Catalase Protein Expression Research Articles

The Protective Effect of Trichosanthes kirilowii Peel Polysaccharide on the Oxidative Damaged HepG2 and HUASMC Cells.

Background Oxidative stress is an important cause of liver disease and atherosclerosis. Natural substances with antioxidant activity are good drugs for treating liver disease and atherosclerosis. Trichosanthes kirilowii Peel Polysaccharide (TKPP) can remove DPPH (2,2-Diphenyl-1-picrylhydrazyl) free radicals and hydroxyl free radicals in vitro, which shows antioxidant activity. Therefore, it is speculated that it can protect human hepatoma cell line (HepG2) and umbilical artery smooth muscle cell (HUASMC) against oxidative damage by hydrogen peroxide (H2O2). Methods Oxidative damage cell models of HepG2 and HUASMC were induced by H2O2. HepG2 and HUASMC were divided into blank group, H2O2 injury group, TKPP treatment group, and glutathione (GSH) positive control group. Cell Counting Kit-8 (CCK-8) was used to detect cell viability. The level of total GSH and the amount of Nitric oxide (NO) secreted by cells were detected by specific kits. The gene and protein expressions of catalase (CAT) and superoxide dismutase (SOD) were detected by fluorescence quantitative PCR and Western Blot. Results In these two kinds of cells, compared with the control group, the survival rate, total GSH level, and NO secretion, CAT and SOD gene and protein expressions were significantly decreased in the H2O2 damaged group. In the TKPP treatment group, the cell survival rate was significantly elevated with the increase of the polysaccharide concentration, and the total GSH level, NO secretion, CAT and SOD gene expression, and protein expression levels were also significantly increased. Conclusion TKPP can improve the activities of HepG2 and HUASMC cells damaged by H2O2 and protect the cellular antioxidant system.

Cessation Restores Blood Pressure Levels and Endothelial Function Affected by Cadmium Exposure on Rats.

Chronic cadmium exposure produces high blood pressure and endothelial damage; however, it is not known whether these effects could be reversed by interrupting the exposure to the metal. Therefore, we evaluate the systolic blood pressure (SBP) and vascular reactivity during and following chronic cadmium-exposure discontinuance. Rats received 100mg.L-1 cadmium chloride (CdCl2) in the drinking water or tap water (Ct) for 30days and/or tap water for 30days more. The cadmium plasma content, blood pressure and vascular reactivity of isolated aorta were evaluated. Cadmium exposure increased cadmium plasma content, SBP and aorta contractile responses to phenylephrine, all reversed after suspending exposure. Endothelial removal and nitric oxide synthase (NOS) inhibition increased phenylephrine response both on control and Cd-discontinuation models. Cd-discontinuation group presented increased CAMKII and PKA protein expression, as peNOSSer1177. Superoxide dismutase (SOD) incubation reduced contractile response on control group, and catalase incubation enhanced the response to phenylephrine in this group. Meanwhile, both SOD2 and catalase protein expression were increased in Cd-cessation rats. Our findings provide evidence that increased SBP and endothelial dysfunction induced by Cd chronic exposure are reversed by suspending the metal exposure probably due to an improvement of antioxidant enzymes and eNOS function.

Effect of Acrylamide Treatment on Cyp2e1 Expression and Redox Status in Rat Hepatocytes.

Acrylamide (AA) toxicity is associated with oxidative stress. During detoxification, AA is either coupled to gluthatione or biotransformed to glycidamide by the enzyme cytochrome P450 2E1 (CYP2E1). The aim of our study was to examine the hepatotoxicity of AA in vivo and in vitro. Thirty male Wistar rats were treated with 25 or 50 mg/kg b.w. of AA for 3 weeks. Qualitative and quantitative immunohistochemical evaluation of inducible nitric oxide synthase (iNOS), CYP2E1, catalase (CAT), superoxide dismutase 1 (SOD1), and SOD2 expression in liver was carried out. Bearing in mind that the liver is consisted mainly of hepatocytes, in a parallel study, we used the rat hepatoma cell line H4IIE to investigate the effects of AA at IC20 and IC50 concentrations on the redox status and the activity of CAT, SOD, and glutathione-S-transferase (GST), their gene expression, and CYP2E1 and iNOS expression. Immunohistochemically stained liver sections showed that treatment with AA25mg induced a significant decrease of CYP2E1 protein expression (p < 0.05), while treatment with AA50mg led to a significant increase of iNOS protein expression (p < 0.05). AA treatment dose-dependently elevated SOD2 protein expression (p < 0.05), while SOD1 protein expression was significantly increased only at AA50mg (p < 0.05). CAT protein expression was not significantly affected by AA treatments (p > 0.05). In AA-treated H4IIE cells, a concentration-dependent significant increase in lipid peroxidation and nitrite levels was observed (p < 0.05), while GSH content and SOD activity significantly decreased in a concentration-dependent manner (p < 0.05). AA IC50 significantly enhanced GST activity (p < 0.05). The level of mRNA significantly increased in a concentration-dependent manner for iNOS, SOD2, and CAT in AA-treated H4IIE cells (p < 0.05). AA IC50 significantly increased the transcription of SOD1, GSTA2, and GSTP1 genes (p < 0.05), while AA IC20 significantly decreased mRNA for CYP2E1 in H4IIE cells (p < 0.05). Obtained results indicate that AA treatments, both in vivo and in vitro, change hepatocytes; drug-metabolizing potential and disturb its redox status.

Chronic treatment with juçara (Euterpe edulis) fruit pulp produces antihypertensive effect and improve on baroreflex sensitivity in Spontaneous Hypertensive Rats (SHR)

Background: Juçara açaí is a Brazilian berry with high anthocyanidin content and high antioxidant properties. A correlation between oxidative stress, hypertension and baroreflex impairment is known, however, no previous studies investigate the effect of juçara fruit pulp on hypertension. Aim: The aim of this study was to investigate antihypertensive effect of juçara (Euterpe edulis) fruit pulp in SHR animals, its antioxidant capacity and influence on baroreflex sensitivity. Methods: Chemical profile was obtained by ESI(-)-FT-ICR-MS. For biological analysis, male SHR were separated into groups: control (S), which received water; captopril (SC), which received captopril (30 mg/kg), and juçara (SJ), which received juçara pulp (4.4mL/kg). Treatments were daily performed by gavage and remained for 8 weeks. Baroreflex evaluation was made by in bolus injection of phenylephrine and sodium nitroprusside, after the analysis the heart was frozen (-80ºC) for biomolecular evaluations (AOPP, TBARS, SOD and catalase activity and protein expression). Results: The treatments promoted decrease in arterial pressure (S: 163.3±2.2; SC: 154.0±0.6; SJ: 149.0±3.8 mmHg) and improved the baroreflex sensitivity after nitroprusside activation. In addition, it promoted a decrease in AOPP and an increase in the activity of antioxidant enzymes. However, no changes were observed in protein expression. Conclusion: It was shown that juçara can promote cardiovascular benefits and its consume should be stimulated for prevention of hypertension.

Salidroside Exerts Beneficial Effect on Testicular Ischemia-Reperfusion Injury in Rats

Testicular torsion-detorsion results in testicular ischemia-reperfusion injury, which is associated with overgeneration of reactive oxygen species. Salidroside, a major bioactive ingredient extracted from Rhodiola rosea, has strong antioxidant activity. The purpose of this study was to examine the effect of salidroside on testicular ischemia-reperfusion injury. Sixty rats were randomly separated into 3 experimental groups: group A = sham-operated control; group B = testicular ischemia-reperfusion; and group C = testicular ischemia-reperfusion treated with salidroside. The rats in the sham-operated control group received all surgical procedures except testicular torsion-detorsion. The testicular ischemia-reperfusion group underwent 2 hours of left testicular torsion followed by detorsion. The rats in the salidroside-treated group received the same surgical procedure as in testicular ischemia-reperfusion group, but salidroside was injected intraperitoneally at reperfusion. Testicular malondialdehyde content (a reliable index of reactive oxygen species) and protein expression of superoxide dismutase and catalase which are primary antioxidant enzymes in testes were measured at 4 hours after reperfusion. Testicular spermatogenesis was evaluated at 3 months after reperfusion. The malondialdehyde content increased significantly, while superoxide dismutase and catalase protein expression and testicular spermatogenesis reduced significantly in ipsilateral testes of testicular ischemia-reperfusion group, as compared with sham-operated control group. Therapy with salidroside significantly reduced malondialdehyde content and significantly enhanced superoxide dismutase and catalase protein expression and spermatogenesis in ipsilateral testes, as compared with testicular ischemia-reperfusion group. The present findings indicate that treatment with salidroside ameliorates testicular ischemia-reperfusion injury by reducing reactive oxygen species level by upregulating superoxide dismutase and catalase protein expression.

The Role of Polyphenols on Blue Light‐Induced Retinal Pigment Cell Damage

The increased exposure of eyes to blue light is a growing concern in our everyday lives. Blue light is responsible for cell damage, cell death, and oxidative stress all of which can lead to vision loss. Resveratrol and pterostilbene are polyphenols found in the skin of various fruits including grapes and blueberries and have been known for antioxidant properties. The aim of this study was to investigate the protective role of resveratrol and pterostilbene on cell death, cell proliferation, and oxidative stress in Human Retinal Pigment Epithelial (ARPE‐19) cells exposed to blue light. ARPE‐19 cells were pre‐incubated with either; 50 or 100uM of resveratrol or 10 or 50uM of pterostilbene for 4 hours followed by exposure to blue light (475nm) for 12 hours. Subsequent to the blue light exposure, cellular viability, oxidative stress, and death were measured in each treatment condition. Blue light exposure resulted in a decrease in cell viability, an increase in caspase 3/7 activation, and cell death. This was accompanied by an increase in protein expression of catalase, and manganese superoxide dismutase (MnSOD) and an increase in 4‐Hydroxynoneal (4HNE) protein adduct formation. In conclusion, resveratrol mitigated the damaging effects of blue light on ARPE‐19 cells by attenuating cell death and oxidative stress. This in‐vitro study is the first step into understanding how polyphenols can possibly be used to reduce the oxidative damage caused by blue light in the eyes and prevent unnecessary vision loss.

FOXO3a-ROS pathway is involved in androgen-induced proliferation of prostate cancer cell

BackgroundAlthough FOXO3a can inhibit the cell proliferation of prostate cancer, its relationship with reactive oxygen species (ROS) in prostate cancer (PCa) has not been reported.MethodsWe analyzed the correlation between the expression of FOXO3a and the antioxidant enzyme catalase in prostate cancer with the TCGA and GEPIA databases. We also constructed a PPI network of FOXO3a via the STRING database. The mRNA and protein expression of FOXO3a and catalase were detected by qRT-PCR or western blotting in LNCaP and 22RV1 cells treated with DHT, R1881, or Enzalutamide. The effects of FOXO3a on catalase expression were tested by over-expressing or knocking down FOXO3a in LNCaP cells. Furthermore, the catalase activity and ROS level were detected in LNCaP cells treated with DHT. Cell proliferation and ROS were also analyzed in LNCaP which was treated with antioxidant.ResultsResults showed that the catalase expression was down-regulated in prostate cancer. A positive correlation between FOXO3a and catalase existed. DHT treatment could significantly reduce FOXO3a and catalase expression at mRNA and protein level in LNCaP cells. Catalase expression partly depended on FOXO3a as over-expression and knockdown of FOXO3a could result in the expresssion change of catalase. DHT treatment was found to inhibit catalase activity and increase ROS level in prostate cancer cell. Our study also demonstrated that antioxidant treatment reduced DHT-induced proliferation and ROS production in prostate cancer cell.ConclusionsWe discovered a novel mechanism by which DHT promotes prostate cancer cell proliferation via suppressing catalase activity and activating ROS signaling via a FOXO3a dependent manner.

PSV-17 The Interaction Between Heat Stress and Zearalenone Toxicity in Porcine Skeletal Muscle

Abstract Heat stress (HS) and Zearalenone (Zen) exposure negatively impact growth, production efficiency, animal welfare, and both can be lethal. Zearalenone, a mycotoxin, can contaminate a variety of feedstuffs. As both HS and Zen independently can cause oxidative stress, we hypothesized that concomitant exposure to HS and Zen would cause oxidative stress in pig skeletal muscle more than either agent alone. Prepubertal gilts (n = 26) were fed placebo or Zen (40 ug/kg) twice daily and exposed to thermoneutral conditions (TN; 21.0 °C) or a 12 h diurnal HS challenge (simulated night - 32.2 °C; simulated day - 35.0 °C) for 7 d. Immediately following the environmental challenge, pigs were euthanized and the oxidative (STR) portion of the semitendinosus muscle was collected. Heat stress increased rectal temperature following the simulated nighttime (TN: 38.9 ± 0.04 °C; HS: 39.3 ± 0.04; P &amp;lt; 0.0001) and daytime (TN: 39.2 ± 0.09 °C; HS: 39.9 ± 0.08 °C; P &amp;lt; 0.0001) conditions. In STR, malondialdehyde-modified protein abundance (P = 0.1) and concentration (P &amp;lt; 0.1), markers of oxidative stress, tended to increase following Zen exposure, largely driven by the HS-Zen group. Heat stress increased protein expression of antioxidant enzymes catalase (p&amp;lt; 0.05) and superoxide dismutase 1 (P &amp;lt; 0.05), while Zen decreased glutathione peroxidase 1 (P &amp;lt; 0.05) transcript (P &amp;lt; 0.05) and protein (P &amp;lt; 0.05) abundance and enzymatic activity (P &amp;lt; 0.05). Although a mild HS paradigm and low Zen exposure were employed, oxidative stress markers were altered suggesting oxidative skeletal muscle is susceptible to HS- and Zen-mediated changes in redox balance. In total, these data raise the possibility that more severe HS exposures and greater Zen doses, similar to those sometimes encountered in production settings, may compromise muscle health.

Effects of a Short-Term Lipopolysaccharides Challenge on Mouse Brain and Liver Peroxisomal Antioxidant and β-oxidative Functions: Protective Action of Argan Oil.

During sepsis, the imbalance between oxidative insult and body antioxidant response causes the dysfunction of organs, including the brain and liver. Exposing mice to bacterial lipopolysaccharides (LPS) results in a similar pathophysiological outcome. The protection offered by argan oil was studied against LPS-induced oxidative stress, dysregulation of peroxisomal antioxidants, and β-oxidation activities in the brain and liver. In a short-term LPS treatment, lipid peroxidation (malonaldehyde assay) increased in the brain and liver with upregulations of proinflammatory tumor necrosis factor (Tnf)-α and anti-inflammatory interleukin (Il)-10 genes, especially in the liver. Although exposure to olive oil (OO), colza oil (CO), and argan oil (AO) prevented LPS-induced lipid peroxidation in the brain and liver, only AO exposure protected against liver inflammation. Remarkably, only exposure to AO prevented LPS-dependent glutathione (GSH) dysregulation in the brain and liver. Furthermore, exposure to AO increased more efficiently than OO and CO in both organs, peroxisomal antioxidant capacity via induction of catalase (Cat) gene, protein and activity expression levels, and superoxide dismutase (Sod1) mRNA and activity levels. Interestingly, LPS decreased protein levels of the peroxisomal fatty acid-ATP binding cassette (ABC) transporters, ABCD1 and ABCD2, and increased acyl-CoA oxidase 1 (ACOX1) protein expression. Moreover, these LPS effects were attenuated for ABCD1 and ACOX1 in the brain of mice pretreated with AO. Our data collectively highlight the protective effects of AO against early oxidative stress caused by LPS in the brain and liver and their reliance on the preservation of peroxisomal functions, including antioxidant and β-oxidation activities, making AO a promising candidate for the prevention and management of sepsis.

Use of the β-Glucan-Producing Lactic Acid Bacteria Strains Levilactobacillus brevis and Pediococcus claussenii for Sourdough Fermentation-Chemical Characterization and Chemopreventive Potential of In Situ-Enriched Wheat and Rye Sourdoughs and Breads.

The aim of the present study was to examine β-glucan production and the potential prebiotic and chemopreventive effects of wheat and rye sourdoughs and breads generated with wild-type and non-β-glucan-forming isogenic mutant strains of Levilactobacillus brevis and Pediococcus claussenii. Sourdough and bread samples were subjected to in vitro digestion and fermentation. Fermentation supernatants (FS) and pellets (FP) were analyzed (pH values, short-chain fatty acids (SCFA), ammonia, bacterial taxa) and the effects of FS on LT97 colon adenoma cell growth, viability, caspase-2 and -3 activity, genotoxic and antigenotoxic effects and on gene and protein expression of p21, cyclin D2, catalase and superoxide dismutase 2 (SOD2) were examined. Concentrations of SCFA were increased and concentrations of ammonia were partly reduced in the FS. The relative abundance of Bifidobacteriaceae was increased in all FPs. Treatment with FS reduced the growth and viability of LT97 cells and significantly increased caspase-2 and -3 activities without exhibiting genotoxic or antigenotoxic effects. The p21 mRNA and protein levels were increased while that of cyclin D2 was reduced. Catalase and SOD2 mRNA and protein expression were marginally induced. The presented results indicate a comparable chemopreventive potential of wheat and rye sourdoughs and breads without an additional effect of the formed β-glucan.

Improving effect of cordycepin on insulin synthesis and secretion in normal and oxidative-damaged INS-1 cells

Diabetes mellitus (DM) has recently become one of the major diseases that have received attention. Cordycepin (molecular formula: C10H13N5O3), is one of the major bioactive components of Cordyceps militaris, decreases blood glucose levels. In this study, the effect and mechanism of cordycepin in normal and oxidative-damaged INS-1 cells were explored by using cell and molecular biology methods. Results showed that cordycepin could enhance insulin synthesis and secretion. The mechanism is possibly related to the elevated ATP content induced membrane depolarisation and increased Ca2+ concentration. At the genetic level, cordycepin upregulated the mRNA level of insulin, pancreatic duodenal homeobox factor-1 (PDX-1) and glucose transporter 1 (GLUT1). At the protein level, cordycepin promoted the expression of PDX-1, GLUT1, serine threonine kinase (Akt) and phosphorylated Akt (P-Akt). These effects may also contribute to the enhancement of insulin synthesis and secretion. Further analysis revealed that cordycepin protected against H2O2-induced damage on INS-1 cells and improved their viability and insulin synthesis/secretion. This effect should be attributed to the reduced intracellular reactive oxygen species (ROS), enhanced mitochondrial membrane potential (MMP), increased activity of superoxide dismutase (SOD) and upregulated genetic and protein expression of catalase (CAT), PDX-1, GLUT1 and P-Akt. In conclusion, cordycepin promotes insulin synthesis and secretion in normal islet β cells and improves this function in oxidative-damaged islet β cells. Given that islet β cells are vulnerable to oxidative stress, the improving effect of cordycepin on the antioxidant capacity and insulin synthesis/secretion of INS-1 cells may be an important mechanism for its hypoglycaemic effect.

Acetyl l-carnitine protects adipose-derived stem cells against serum-starvation: regulation on the network composed of reactive oxygen species, autophagy, apoptosis and senescence.

Adipose-derived stem cells (ADSCs) play an important role in cell therapy and regenerative medicine. However, local nutritional deficiency often limits therapeutical effect of the transplanted cells. Acetyl l-carnitine (ALC) is a common energy metabolism regulator and free radical scavenger. This study investigated the effect of ALC on ADSCs exposed to severe serum-deprivation and explored the relative machanisms. Treating with 1mM ALC improved proliferation and alleviated senescence of starved cells, accompanied with reduced reactive oxygen species (ROS) and increased protein expression of SOD1 and catalase. In addition, ALC inhibited apoptosis but increased starvation-induced autophagy, which might be related to the regulation of phases of dissociation of Bcl-2-Beclin1 and Bcl-2-Bax complexes. Evidence obtained by replacing ALC with N-acetylcysteine (N-AC) suggested that ROS might be the central inducer of autophagy, apoptosis and senescence. There was a difference between ALC and N-AC in the protection mechanism, that was, compared with N-AC, ALC maintained autophagy well at the same time as anti-oxidation. Inhibition of autophagy by 3-methyladenine (3-MA) partially offset the protective effect of ALC. However, despite low-level ROS and enhanced autophagy, ALC with high concentration (10mM) markedly aggravated cell apoptosis and senescence, thus losing cytoprotection and even causing damage.

Blueberry extract attenuates norepinephrine-induced oxidative stress and apoptosis in H9c2 cardiac cells.

Enhanced sympathetic system activation mediated by norepinephrine (NE) contributes to adverse cardiac remodeling leading to oxidative stress and cell death, progressing to heart failure. Natural antioxidants may help maintain redox balance, attenuating NE-mediated cardiac cell damage. In the present study, we evaluated the effect of a blueberry extract (BBE) on H9c2 cardiac cells exposed to NE on cell death, oxidative stress status and its major signaling pathways. H9c2 cells were pre-incubated with 50μg/ml of BBE for 4h and maintained in the presence of 100μM NE for 24h. NE exposure resulted in increased caspase 3/7 activity. This was associated with reduced protein expression of antioxidants catalase, superoxide dismutase and glutathione peroxidase and increase in 4-hydroxynonenal adduct formation. NE led to increased activity of Protein kinase B (Akt), Forkhead box O3a and AMP-activated protein kinase alpha and decreased activity of Signal transducer and activator of transcription 3. BBE prevented caspases activation and abrogated NE-induced increase in oxidative stress, as well as attenuated the increase in Akt. Based on these findings, it is concluded that BBE promoted cardioprotection of H9c2 cells in an in vitro model of NE-induced oxidative damage, suggesting a cardioprotective role for BBE in response to NE exposure.

HGF AND PROTECTIVE ROLL IN THE INTESTINAL COLLATERAL DAMAGE BY Α-NAFTILISOTIOCIANATO- INDUCE CHOLESTASIS.

he prevalence of cholestasis has been increasing in recent years; The excretion of bile acids via basolateral has been demonstrated to prevent the excessive accumulation in the hepatocyte, and the liver-intestine axis has been seen affected by enterohepatic circulation deregulation. The epithelial permeability loss caused by the tight junction ruptures leads to inflammation and reactive oxygen species (ROS) production. The hepatocyte growth factor is an essential cellular redox regulator and repair growth factor; it has been reported in its relevance in the intestinal mucosa regeneration and proliferative proprieties. This study aims to evaluate the protective effect of HGF in the intestine of animals subjected to cholestatic damage induced by ANIT. Material and methods: Twenty 10-12 weeks-old male CD-1 mice were used. ANIT (60mg/kg) was administrated at the beginning, 24 h later HGF (10 µg/kg) was injected, and 48 h later, the animals were subjected to euthanasia under anesthesia, and serum and intestines were collected. According to the National Institutes of Health of United Stated (NIH) guide, All mice have been cared for and the Norma Oficial Mexicana (NOM), NOM-062-ZOO-1999. The intestinal tissue was fixed and embedded in paraffin for the histological assessment, followed by routine H&E staining. The expression analysis of TNF- alpha, IL-1b and IL.6 were performed by RT-qPCR using a CFX96 Touch thermocycler with 5μg 2x SYBER Green, which included 1000ng of cDNA and 2μl of forward and reverse primers. The protein quantification was evaluated by Western Blot analysis; using 12% polyacrylamide gels, and the primary antibodies for anti-SOD-1, anti-GPx4, anti-Catalase were incubated. Data are presented as the average ± standard error media (SEM) using GrandPad (Prism 8) software. Variance analysis (ANOVA) was used for the statistical analysis and was considered p <0.005 to indicate a statistical significance. Macroscopic changes reveal no apparent effect. Microscopic studies carried out by H&E staining showed a reduction of the intestinal lumen diameter in mice under ANIT treatment compared with Not treated control (NT). Interestingly, ANIT + HGF-treated group showed protective effects preserving lumen and tissue architecture. To corroborate the potential repair effect of HGF treatment to maintain the tissue and thus digestive process, the excreted stool for every group was addressed. The stools excretion level of ANIT- treated mice was significantly reduced compared with the control and co-treated mice. These results indicate that ANIT-cholestasis induce damage in the small intestine. However, results also found a vulnerability in the colon and ileum to cholestasis damage. To determine whether these sections received damage in ANIT- acute cholestasis model, by RT q-PCR, we examined the mRNA expression of inflammatory cytokines, which were increased in ANIT- treatment. By comparison, HGF co-treatment decreases inflammation like the control group. To check if this regulation of inflammation was for the HGF-induced redox regulation we evaluated, the protein expression of SOD-1, GPx4, and catalase. The treatment with HGF increased the expression of antioxidant enzymes of the intestine tissue. These results suggest that the damage in the intestine is supported by the regulation of ROS induced by cholestasis disease. The current study demonstrated how HGF exerts a protective effect in the intestine triggered by ANIT. This effect seems to be the cellular redox regulation seen in the liver and renal tissue. CONACYT: CB-A1-S-38154. The authors declare that there is no conflict of interest.

Catalase Mediates the Inhibitory Actions of PPARδ against Angiotensin II-Triggered Hypertrophy in H9c2 Cardiomyocytes.

Hypertrophy of myocytes has been implicated in cardiac dysfunctions affecting wall stress and patterns of gene expression. However, molecular targets potentially preventing cardiac hypertrophy have not been fully elucidated. In the present study, we demonstrate that upregulation of catalase by peroxisome proliferator-activated receptor δ (PPARδ) is involved in the anti-hypertrophic activity of PPARδ in angiotensin II (Ang II)-treated H9c2 cardiomyocytes. Activation of PPARδ by a specific ligand GW501516 significantly inhibited Ang II-induced hypertrophy and the generation of reactive oxygen species (ROS) in H9c2 cardiomyocytes. These effects of GW501516 were almost completely abolished in cells stably expressing small hairpin (sh)RNA targeting PPARδ, indicating that PPARδ mediates these effects. Significant concentration and time-dependent increases in catalase at both mRNA and protein levels were observed in GW501516-treated H9c2 cardiomyocytes. In addition, GW501516-activated PPARδ significantly enhanced catalase promoter activity and protein expression, even in the presence of Ang II. GW501516-activated PPARδ also inhibited the expression of atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP), which are both marker proteins for hypertrophy. The effects of GW501516 on the expression of ANP and BNP were reversed by 3-amino-1,2,4-triazole (3-AT), a catalase inhibitor. Inhibition or downregulation of catalase by 3-AT or small interfering (si)RNA, respectively, abrogated the effects of PPARδ on Ang II-induced hypertrophy and ROS generation, indicating that these effects of PPARδ are mediated through catalase induction. Furthermore, GW501516-activated PPARδ exerted catalase-dependent inhibitory effects on Ang II-induced hypertrophy by blocking p38 mitogen-activated protein kinase. Taken together, these results indicate that the anti-hypertrophic activity of PPARδ may be achieved, at least in part, by sequestering ROS through fine-tuning the expression of catalase in cardiomyocytes.

Illuminating the effect of beneficial blue light and ROS-modulating enzymes in Dupuytren's disease.

Dupuytren's disease (DD) is a fibroproliferative disorder of the palmar aponeurosis, which is characterized by a compound myofibrogenesis and evidenced by an increased expression of α-smooth muscle actin (α-SMA). In Dupuytren's tissue, higher levels of reactive oxygen species (ROS) are documented, stimulating the proliferation and differentiation of myofibroblasts. Our preliminary study demonstrates that α-SMA-expression is significantly inhibited by blue light irradiation in DD. The objective of this study was to investigate the beneficial effect of blue light irradiation and to elucidate the influence of ROS on myofibrogenesis in the pathogenesis of DD. Therefore, an in-vitro model of human DD fibroblasts was used. DD fibroblasts and control fibroblasts isolated from carpal tunnel syndrome (CTS) were daily irradiated with 40 J/cm2 (λ = 453 nm, 38 mW/cm2). Protein expression of ROS-modulating enzymes (Catalase, NOX4, SOD1, MnSOD) and α-SMA were determined, and additionally analysed after a pharmacological inhibition of the TGF-β1-signaling with SB431542. Furthermore, the protein expression of α-SMA as surrogate parameter for myofibrogenesis was evaluated after applying different concentrations of long-lasting ROS. It could be determined that the beneficial blue light irradiation, which inhibited myofibrogenesis, is mediated by a significant inhibition of catalase protein expression. This effect should be accompanied with an increased intracellular ROS level. Proof of evidence was an H2O2-application on DD fibroblasts, also leading to a decreased myofibrogenesis. Furthermore, it could be demonstrated that endogenous MnSOD was significantly downregulated in resting DD fibroblasts. If DD fibroblasts were treated with the pharmacological inhibitor SB431542, myofibrogenesis was inhibited, but MnSOD expression was simultaneously elevated, which ought to affect ROS level by raising intracellular H2O2 amount. Blue light irradiation as well as the pharmacological action of SB431542 in consequence mediates their beneficial effect on disturbed myofibrogenesis in DD by further increasing ROS level. The present study demonstrates the importance of intracellular ROS homeostasis in DD and illuminates the beneficial effect of blue light as a promising therapy option for DD.

Oxandrolone treatment in juvenile rats induced anxiety-like behavior in young adult animals

AimsOxandrolone (OXA) is a synthetic steroid used for the treatment of clinical conditions associated with catabolic states in humans, including children. However, its behavioral effects are not well known. Our goal was to evaluate the anxiety-like behavior induced in young adult rats after the treatment of juvenile animals with OXA. MethodsFour-week-old male rats were separated into three groups: Control (CON), therapeutic-like OXA dose (TD), and excessive OXA dose (ED), in which 2.5 and 37.5 mg/kg/day of OXA were administered via gavage for four weeks for TD and ED, respectively. Behavior was evaluated through the elevated plus maze (EPM) and open field (OF) tests. Protein expression of catalase (CAT), superoxide dismutase (SOD), Tumor necrosis factor-α (TNF-α), and dopamine receptor 2 (DrD2) were analyzed in tissue samples of the hippocampus, amygdala, and prefrontal cortex by Western Blot. ResultsOXA induced anxiety-like behaviors in both TD and ED animals; it decreased the time spent in the open arms of the EPM in both groups and reduced the time spent in the central zone of the OF in the TD group. In the hippocampus, CAT expression was higher in TD compared with both control and ED animals. No differences were found in the amygdala and prefrontal cortex. TNF-α, SOD, and DrD2 levels were not altered in any of the assessed areas. ConclusionsTreatment of juvenile rats with OXA led to anxiety-like behavior in young adult animals regardless of the dose used, with minor changes in the antioxidant machinery located in the hippocampus.

Melatonin improves the antioxidant capacity in cardiac tissue of Wistar rats after exhaustive exercise

We investigated the effects of melatonin on the onset and resolution of the oxidative stress in the cardiac muscle in melatonin-treated and nontreated rats subjected to an exhaustive exercise session. Forty male rats were divided into: melatonin-treated (20 mg/kg supplemented for 10 d) and control. On the 10th day, each group was subdivided according to euthanasia moments: control or melatonin-treated not exercised (C0h and M0h); immediately after the exercise (CIA and MIA); and 2 h after exercise (C2h and M2h). The heart of animals was removed and the levels of oxidative stress index (OSI) and the formation of thiobarbituric acid reactive substances (TBARS), protein carbonyl, and the activities of aconitase, catalase (CAT), glutathione peroxidase (GPx), and superoxide dismutase (SOD) were evaluated. Total antioxidant status (TAS), total oxidant status (TOS), and the protein expression of CAT, GPx, and SOD was also measured. Our data revealed significant differences on: (i) OSI (p=.029), CAT activity (p=.016), CAT content (p<.001), GPx content (p=.014), reduced glutathione levels (p<.001), and aconitase activity (p<.001) for interaction of melatonin; (ii) GPx activity (p=.005), reduced glutathione (p=.004), protein carbonyl (p=.035), and TBARS levels (p=.028) between groups, and (iii) TBARS levels (p=.016) for significance between moments. Although the exhaustive exercise protocol imposed mild oxidative stress on the cardiac tissue of rats, melatonin induced antioxidant responses that rebalanced the redox status of the cardiac tissue, especially after exhaustive exercise.

Hydrogen peroxide accelerates ventricular arrhythmogenesis by inactivating N‐type calcium channels in cardiac vagal postganglionic neurons in type 2 diabetic rats

Withdrawal of cardiac vagal activity is a common complication in patients with type 2 diabetes mellitus (T2DM), and is associated with arrhythmia-related sudden cardiac death and high mortality in T2DM patients. Although our recent study has demonstrated that reduction of N-type calcium (Cav2.2) currents and cell excitability of cardiac vagal postganglionic (CVP) neurons exacerbates myocardial infarction-evoked ventricular arrhythmias and mortality in T2DM rats, the mechanisms responsible for T2DM-reduced Cav2.2 channel activity in CVP neurons remain unclear. Since reactive oxygen species (ROS) has been reported to modulate calcium channel activity in peripheral postganglionic neurons, here we tested if hydrogen peroxide (H2O2, a type of ROS) overproduction inactivates Cav2.2 channels and further contributes to ventricular vagal dysfunction and ventricular arrhythmogenesis in T2DM. Rat T2DM was induced by a high-fat diet plus streptozotocin injection. Adenoviral catalase gene (Ad.CAT, 2 µl, 1 x 1010 pfu/ml) was microinjected into CVP neurons to scavenge local H2O2 production. Reduced catalase expression accompanied by overproduction of H2O2 was detected in the CVP neurons in T2DM rats. In vivo transfection of Ad.CAT not only increased protein expression of catalase but also attenuated H2O2 production in CVP neurons in T2DM rats. Data from reverse-phase protein array also demonstrated that overexpression of catalase by transfection of Ad.CAT into CVP neurons markedly increased T2DM-reduced protein expression of Cav2.2 in CVP neurons. Local microinjection of Ad.CAT into CVP neurons also significantly restored T2DM-blunted ventricular vagal activity, as demonstrated by an improvement in the response of left ventricular systolic pressure to left vagal efferent nerve stimulation. Additionally, in the power spectral analysis of heart rate variability (HRV), transfection of Ad.CAT into CVP neurons increased the T2DM-attenuated high frequency power (an index of vagal activity), leading to further restoration of the low frequency to high frequency ratio. Moreover, the inducibility of ventricular arrhythmia measured in anesthetized condition was significantly reduced in T2DM rats transfected with Ad.CAT, compared with T2DM rats (0.75±0.48 in T2DM+Ad.CAT group vs. 3.0±1.38 in T2DM group). Transfection of Ad. vector into CVP neurons had no effects on protein expression of catalase and Cav2.2, ventricular vagal activity, HRV, as well as susceptibility to ventricular arrhythmias in T2DM. Based on above data, we conclude that H2O2 overproduction inactivates Cav2.2 channels and further contributes to ventricular vagal dysfunction and ventricular arrhythmogenesis in T2DM, suggesting that H2O2 signaling pathway might be an effective therapeutic target to suppress ventricular arrhythmias in patients with T2DM.

Imperatorin Interferes with LPS Binding to the TLR4 Co-Receptor and Activates the Nrf2 Antioxidative Pathway in RAW264.7 Murine Macrophage Cells.

Imperatorin (IMP) could downregulate several inflammatory transcription factor signaling pathways. Some studies have pointed out that IMP could interfere with toll-like receptor 4 (TLR4) signaling. This study evaluates how IMP interferes with the TLR4 co-receptors signaling through the protein-ligand docking model, Western blotting, immunofluorescence (IF), and atomic force microscopy (AFM) assays in lipopolysaccharide (LPS) stimulated macrophage-like RAW264.7 cells in vitro. The results of the protein-ligand docking demonstrate that IMP interferes with LPS binding to the LPS-binding protein (LBP), the cluster of differentiation 14 (CD14), and the toll-like receptor 4/myeloid differentiation factor 2 (TLR4/MD-2) co-receptors in LPS-stimulated RAW264.7 cells. Compared with TLR4 antagonist CLI-095 or dexamethasone, IMP could suppress the protein expressions of LBP, CD14, and TLR4/MD-2 in LPS-stimulated cells. Furthermore, the three-dimensional (3D) image assay of the AFM showed IMP could prevent the LPS-induced morphological change in RAW264.7 cells. Additionally, IMP could activate the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, and it increased the antioxidative protein expression of heme oxygenase-1 (HO-1), superoxidase dismutase (SOD), and catalase (CAT). Our results are the first to reveal that the anti-inflammatory effect of IMP interferes with LPS binding to TLR4 co-receptor signaling and activates the antioxidative Nrf2 signaling pathway.