) and SMC Rac1-deficient mice (SM-Rac1-KO mice) to chronic hypoxia (10% O 2 , 4 weeks). PAH is assessed by the measurement of RV systolic pressure and hypertrophy. The proliferation of pulmonary arterial SMC (PASMC) in hypoxic condition is measured by EdU staining. Oxidative stress is assessed by immunofluorescence using the DHE probe and by electron spin resonance (ESR). In biopsies from PAH patients, we detected an increase (3-fold) of Rac-GTP in PA compared to control ( n = 4–6). To decipher the role of Rac activation in PAH, WT and SM-Rac1-KO mice were exposed to hypoxia to induce PAH. The elevation of systolic pressure in RV and RV hypertrophy were decreased by 32.5% and 60% ( n = 5–7) in SM-Rac1-KO mice. Deletion of Rac1 in SMC reduced the hypoxia-induced remodeling. In vitro, we observed that hypoxia-induced PASMC proliferation is decreased by 50% by the Rac1 inhibitor, EHT1864. We demonstrated that Rac1 inhibition or deletion in SMC prevent ROS production in PASMC and limit PASMC proliferation and vascular remodeling. Our results suggest that Rac1 activity in PASMC plays a causal role in PAH by favoring ROS dependent PA remodelling induced by chronic hypoxia. Therefore, Rac1 deletion or inhibition protect against hypoxic PAH.

Oral administration of NSP-116, a free radical scavenger, suppresses the symptoms of retinal vein occlusion in the murine model

Retinal vein occlusion (RVO) is an intractable eye disease that results in reduced visual acuity, associated with retinal ischemia, hemorrhage, and edema. RVO results in excessive ROS production in the retina, causing inflammation and retinal edema. A free radical scavenger, 4-(4-acetylpiperazin-1-yl)-2-(1H-imidazole-1-yl) aniline (NSP-116), has been reported to demonstrate antioxidative effects and prevent ROS production in the retina. Therefore, NSP-116 may represent a useful drug for treating the pathological symptoms of RVO, such as retinal edema and ischemic symptoms. This study aimed to investigate the effects of NSP-116 in a murine model of RVO. We evaluated the thickness of the retinal layer and the size of the non-perfused area following the oral administration of NSP-116. Moreover, we used western blot analysis to examine the expression levels of vascular endothelial growth factor (VEGF) and tumor necrosis factor (TNF)-α, after NSP-116 administration, and examined the localization of 8-hydroxy-2′-deoxyguanosine (8-OHdG), by immunostaining. The findings indicate that NSP-116 suppressed retinal edema and expansion the non-perfused area by suppressing the increased expression of VEGF, TNF-α, and 8-OHdG in the murine RVO model. In conclusion, the oral administration of NSP-116 may serve as an effective pharmacological treatment for the pathological symptoms of RVO.

Celecoxib Exerts Neuroprotective Effects in β-Amyloid-Treated SH-SY5Y Cells Through the Regulation of Heme Oxygenase-1: Novel Insights for an Old Drug.

The formation and aggregation of amyloid-β-peptide (Aβ) into soluble and insoluble species represent the pathological hallmarks of Alzheimer’s disease (AD). Over the last few years, however, soluble Aβ (sAβ) prevailed over fibrillar Aβ (fAβ) as determinant of neurotoxicity. One of the main therapeutic strategies for challenging neurodegeneration is to fight against neuroinflammation and prevent free radical-induced damage: in this light, the heme oxygenase/biliverdin reductase (HO/BVR) system is considered a promising drug target. The aim of this work was to investigate whether or not celecoxib (CXB), a selective inhibitor of the pro-inflammatory cyclooxygenase-2, modulates the HO/BVR system and prevents lipid peroxidation in SH-SY5Y neuroblastoma cells. Both sAβ (6.25–50 nM) and fAβ (1.25–50 nM) dose-dependently over-expressed inducible HO (HO-1) after 24 h of incubation, reaching statistical significance at 25 and 6.25 nM, respectively. Interestingly, CXB (1–10 μM, for 1 h) further enhanced Aβ-induced HO-1 expression through the nuclear translocation of the transcriptional factor Nrf2. Furthermore, 10 μM CXB counteracted the Aβ-induced ROS production with a mechanism fully dependent on HO-1 up-regulation; nevertheless, 10 μM CXB significantly counteracted only 25 nM sAβ-induced lipid peroxidation damage in SH-SY5Y neurons by modulating HO-1. Both carbon monoxide (CORM-2, 50 nM) and bilirubin (50 nM) significantly prevented ROS production in Aβ-treated neurons and favored both the slowdown of the growth rate of Aβ oligomers and the decrease in oligomer/fibril final size. In conclusion, these results suggest a novel mechanism through which CXB is neuroprotective in subjects with early AD or mild cognitive impairment.

Effects of hydrogen peroxide, doxorubicin and ultraviolet irradiation on senescence of human dental pulp stem cells

ObjectiveThe objective of this study was to evaluate the ability of three distinct agents on the induction of senescence in human dental pulp stem cells (DPSCs). DesignDPSCs from three separate donors were treated with H2O2, doxorubicin and ultraviolet (UV) irradiation. The response of the cells to the three agents was assayed by specific staining for SA-βGal, RT-qPCR and flow cytometry. ResultsThe results showed that incubation with 100 μM H2O2 and 20 nM Doxorubicin for seven days led to senescence in all donors’ cells equally. Interestingly, UV irradiation for just one minute was sufficient to induce senescence in the cells. The SA-βGal positive senescent cells were arrested in G1 phase and their S phase was significantly reduced as analyzed by flow cytometry. Significant increment in p21 and BTG1 expression and decrement in CCND1 expression also confirmed the cells have been arrested and get senescent via p53-p21 pathway. ConclusionAll three agents successfully triggered senescence in the cells. There was no significant difference in the capacity of the three donor’s cells for senescence. To avoid premature senescence in stem cell in vitro, it is recommended to avoid unnecessary exposure of the cell to fluorescent and UV light. Moreover, to prevent ROS production, we recommend using a separate incubator with low oxygen content for cell culture, if possible.

IMPAD1 functions as mitochondrial electron transport inhibitor that prevents ROS production and promotes lung cancer metastasis through the AMPK-Notch1-HEY1 pathway.

The tumor microenvironment (TME) and metabolic reprogramming have been implicated in cancer development and progression. However, the link between TME, metabolism, and cancer progression in lung cancer is unclear. In the present study, we identified IMPAD1 from the conditioned medium of highly invasive CL1-5. High expression of IMPAD1 was associated with a poorer clinical phenotype in lung cancer patients, with reduced survival and increased lymph node metastasis. Knockdown of IMPAD1 significantly inhibited migration/invasion abilities and metastasis in vitro and in vivo. Upregulation of IMPAD1 and subsequent accumulation of AMP in cells increased the pAMPK, leading to Notch1 and HEY1 upregulation. As AMP is an ADORA1 agonist, treatment with ADORA1 inhibitor reduced the expression of pAMPK and HEY1 expression in IMPAD1-overexpressing cells. IMPAD1 caused mitochondria dysfunction by inhibiting mitochondrial Complex I activity, which reduced mitochondrial ROS levels and activated the AMPK-HEY1 pathway. Collectively this study supports the multipotent role of IMPAD1 in promotion of lung cancer metastasis by simultaneously increasing AMP levels, inhibition of Complex I activity to decrease ROS levels, thereby activating AMPK-Notch1-HEY1 signaling, and providing an alternative metabolic pathway in energy stress conditions.

Mitochondrial respirasome works as a single unit and the cross-talk between complexes I, III2 and IV stimulates NADH dehydrogenase activity

Ustilago maydis is an aerobic basidiomycete that depends on oxidative phosphorylation for its ATP supply, pointing to the mitochondrion as a key player in its energy metabolism. Mitochondrial respiratory complexes I, III2, and IV occur in supramolecular structures named respirasome. In this work, we characterized the subunit composition and the kinetics of NADH:Q oxidoreductase activity of the digitonine-solubilized respirasome (1600 kDa) and the free-complex I (990 kDa). In the presence of 2,6-dimethoxy-1,4-benzoquinone (DBQ) and cytochrome c, both the respirasome NADH:O2 and the NADH:DBQ oxidoreductase activities were inhibited by rotenone, antimycin A or cyanide. A value of 2.4 for the NADH oxidized/oxygen reduced ratio was determined for the respirasome activity, while ROS production was less than 0.001% of the oxygen consumption rate. Analysis of the NADH:DBQ oxidoreductase activity showed that respirasome was 3-times more active and showed higher affinity than free-complex I. The results suggest that the contacts between complexes I, III2 and IV in the respirasome increase the catalytic efficiency of complex I and regulate its activity to prevent ROS production.

Pressure-dependent NOS activation contributes to endothelial hyperpermeability in a model of acute heart failure

Aims: Acute increases in left ventricular end diastolic pressure (LVEDP) can induce pulmonary edema (PE). The mechanism(s) for this rapid onset edema may involve more than just increased fluid filtration. Lung endothelial cell permeability is regulated by pressure-dependent activation of nitric oxide synthase (NOS). Herein, we demonstrate that pressure-dependent NOS activation contributes to vascular failure and PE in a model of acute heart failure (AHF) caused by hypertension.Methods and results: Male Sprague–Dawley rats were anesthetized and mechanically ventilated. Acute hypertension was induced by norepinephrine (NE) infusion and resulted in an increase in LVEDP and pulmonary artery pressure (Ppa) that were associated with a rapid fall in PaO2, and increases in lung wet/dry ratio and injury scores. Heart failure (HF) lungs showed increased nitrotyrosine content and ROS levels. L-NAME pretreatment mitigated the development of PE and reduced lung ROS concentrations to sham levels. Apocynin (Apo) pretreatment inhibited PE. Addition of tetrahydrobiopterin (BH4) to AHF rats lung lysates and pretreatment of AHF rats with folic acid (FA) prevented ROS production indicating endothelial NOS (eNOS) uncoupling.Conclusion: Pressure-dependent NOS activation leads to acute endothelial hyperpermeability and rapid PE by an increase in NO and ROS in a model of AHF. Acute increases in pulmonary vascular pressure, without NOS activation, was insufficient to cause significant PE. These results suggest a clinically relevant role of endothelial mechanotransduction in the pathogenesis of AHF and further highlights the concept of active barrier failure in AHF. Therapies targetting the prevention or reversal of endothelial hyperpermeability may be a novel therapeutic strategy in AHF.

Frontispiece: N4‐Tetradentate Chelators Efficiently Regulate Copper Homeostasis and Prevent ROS Production Induced by Copper‐Amyloid‐β1–16

Frontispiece: N₄‐Tetradentate Chelators Efficiently Regulate Copper Homeostasis and Prevent ROS Production Induced by Copper‐Amyloid‐β_1–16

Anthocyanins Improve Hippocampus-Dependent Memory Function and Prevent Neurodegeneration via JNK/Akt/GSK3β Signaling in LPS-Treated Adult Mice.

Microglia plays a critical role in the brain and protects neuronal cells from toxins. However, over-activation of microglia leads to deleterious effects. Lipopolysaccharide (LPS) has been reported to affect neuronal cells via activation of microglia as well as directly to initiate neuroinflammation. In the present study, we evaluated the anti-inflammatory and anti-oxidative effect of anthocyanins against LPS-induced neurotoxicity in an animal model and in cell cultures. Intraperitoneal injections of LPS (250μg/kg/day for 1week) induce ROS production and promote neuroinflammation and neurodegeneration which ultimately leads to memory impairment. However, anthocyanins treatment at a dose of 24mg/kg/day for 2weeks (1week before and 1week co-treated with LPS) prevented ROS production, inhibited neuroinflammation and neurodegeneration, and improved memory functions in LPS-treated mice. Both histological and immunoblot analysis indicated that anthocyanins reversed the activation of JNK, prevented neuroinflammation by lowering the levels of inflammatory markers (p-NF-kB, TNF-α, and IL-1β), and reduced neuronal apoptosis by reducing the expression of Bax, cytochrome c, cleaved caspase-3, and cleaved PARP-1, while increasing the level of survival proteins p-Akt, p-GSK3β, and anti-apoptotic Bcl-2 protein. Anthocyanins treatment increased the levels of memory-related pre- and post-synaptic proteins and improved the hippocampus-dependent memory in the LPS-treated mice. Overall, this data suggested that consumption of naturally derived anti-oxidant agent such as anthocyanins ameliorated several pathological events in the LPS-treated animal model and we believe that anthocyanins would be a safe therapeutic agent for slowing the inflammation-induced neurodegeneration in the brain against several diseases such as Alzheimer's disease and Parkinson's disease.

N4 -Tetradentate Chelators Efficiently Regulate Copper Homeostasis and Prevent ROS Production Induced by Copper-Amyloid-β1-16.

The disruption of copper homeostasis and the oxidative stress induced by Cu-amyloids are crucial features of Alzheimer's disease pathology. The copper specific N4 -tetradendate ligands TDMQ20 and 1 are able to fully inhibit in vitro the aerobic oxidation of ascorbate induced by Cu-Aβ1-16 , even in the presence of 100 molar equivalents of ZnII with respect to CuII , whereas other ligands with N2 O2 or N3 O2 coordination spheres failed to do so. This essential result indicates that, in addition to metal selectivity, the coordination sphere of copper chelators should exhibit a N4 -tetradendate motif to be able to reduce an oxidative stress in the zinc-rich physiological environment of brain. The N4 -scaffolds of these two aminoquinoline-based ligands, TDMQ20 or 1, suitable for a square-planar coordination of copper(II), allowed them to enhance both the selectivity for copper and the ability to reduce the oxidative stress induced by copper-amyloid in a zinc-rich environment.

Chamazulene prevents ROS production in human dermal fibroblast and bovine aortic endothelial cells exposed to oxidative stress

Chamazulene prevents ROS production in human dermal fibroblast and bovine aortic endothelial cells exposed to oxidative stress

Ascorbic acid as DNase I inhibitor in prevention of male infertility

Apoptotic and/or ROS-induced DNA fragmentation in sperm cells may contribute to the development of male infertility. As the known dietary antioxidant, ascorbic acid prevents ROS production and protects sperm cells from DNA damage. Here, we found that ascorbic acid has the ability to inhibit DNase I, one of the main endonucleases involved in DNA fragmentation during apoptosis. Site Finder and Molecular docking defined the ascorbic acid interactions with the most important residues of DNase I, including H-donor interactions with Asp 168 and Asn 170, and H-acceptor interaction with Asn 170. As a furan derivative, ascorbic acid could be considered a pioneer of substrate-based DNase I inhibitors. The results indicate to another possible mechanism for prevention of male infertility by ascorbic acid.

Melatonin inhibits apoptotic cell death induced by Vibrio vulnificus VvhA via melatonin receptor 2 coupling with NCF-1

Melatonin, an endogenous hormone molecule, has a variety of biological functions, but a functional role of melatonin in the infection of Gram-negative bacterium Vibrio vulnificus has yet to be described. In this study, we investigated the molecular mechanism of melatonin in the apoptosis of human intestinal epithelial (HCT116) cells induced by the hemolysin (VvhA) produced by V. vulnificus. Melatonin (1 μM) significantly inhibited apoptosis induced by the recombinant protein (r) VvhA, which had been inhibited by the knockdown of MT2. The rVvhA recruited caveolin-1, NCF-1, and Rac1 into lipid rafts to facilitate the production of ROS responsible for the phosphorylation of PKC and JNK. Interestingly, melatonin recruited NCF-1 into non-lipid rafts to prevent ROS production via MT2 coupling with Gαq. Melatonin inhibited the JNK-mediated phosphorylation of c-Jun responsible for Bax expression, the release of mitochondrial cytochrome c, and caspase-3/-9 activation during its promotion of rVvhA-induced apoptotic cell death. In addition, melatonin inhibited JNK-mediated phosphorylation of Bcl-2 responsible for the release of Beclin-1 and Atg5 expression during its promotion of rVvhA-induced autophagic cell death. These results demonstrate that melatonin signaling via MT2 triggers recruitment of NCF-1 into non-lipid rafts to block ROS production and JNK-mediated apoptotic and autophagic cell deaths induced by rVvhA in intestinal epithelial cells.

CoQ0-induced mitochondrial PTP opening triggers apoptosis via ROS-mediated VDAC1 upregulation in HL-60 leukemia cells and suppresses tumor growth in athymic nude mice/xenografted nude mice.

Coenzyme Q (CoQ) analogs with variable numbers of isoprenoid units have been demonstrated as anticancer and antioxidant/pro-oxidant molecules. This study examined the in vitro and in vivo antitumor and apoptosis activities of CoQ0 (2,3-dimethoxy-5-methyl-1,4-benzoquinone, zero isoprenoid side-chains) through upregulation of the Voltage-dependent anion channel 1 (VDAC1) signaling pathway on human promyelocytic leukemia. CoQ0 (0-40μg/mL) treatment significantly reduced HL-60 cell viability, and up-regulated mitochondrial VDAC1 expression. CoQ0 treatment triggers intracellular ROS generation, calcium release, ΔΨm collapse and PTP opening in HL-60 cells. CoQ0 treatment induced apoptosis, which was associated with DNA fragmentation, cytochrome c release, caspase-3 and PARP activation, and Bax/Bcl-2 dysregulation. Annexin V-PI staining indicated that CoQ0 promotes late apoptosis. Furthermore, the blockade of CoQ0-induced ROS production by antioxidant NAC pretreatment substantially attenuated CoQ0-induced apoptosis. The activation of p-GSK3β expression, cyclophilin D inhibition, and p53 activation through ROS are involved in CoQ0-induced HL-60 apoptotic cell death. Notably, ROS-independent p38 activation is involved in CoQ0-mediated apoptosis in HL-60 cells. In addition, the silencing of VDAC1 also prevented CoQ0-induced mitochondrial translocation of Bax, activation of caspase-3, and reduction in Bcl-2. Intriguingly, VDAC1 silencing did not prevent ROS production induced by CoQ0, which in turn indicates that CoQ0 induced ROS-mediated VDAC1 and then mitochondrial apoptosis in HL-60 cells. In vivo results revealed that CoQ0 is effective in delaying tumor incidence and reducing the tumor burden in HL-60-xenografted nude mice. Taken together, CoQ0 could be a promising anticancer agent for the treatment of human promyelocytic leukemia through upregulation of VDAC1 signaling pathways.

Debaryomyces Hansenii: Adaptation Mechanisms to Different Carbon Sources and Oxygen Concentrations

Debaryomyces hansenii is an oleaginous, mostly aerobic yeast. Its mitochondrial respiratory chain contains complejxes I, II, III and IV, an alternative oxidase, aln alternative external NADH dehydrogenase an a glycerol-phosphate dehydrogenase (Cabrera-Orefice et al, 2014, Biochim Biophys Acta 1837(1): 73-84; 2014, Biochimie 102: 124-136). Alternative dehydrogenases, seem to physiologically uncouple mitochondria in order to prevent ROS production (Guerrero-Castillo et al, 2011, Biochim Biophys Acta 1817(2): 353-62). In addition, the opening of a unspecific channel releases NAD+, thus inactivating complex I and decreasing the proton-pumping stoichiometry in the system. In regard to the carbon source, lactate or succinate led to the expression of AOX at all growth phases. In contrast, in the presence of glucose or galactose, AOX was only expressed at the late exponential phase. In contrast, in glycerol AOX was never expressed and biomass growth was larger. In addition, large lipidic production was observed. It is suggested that D. hansenii is useful to study cell adaptations to different growth conditions. Also its lipid production mechanisms might be interesting from a biotechnological point of view. The regulation of redox enzyme expression maintains a high respiratory activity even when ATP consumption is low, probably preventing overproduction of ROS.

Regulation of cellular oxidative stress and apoptosis by G protein-coupled receptor kinase-2; The role of NADPH oxidase 4

Cardiac myocyte oxidative stress and apoptosis are considered important mechanisms for the development of heart failure (HF). Chronic HF is characterized by increased circulating catecholamines to augment cardiac output. Long-term stimulation of myocardial β-adrenergic receptors (β-ARs) is deleterious in cardiac myocytes, however, the potential mechanisms underlying increased cell death are unclear. We hypothesize that GRK2, a critical regulator of myocardial β-AR signaling, plays an important role in mediating cellular oxidative stress and apoptotic cell death in response to β-agonist stimulation. Stimulation of H9c2 cells with a non-selective β-agonist, isoproterenol (Iso) caused increased oxidative stress and apoptosis. There was also increased Nox4 expression, but no change in Nox2, the primary NADPH isoforms and major sources of ROS generation in cardiac myocytes. Adenoviral-mediated overexpression of GRK2 led to similar increases in ROS production and apoptosis as seen with Iso stimulation. These increases in oxidative stress were abolished by pre-treatment with the non-specific Nox inhibitor, apocynin, or siRNA knockdown of Nox4. Adenoviral-mediated expression of a GRK2 inhibitor prevented ROS production and apoptosis in response to Iso stimulation. β-Arrestins are signaling proteins that function downstream of GRK2 in β-AR uncoupling. Adenoviral-mediated overexpression of β-arrestins increased ROS production and Nox4 expression. Chronic β-agonist stimulation in mice increased Nox4 expression and apoptosis compared to PBS or AngII treatment. These data demonstrate that GRK2 may play an important role in regulating oxidative stress and apoptosis in cardiac myocytes and provides an additional novel mechanism for the beneficial effects of cardiac-targeted GRK2 inhibition to prevent the development of HF.