Reactive oxygen species impair Na+ transport and renal components of the renin-angiotensin-aldosterone system after paraquat poisoning.

Paraquat (1,1'-dimethyl-4,4'-bipyridyl dichloride) is an herbicide widely used worldwide and officially banned in Brazil in 2020. Kidney lesions frequently occur, leading to acute kidney injury (AKI) due to exacerbated reactive O2 species (ROS) production. However, the consequences of ROS exposure on ionic transport and the regulator local renin-angiotensin-aldosterone system (RAAS) still need to be elucidated at a molecular level. This study evaluated how ROS acutely influences Na+-transporting ATPases and the renal RAAS. Adult male Wistar rats received paraquat (20 mg/kg; ip). After 24 h, we observed body weight loss and elevation of urinary flow and serum creatinine. In the renal cortex, paraquat increased ROS levels, NADPH oxidase and (Na++K+)ATPase activities, angiotensin II-type 1 receptors, tumor necrosis factor-α (TNF-α), and interleukin-6. In the medulla, paraquat increased ROS levels and NADPH oxidase activity but inhibited (Na++K+)ATPase. Paraquat induced opposite effects on the ouabain-resistant Na+-ATPase in the cortex (decrease) and medulla (increase). These alterations, except for increased serum creatinine and renal levels of TNF-α and interleukin-6, were prevented by 4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl (tempol; 1 mmol/L in drinking water), a stable antioxidant. In summary, after paraquat poisoning, ROS production culminated with impaired medullary function, urinary fluid loss, and disruption of Na+-transporting ATPases and angiotensin II signaling.

Curcumin Modulates NOX Gene Expression and ROS Production via P-Smad3C in TGF-ß-Activated Hepatic Stellate Cells.

Background: Liver fibrosis, associated with hepatic stellate cells (HSCs), occurs when a healthy liver sustains damage, thereby impairing its function. NADPH oxidases (NOXs), specifically isoforms 1, 2, and 4, play a role in reactive oxygen species (ROS) production during hepatic injuries, resulting in fibrosis. Curcumin has shown strong potential in mitigating liver fibrosis. Our research aimed to investigate the effects of curcumin on lowering NOX and ROS levels. This compound was also studied for its effects on NOXs, ROS concentrations through the inhibition of Smad3 phosphorylation in transforming growth factor beta (TGF-ß)-activated human HSCs. Methods: MTT assay investigated the cytotoxic effects of curcumin on HSCs. The cells were activated by exposure to TGF-ß (2 ng/mL) for 24 hours. After activating, the cells were treated with curcumin at 25-150 µM concentrations. After administering curcumin to the cells, we employed RT-PCR and Western blot techniques to evaluate the related gene and protein expression levels. This evaluation was primarily focused on the mRNA expression levels of NOX1, NOX2, NOX4 and phosphorylated Smad3C. Results: The mRNA expression level of aforesaid NOXs as well as α-smooth muscle actin (α-SMA), collagen1-α, and ROS levels were significantly reduced following 100 µM curcumin treatment. Furthermore, curcumin significantly decreased the p-Smad3C protein level in TGF-ß-activated cells, with fold changes of 3 and 2 observed at 75 and 100 µM, respectively. Conclusion: Curcumin decreased the levels of ROS and NOX, as well as the expression of α-SMA and collagen1-α. The primary mechanism for this reduction could be linked to the level of p-Smad3C. Hence, curcumin could serve as an effective therapeutic agent for liver fibrosis.

A NAD(P)H oxidase mimic for catalytic tumor therapy via a deacetylase SIRT7-mediated AKT/GSK3ß pathway.

Nicotinamide adenine dinucleotide (NADH) and its phosphorylated form, NADPH, are essential cofactors that play critical roles in cell functions, influencing antioxidation, reductive biosynthesis, and cellular pathways involved in tumor cell apoptosis and tumorigenesis. However, the use of nanomaterials to consume NAD(P)H and thus bring an impact on signaling pathways in cancer treatment remains understudied. In this study, we employed a salt template method to synthesize a carbon-coated-cobalt composite (C@Co) nanozyme, which exhibited excellent NAD(P)H oxidase (NOX)-like activity and mimicked the reaction mechanism of natural NOX. The C@Co nanozyme efficiently consumed NAD(P)H within cancer cells, leading to increased production of reactive oxygen species (ROS) and a reduction in mitochondrial membrane potential. Meanwhile, the generation of the biologically active cofactor NAD(P)+ promoted the expression of the deacetylase SIRT7, which in turn inhibited the serine/threonine kinase AKT signaling pathway, ultimately promoting apoptosis. This work sheds light on the influence of nanozymes with NOX-like activity on cellular signaling pathways in tumor therapy and demonstrates their promising antitumor effects in a tumor xenograft mouse model. These findings contribute to a better understanding of NAD(P)H manipulation in cancer treatment and suggest the potential of nanozymes as a therapeutic strategy for cancer therapy.

Ellagic acid protects against angiotensin II-induced hypertrophic responses through ROS-mediated MAPK pathway in H9c2 cells.

The early myocardial response of hypertension is an elevation of angiotensin-II (Ang-II) concentration, leading to heart failure and cardiac hypertrophy. This hypertrophic event of the heart is mediated by the interaction of Ang type 1 receptors (AT-R1), thereby modulating NADPH oxidase activity in cardiomyocytes, which alters redox status in cardiomyocytes. Ellagic acid (EA) has anti-inflammatory and anti-oxidative capacities. Thus, EA has potential preventive effects on cardiovascular diseases and diabetes. In the last decades, because the protective effect of EA on Ang-II-induced hypertrophic responses is unclear, this study aims to investigate the protective effect of EA in cardiomyocytes. H9c2 cells were treated to Ang-II 1 µM for 24 h to induce cellular damage. We found that EA protected against Ang-II-increased cell surface area and pro-hypertrophic gene expression in H9c2. EA reduced Ang-II-caused AT-R1 upregulation, thereby inhibiting oxidative stress NADPH oxidase activation. EA mitigated Ang-II-enhanced p38 and extracellular-signal-regulated kinase (ERK) phosphorylation. Moreover, EA treatment under Ang-II stimulation also reversed NF-κB activity and iNOS expression. This study shows that EA protects against Ang-II-induced myocardial hypertrophy and attenuates oxidative stress through reactive oxygen species-mediated mitogen-activated protein kinase signaling pathways in H9c2 cells. Thus, EA may be an effective compound for preventing Ang-II-induced myocardial hypertrophy.

Ophthalmic wild olive (ACEBUCHE) oil nanoemulsions exert oculoprotective effects against oxidative stress induced by arterial hypertension.

Oxidative stress plays a key role in several systemic and ocular diseases, including hypertensive eye diseases. In this context, we previously showed that oral administration of wild olive (acebuche, ACE) oil from Olea europaea var. sylvestris can counteract ocular damage secondary to arterial hypertension by modulating excess reactive oxygen species (ROS) produced by the enzyme nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Therefore, this work describes the development of an ACE oil-based formulation for ocular administration as a local therapy to counteract hypertension-related oxidative damage. Specifically, ACE oil nanoemulsions (NEs) were successfully produced and characterized, exhibiting appropriate features for ophthalmic administration, including a nanometer size (<200 nm), moderate negative ZP, adequate osmolality and pH, and colloidal stability in biorelevant fluids. Likewise, the NEs presented a shear thinning behavior, especially convenient for ocular instillation. In vivo evaluation was performed through either intravitreal injection or topical ophthalmic administration in mice with hypertension induced via administration of Nω-nitro-L-arginine-methyl-ester (L-NAME). Both routes of administration reduced hypertensive morphological alterations and demonstrated a noticeable antioxidant effect thanks to the reduction of the activity/expression of NADPH oxidase in cornea and retina. Thus, an ACE oil ophthalmic formulation represent a promising therapy for ocular pathologies associated with arterial hypertension.

ROS produced by NOX promote the neurite growth in a PI3K/Akt independent manner.

Reactive oxygen species (ROS) function as signaling molecules in several physiologic and pathologic processes. In central nervous system, ROS are critical for differentiation, migration, polarization, and neurite growth. These actions are mediated by reversible oxidation of target proteins. On the other hand, PI3K/Akt signaling pathway is susceptible to be modulated by ROS and it has been implicated in neurite growth. In this study, we evaluated the participation of ROS in the neurite growth of cultured rat cerebellar granule neurons (CGN), as well as the possible regulation of the PI3K/Akt pathway by ROS during neurite outgrowth. For this purpose, CGN were treated with cellular or mitochondrial antioxidants, or an NOX inhibitor and neurite growth was evaluated. Moreover, to assess the participation Akt in this process, the p-Akt levels were measured in CGN treated with antioxidants or a NOX inhibitor. The effect of antioxidants on the neurite growth in the presence of a PI3K inhibitor was also measured. We found that cellular antioxidants and the NOX inhibitor decreased the neurite growth, but not the mitochondrial antioxidant. Interestingly, the antioxidants increased the p-Akt levels; however, the effect of antioxidants on neurite growth was no dependent on the Akt activity since the inhibitor of PI3K did not modify the antioxidant action on neurite growth. Our results show that the PI3K/Akt pathway participates in neurite growth and that ROS produced by NOX could function as signals in this process; however, this action is not mediated by a redox regulation of Akt activity.

GM1 ganglioside protects against LPS-induced neuroinflammatory and oxidative responses by inhibiting the activation of Akt, TAK1 and NADPH oxidase in MG6 microglial cells.

GM1 is a major brain ganglioside that exerts neurotrophic, neuroprotective and antineuroinflammatory effects. The aim of this study was to obtain insights into the antineuroinflammatory mechanisms of exogenous GM1 in lipopolysaccharide (LPS)-stimulated MG6 mouse transformed microglial cell line. First, we found that GM1 prevented the LPS-induced transformation of microglia into an amoeboid-like shape. GM1 treatment inhibited LPS-induced expression of inducible nitric oxide synthase, cyclooxygenase-2 (COX-2), and proinflammatory cytokines such as TNF-α, IL-1ß and IL-6 in MG6 cells. In LPS-treated mice, GM1 also reduced striatal microglia activation and attenuated COX-2 expression. Subsequent mechanistic studies showed that GM1 suppressed LPS-induced nuclear translocation of nuclear factor κB (NF-κB) and activator protein-1 (AP-1), two critical transcription factors responsible for the production of proinflammatory mediators. GM1 exhibited antineuroinflammatory properties by suppressing Akt/NF-κB signaling and the activation of mitogen-activated protein kinases (MAPKs), including p38 MAPK, extracellular signal-regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK). Furthermore, GM1 suppressed LPS-induced activation of transforming growth factor-ß-activated kinase 1 (TAK1) and NADPH oxidase 2 (NOX2), upstream regulators of the IκBα/NF-κB and MAPK/AP-1 signaling pathways. GM1 also inhibited NOX-mediated reactive oxygen species (ROS) production and protected against LPS-induced MG6 cell death, suggesting an antioxidant role of GM1. In conclusion, GM1 exerts both antineuroinflammatory and antioxidative effects by inhibiting Akt, TAK1 and NOX2 activation.

SARS-CoV-2 primed platelets-derived microRNAs enhance NETs formation by extracellular vesicle transmission and TLR7/8 activation.

BACKGROUND: Hyperactive neutrophil extracellular traps (NETs) formation plays a key role in the pathogenesis of severe COVID-19. Extracellular vesicles (EVs) are vehicles which carry cellular components for intercellular communication. The association between COVID-19 patients-derived EVs and NETs formation remains elusive. METHODS: We explored the roles of EVs in NETs formation from 40 COVID-19 patients with different disease severities as well as 30 healthy subjects. The EVs-carried microRNAs profile was analyzed using next generation sequencing approach which was validated by quantitative reverse transcription PCR. The regulatory mechanism of EVs on NETs formation was investigated by using an in vitro cell-based assay, including immunofluorescence assay, flow cytometry, and immunoblotting. RESULTS: COVID-19 patient-derived EVs induced NETs formation by endocytosis uptake. SARS-CoV-2 spike protein-triggered NETs formation was significantly enhanced in the presence of platelet-derived EVs (pEVs) and this effect was Toll-like receptor (TLR) 7/8- and NADPH oxidase-dependent. Increased levels of miR-21/let-7b were revealed in EVs from COVID-19 patients and were associated with disease severity. We demonstrated that the spike protein activated platelets directly, followed by the subsequent intracellular miR-21/let-7b upregulation and then were loaded into pEVs. The pEVs-carried miR-21 interacted with TLR7/8 to prime p47phox phosphorylation in neutrophils, resulting in NADPH oxidase activation to promote ROS production and NETs enhancement. In addition, miR-21 modulates NF-κB activation and IL-1ß/TNFα/IL-8 upregulation in neutrophils upon TLR7/8 engagement. The miR-21 inhibitor and TLR8 antagonist could suppress efficiently spike protein-induced NETs formation and pEVs primed NETs enhancement. CONCLUSIONS: We identified SARS-CoV-2 triggered platelets-derived GU-enriched miRNAs (e.g., miR-21/let-7b) as a TLR7/8 ligand that could activate neutrophils through EVs transmission. The miR-21-TLR8 axis could be used as a potential predisposing factor or therapeutic target for severe COVID-19.

Neuronal NADPH oxidase is required for neurite regeneration of Aplysia bag cell neurons.

NADPH oxidase (Nox), a major source of reactive oxygen species (ROS), is involved in neurodegeneration after injury and disease. Nox is expressed in both neuronal and non-neuronal cells and contributes to an elevated ROS level after injury. Contrary to the well-known damaging effect of Nox-derived ROS in neurodegeneration, recently a physiological role of Nox in nervous system development including neurogenesis, neuronal polarity, and axonal growth has been revealed. Here, we tested a role for neuronal Nox in neurite regeneration following mechanical transection in cultured Aplysia bag cell neurons. Using a novel hydrogen peroxide (H2 O2 )-sensing dye, 5'-(p-borophenyl)-2'-pyridylthiazole pinacol ester (BPPT), we found that H2 O2 levels are elevated in regenerating growth cones following injury. Redistribution of Nox2 and p40phox in the growth cone central domain suggests Nox2 activation after injury. Inhibiting Nox with the pan-Nox inhibitor celastrol reduced neurite regeneration rate. Pharmacological inhibition of Nox is correlated with reduced activation of Src2 tyrosine kinase and F-actin content in the growth cone. Taken together, these findings suggest that Nox-derived ROS regulate neurite regeneration following injury through Src2-mediated regulation of actin organization in Aplysia growth cones.

MARCH9 Mediates NOX2 Ubiquitination to Alleviate NLRP3 Inflammasome-Dependent Pancreatic Cell Pyroptosis in Acute Pancreatitis.

OBJECTIVE: The pathogenesis of acute pancreatitis mainly involves NLRP3 inflammasome-mediated pancreatic cell injury, although regulators of this inflammasome machinery are still not fully identified. Membrane-associated RING-CH 9 (MARCH9) is a member of MARCH-type finger proteins, which regulates innate immunity through catalyzing polyubiquitination of critical immune factors. The aim of present research is to examine the function of MARCH9 in acute pancreatitis. METHODS: Cerulein-induced acute pancreatitis was established on pancreatic cell line AR42J and rat model. Reactive oxygen species (ROS) accumulation and NLRP3 inflammasome-dependent cell pyroptosis in pancreas were examined by flow cytometry. RESULTS: MARCH9 was downregulated by cerulein, but overexpressing MARCH9 could inhibit NLRP3 inflammasome activation and ROS accumulation, thus suppressing pancreatic cell pyroptosis and mitigating pancreatic injury. We further uncovered that the mechanism underlying such an effect of MARCH9 is through mediating the ubiquitination of NADPH oxidase-2, whose deficiency reduces cellular ROS accumulation and inflammasome formation. CONCLUSIONS: Our results suggested that MARCH9 suppresses NLRP3 inflammasome-mediated pancreatic cell injury through mediating the ubiquitination and degradation of NADPH oxidase-2, which compromises ROS generation and NLRP3 inflammasomal activation.

Dexamethasone Induces Senescence-Associated Changes in Trabecular Meshwork Cells by Increasing ROS Levels Via the TGFß/Smad3-NOX4 Axis.

Glaucoma is a serious complication of glucocorticoid (GC) therapy arising through elevations in intraocular pressure (IOP). Dexamethasone (DEX) is reported to contribute to elevated IOP through different effects on the trabecular meshwork but whether DEX contributes to glaucoma development through the induction of cellular senescence is still unclear. We explored the actions of DEX on transformed human trabecular meshwork cells (HTMCs) using RNA-seq and conducted bioinformatic analyses to determine the affected pathways. Among the 4,103 differentially expressed genes identified in transformed HTMCs treated with 400 nM DEX (2,036 upregulated and 2,067 downregulated genes, respectively), bioinformatic analyses revealed significant enrichment and potential interplay between the transforming growth factor beta (TGFß)41; signaling and cellular senescence pathways. DEX treatment induced senescence changes in primary and transformed HTMCs as indicated by increases in SA-ß-gal positivity, interleukin (IL)-6 secretion, and senescence-associated heterochromatin foci (SAHF) along with selective accumulation of senescence marker p15 and elevations in reactive oxygen species (ROS) levels. Notably, the DEX-induced senescence changes were rescued by treatment with the TGFß/Smad3 pathway inhibitor SIS3. Furthermore, we show that DEX increases cellular ROS levels via upregulation of NADPH oxidase 4 (NOX4) through activation of Smad3, and that SIS3 decreases ROS levels by downregulating NOX4. Instructively, inhibiting NOX4 with GLX351322 and scavenging ROS with NAC were both effective in preventing DEX-induced senescence changes. Similarly, we found in the mouse model that DEX-ac upregulated p15 and NOX4 expression in the trabecular meshwork, with cotreatment with GLX351322 alleviating elevations in IOP. We establish that DEX induces senescence changes in HTMCs by increasing ROS levels via the TGFß/Smad3/NOX4 axis, increasing IOP and contributing to glaucoma development.

All-trans retinoic acid inhibits oxidative stress via ACE2/Ang (1-7)/MasR pathway in renal tubular epithelial cells stimulated with high glucose.

The aim of this study was to investigate the effects of all-trans retinoic acid (atRA) on oxidative stress in renal tubular epithelial cells induced by high glucose (HG) and its potential mechanism. We investigated the effects of atRA in HG-induced renal epithelial cell line HK-2. Seven groups were designed for this experiment: negative control, mannitol, high-glucose (HG), HG combined with a low concentration of atRA, HG combined with a middle concentration of atRA, HG combined with a high concentration of atRA, and HG combined with captopril. After 48 h of incubation, oxidative stress factor expression in the supernatant was detected by enzyme-linked immunosorbent assay. Reactive oxygen species and cell apoptosis expression were assessed by flow cytometry. NADPH oxidase, fibrosis factor, and angiotensin-converting enzyme 2/angiotensin (1-7)/mas receptor (ACE2/Ang (1-7)/MasR) pathway-related protein expressions were determined by western blot analysis. The expressions of oxidative stress factors, NADPH oxidase components, and fibrosis factors were significantly higher after HG treatment. Apoptosis of HK2 cells in the HG group was also significantly higher. AtRA could reverse the above abnormal changes in a concentration-dependent manner. HG significantly promoted the expression of ACE, Ang II, and Ang II type 1 receptor (AT1R), whereas it inhibited the expression of ACE2, Ang (1-7), and MasR. With the elevation of concentration, atRA could gradually suppress the expression of ACE, Ang II, and AT1R, but facilitate ACE2, Ang (1-7), and MasR. These results were statistically significant. AtRA could significantly inhibit oxidative stress and apoptosis of renal tubular epithelial cells induced by HG. The mechanism may inhibit the ACE/Ang II/AT1R pathway and/or activate ACE2/Ang (1-7)/MasR pathway.

Topically Administered NOX4 Inhibitor, GLX7013114, Is Efficacious in Treating the Early Pathological Events of Diabetic Retinopathy.

NADPH oxidases (NOXs) are major players in generating reactive oxygen species (ROS) and are implicated in various neurodegenerative ocular pathologies. The aim of this study was to investigate the role of a NOX4 inhibitor (GLX7013114) in two in vivo, experimental streptozotocin (STZ) paradigms depicting the early events of diabetic retinopathy (DR). Animals in the diabetic treated group received GLX7013114 topically (20 µL/eye, 10 mg/mL, once daily) for 14 days (paradigm A: preventive) and 7 days (paradigm B: treated) at 48 h and 4 weeks after STZ injection, respectively. Several methodologies were used (immunohistochemistry, Western blot, real-time PCR, ELISA, pattern electroretinography [PERG]) to assess the diabetes-induced early events of DR, namely oxidative stress, neurodegeneration, and neuroinflammation, and the effect of GLX7013114 on the diabetic insults. GLX7013114, administered as eye drops (paradigms A and B), was beneficial in treating the oxidative nitrative stress, activation of caspase-3 and micro- and macroglia, and attenuation of neuronal markers. It also attenuated the diabetes-induced increase in vascular endothelial growth factor, Evans blue dye leakage, and proinflammatory cytokine (TNF-α protein, IL-1ß/IL-6 mRNA) levels. PERG amplitude values suggested that GLX7013114 protected retinal ganglion cell function (paradigm B). This study provides new findings regarding the pharmacological profile of the novel NOX4 inhibitor GLX7013114 as a promising therapeutic candidate for the treatment of the early stage of DR. ARTICLE HIGHLIGHTS: NADPH oxidases (NOXs) are implicated in the early pathological events of diabetic retinopathy (DR). The NOX4 inhibitor GLX7013114, topically administered, reduced oxidative damage and apoptosis in the rat streptozotocin model of DR. GLX7013114 protected retinal neurons and retinal ganglion cell function and reduced the expression of pro-inflammatory cytokines in the diabetic retina. GLX7013114 diminished the diabetes-induced increase in vascular endothelial growth factor levels and Evans blue dye leakage in retinal tissue. GLX7013114 exhibits neuroprotective, anti-inflammatory, and vasculoprotective properties that suggest it may have a role as a putative therapeutic for the early events of DR.

Blueberry treatment administered before and/or after lipopolysaccharide stimulation attenuates inflammation and oxidative stress in rat microglial cells.

ABSTRACTMicroglia are key regulators of inflammation and oxidative stress (OS) in the CNS. Microglia activation can lead to chronic inflammation, OS, and neurodegeneration. Blueberries (BB) reduce inflammation and OS when administered to microglia before stressors such as lipopolysaccharide (LPS), but the therapeutic value of BBs administered after activation by stressors has not been examined. Therefore, this study investigated the differential effects of pre-, post-, and pre-/post-BB on inflammation and OS in LPS-activated microglia. Rat microglia were pretreated with BB (0.5 mg/mL) or control media (C) for 24 hours, incubated overnight with LPS (0 or 200 ng/mL), and post-treated with BB or C for 24 hours. Biomarkers of inflammation (e.g. nitrite [NO2-], tumor necrosis factor-ɑ [TNFɑ], inducible nitric oxide synthase [iNOS], cyclooxygenase-2 [COX-2], phosphorylated IκB-α [pIκB-ɑ]) and OS (e.g. NADPH oxidase [NOX2]) were assessed. LPS increased NO2-, TNFɑ, COX-2, iNOS, pIκB-ɑ, and NOX2 compared to non-stressed conditions (P < 0.05), however BB before and/or after LPS significantly reduced these markers compared to no BB (P < 0.05). Pre-BB was more effective than post-BB at reducing LPS-induced NO2-, TNFɑ, and COX-2 (P < 0.05). Pre-BB was also more effective than pre-/post-BB at attenuating LPS-induced NO2- and TNFɑ (P < 0.05). All BB treatments were equally effective in reducing LPS-induced iNOS, pIκB-ɑ, and NOX2. Results suggest that BBs can target the downstream events of LPS-induced microglial activation and prevent stressor-induced neuroinflammation and OS. Furthermore, BBs may not need to be present prior to microglial activation for beneficial effects, suggesting that dietary interventions may be effective even after initiation of disease processes.Graphical Abstract. Cascade of inflammatory and OS-inducing events associated with self-propelling microglial activation by LPS and the effects of blueberry (0.5 mg/mL) administered before and/or after LPS on these processes (blue arrows). BB, blueberry; COX2, cyclooxygenase-2; IκB-ɑ, inhibitor kappa-B-ɑ; iNOS, inducible nitric oxide synthase; LPS, lipopolysaccharide; NF-κB, nuclear factor kappa-B; NO, nitric oxide; NOX2, NADPH oxidase; OS, oxidative stress; ROS, reactive oxygen species; TNFɑ, tumor necrosis factor-ɑ.

Surfactin induces autophagy, apoptosis, and cell cycle arrest in human oral squamous cell carcinoma.

OBJECTIVES: To investigate the anticancer effects and underlying mechanisms of surfactin on human oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: The capacity of surfactin to induce apoptosis, autophagy, and cell cycle arrest of two different human OSCC cell lines was investigated by cell viability, acridine orange staining, and cell cycle regulatory protein expression, respectively. The signaling network underlying these processes were determined by the analysis of reactive oxygen species (ROS) generation, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity, endoplasmic reticulum (ER) stress-related protein levels, calcium release, mitogen-activated protein kinases activation, and cell cycle regulatory protein expression through corresponding reagents and experiments under various experimental conditions using specific pharmaceutical inhibitors or small interfering RNAs. RESULTS: Surfactin was able to induce apoptosis through NADPH oxidase/ROS/ER stress/calcium-downregulated extracellular signal-regulated kinases 1/2 pathway. Surfactin could also lead to autophagy that shared the common regulatory signals with apoptosis pathway until calcium node. Cell cycle arrest at G2 /M phase caused by surfactin was demonstrated through p53 and p21 accumulation combined p34cdc2 , phosphorylated p34cdc2 , and cyclin B1 inhibition, which was regulated by NADPH oxidase-derived ROS. CONCLUSION: Surfactin could induce apoptosis, autophagy, and cell cycle arrest in ROS-dependent manner, suggesting a multifaced anticancer agent for OSCC.

Thymoquinone ameliorates amikacin induced oxidative damage in rat brain tissue.

We investigated the potential neuroprotective effects of thymoquinone (TQ) on amikacin (AK) induced oxidative damage in rat brain. We used 21 male rats divided randomly into three equal groups. The control group was injected intraperitoneally (i.p.) with 0.5 ml 0.9% aqueous NaCl and given 1 ml 0.9% aqueous NaCl orally. The AK group was administered 1.2 g/kg aqueous AK i.p. as a single dose on the day 3 of the study. The AK + TQ group was given a single 1.2 g/kg dose of AK i.p. on the day 3 of the study plus 40 mg/kg/day TQ by oral gavage daily. Treatment with TQ increased serum ferritin and decreased serum calcium levels significantly. TQ also decreased NADPH oxidase-2, NADPH oxidase-4, and caspase-3 levels. Decreased malondialdehyde (MDA) levels and increased superoxide dismutase (SOD) and catalase (CAT) activities were detected in the AK + TQ group compared to the AK group. TQ administration inhibited lipid peroxide formation and blocked oxidative reactions, which reduced the MDA level and increased SOD and CAT activities induced by AK. Oxidative damage caused by AK was ameliorated by TQ treatment owing to its antioxidative and anti-apoptotic effects. TQ may be a potential therapeutic agent for reducing the severity of AK induced oxidative damage to the brain.

Superoxide Release by Macrophages through NADPH Oxidase Activation Dominating Chemistry by Isoprene Secondary Organic Aerosols and Quinones to Cause Oxidative Damage on Membranes.

Oxidative stress mediated by reactive oxygen species (ROS) is a key process for adverse aerosol health effects. Secondary organic aerosols (SOA) account for a major fraction of fine particulate matter, and their inhalation and deposition into the respiratory tract causes the formation of ROS by chemical and cellular processes, but their relative contributions are hardly quantified and their link to oxidative stress remains uncertain. Here, we quantified cellular and chemical superoxide generation by 9,10-phenanthrenequinone (PQN) and isoprene SOA using a chemiluminescence assay combined with electron paramagnetic resonance spectroscopy as well as kinetic modeling. We also applied cellular imaging techniques to study the cellular mechanism of superoxide release and oxidative damage on cell membranes. We show that PQN and isoprene SOA activate NADPH oxidase in macrophages to release massive amounts of superoxide, overwhelming the superoxide formation by aqueous chemical reactions in the epithelial lining fluid. The activation dose for PQN is 2 orders of magnitude lower than that of isoprene SOA, suggesting that quinones are more toxic. While higher exposures trigger cellular antioxidant response elements, the released ROS induce oxidative damage to the cell membrane through lipid peroxidation. Such mechanistic and quantitative understandings provide a basis for further elucidation of adverse health effects and oxidative stress by fine particulate matter.

Bisphenol A: Potential Factor of Miscarriage in Women in the Context of the Phenomenon of Neutrophil Extracellular Traps.

Humans are exposed to a number of environmental pollutants every day. Among them, endocrine disruptors are particularly harmful to human health. Bisphenol A (BPA) is a xenoestrogen that has been shown to disrupt the endocrine system and cause reproductive toxicity. In this study, we aimed to verify the potential relationship between BPA and miscarriage involving the formation of neutrophil extracellular traps (NETs). Blood samples were collected from healthy women and women who had miscarriage in the first trimester of pregnancy. The serum levels of cytoplasmic anti-PR3 antibody and perinuclear anti-MPO antibody were determined using an immunoenzymatic method. The concentrations of key proinflammatory proteins TNF-α and MCP-1, as well as NADPH oxidase subunits NOX1 and NCF2, were also measured in the serum samples. The serum concentration of BPA was determined using gas chromatography. The results showed that the concentrations of BPA were significantly elevated in the serum of women who had miscarriage compared to the control group, with the highest concentration found in the "NETs-positive" group. The levels of MCP-1 and TNF-α were significantly higher in the "NETs-positive" group compared to the "NETs-negative" and control group. The levels of NOX1 and NCF2 were also higher in the "NETs-positive" group compared to the "NETs-negative" group. The study showed that BPA could play a role in the course of miscarriage through the formation of NETs. The results indicate the need to limit the exposure of women planning pregnancy to xenoestrogens, including BPA.

High-salt diet contributes to excess oxidative stress and abnormal metabolism in mouse ovaries.

High-salt diets (HSDs) are associated with elevated levels of reactive oxygen species (ROS), which play a key role in ovarian disorders. However, it is not yet clear whether HSDs impact ovarian redox balance and metabolism. Accordingly, in this study, we analyzed the effect of HSDs on ovarian redox balance by biochemical analysis and further dissected its possible mechanism by metabolic analysis combined with the correlation network method. We found that ROS and H2 O2 levels significantly increased in the ovarian tissue of mice receiving an HSD for 4 weeks. The enhanced activity of NADPH oxidase may contribute to an increase in ROS in ovarian tissue after excessive salt consumption. Meanwhile, the activities of key antioxidant enzymes, including superoxide dismutase 2, glutathione peroxidase, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase, increased significantly. The apparent activation of antioxidant defense appeared insufficient as the glutathione, GSH/GSSG ratio, and NADPH/NADP+ ratio decreased significantly. In addition, HSDs significantly altered the metabolic patterns of ovarian tissue in mice, and pathways were mainly enriched in fatty acid metabolism. Arachidonic acid was an altered hub metabolite according to Pearson correlation network analysis. Mechanistically, an HSD increased the concentration of arachidonic acid in ovarian tissue, inducing high NADPH oxidase activity, which increased the levels of ROS and H2 O2 . Our results indicate that HSDs can lead to increased oxidative stress and dramatically alter the metabolic patterns in mouse ovarian tissues.

Prenatal hypothyroidism diminished exogenous NO-mediated diastolic effects in fetal rat thoracic aorta smooth muscle via increased oxidative stress.

Maternal hypothyroidism is an important problem of modern healthcare and is reported to increase the risk of cardiovascular diseases in the offspring later in life. However, it is unclear whether hypothyroidism during pregnancy causes vascular damage in the fetal period. We established the prenatal hypothyroidism rat model and collected the fetuses at the 21th day of gestation (GD21). Thyroid hormone concentrations in maternal and offspring blood serum were assessed by enzyme-linked immunosorbent assay (ELISA). The thoracic aortas of the fetuses were isolated for microvessel functional testing and histochemical stainings. qPCR and Western blot were performed to access mRNA and protein expression. We found that the concentrations of thyroid hormones in the serum of pregnant rats and fetuses were significantly suppressed at GD21. The responses of the fetal thoracic aortas to SNP were significantly attenuated in the PTU group. However, no statistical difference was found between the two groups when treated with either inhibitor (ODQ) or activator (BAY58-2667) of sGC. The production of O2-• in the arterial wall was significantly increased in hypothyroid fetuses. Moreover, the level of NADPH oxidase (NOX) was increased, while superoxide dismutase 2 (SOD2) was down-regulated in the PTU group, ultimately contributing to the increased production of superoxide. Additionally, decreased SNP-mediated vasodilation found in fetal vessels was improved by either NOX inhibitor (Apocynin) or SOD mimic (Tempol). These results indicate that increased oxidative stress is probably the cause of the diminished diastolic effect of exogenous NO in the thoracic artery of prenatal hypothyroidism exposed fetuses.