SOD2 Protein Expression Research Articles

Lactobacillus plantarum CQPC11 Isolated from Sichuan Pickled Cabbages Antagonizes d-galactose-Induced Oxidation and Aging in Mice.

Chinese pickled cabbage is a traditional fermented food that contains abundant microbes produced during the process of fermentation. In this work, an in vivo animal study was conducted to investigate the effects of a newly isolated lactic acid bacterium (Lactobacillus plantarum CQPC11, LP-CQPC11) on d-galactose-induced oxidation and aging in mice. Analysis of the serum and tissue samples of these mice using molecular biology approaches showed that LP-CQPC11 suppressed the decrease in thymus, brain, heart, liver, spleen, and kidney indices caused by oxidation and aging. Furthermore, LP-CQPC11 increased the levels of SOD (superoxide dismutase), GSH-Px (glutathione peroxidase), and GSH (glutathione), whereas it reduced the levels of NO (nitric oxide) and MDA (malondialdehyde) in the serum, liver, and spleen of oxidation and aging mouse models. Pathological observation indicated that LP-CQPC11 alleviated the damage caused by oxidation and aging on the liver and spleen of mice. qPCR analysis indicated that LP-CQPC11 effectively upregulated the expression of nNOS (neuronal nitric oxide synthase), eNOS (endothelial nitric oxide synthase), Cu/Zn-SOD (cuprozinc-superoxide dismutase), Mn-SOD (manganese superoxide dismutase), CAT (catalase), HO-1 (heme oxygenase-1), Nrf2 (nuclear factor-erythroid 2 related factor 2), γ-GCS (γ-glutamylcysteine synthetase), and NQO1 (NAD(P)H dehydrogenase [quinone] 1), but downregulated the expression of iNOS (inducible nitric oxide synthase) in the mouse liver and spleen. Western blot analysis showed that LP-CQPC11 effectively upregulated SOD1 (Cu/Zn-SOD), SOD2 (Mn-SOD), CAT, GSH1 (c-glutamylcysteine synthetase), and GSH2 (glutathione synthetase) protein expression in mouse liver and spleen tissues. These findings suggest that LP-CQPC11 can effectively prevent d-galactose-induced oxidation and aging in mice, and the effect is even better than that of the commonly used Lactobacillus delbruechii subsp. bulgaricus (LDSB) and vitamin C in the industry. Thus, LP-CQPC11 may be potentially employed as a probiotic strain.

140 - Increased expression and activity of Sod2 promotes transcoelomic metastasis in ovarian cancer

Ovarian cancer is the leading cause of death from gynecologic malignancy due to asymptomatic progression of the disease to advanced stages. Patients with widespread metastases are characterized by a large volume of ascites in the peritoneal cavity within which floating cancer cell populations are found in malignant cases. Metastasis of ovarian cancer occurs when cancer cells escape the primary tumor and adapt to survive under stress conditions associated with anchorage independence in the ascites. We demonstrate that ovarian cancer cells overcome matrix detachment-induced oxidative stress by upregulating expression and activity of the mitochondrial antioxidant enzyme Manganese Superoxide Dismutase 2 (Sod2), and demonstrate that increased expression of Sod2 in anchorage independence is necessary for survival of ovarian cancer cell in vitro and for successful omental metastasis in vivo. Our data reveal a multi-modal regulation of Sod2 in anchorage independence, that includes transcriptional, translational and post-translational regulation at different stages following matrix detachment. In early anchorage independence, a rapid increase in Sod2 protein translation is observed within 30 minutes after cell detachment, without change in mRNA expression. Concomitantly, Sod2 activity is rapidly increased following matrix detachment in a Sirtuin-3 (SIRT3)-dependent manner. We demonstrate that SIRT3, a mitochondrial NAD+ deacetylase, which activates Sod2 by deacetylating key lysine residues including K68, is also necessary for anchorage independent survival and metastasis in vivo. This early post-transcriptional/post-translational regulation of Sod2 is followed by increased mRNA transcription and protein expression of Sod2 within 24 hours of anchorage independence. Concomitantly, we observe increased expression of the stress-response transcription factor Nrf2, a master regulator of antioxidant genes, including Sod2. Collectively, our data suggest that ovarian cancer cells exhibit a multi-modal regulatory system to ensure rapid and sustained expression and activity of Sod2 in response to anchorage independence, which provides protection from mitochondrial oxidative stress associated with transcoelomic metastasis.

Superoxide Dismutase Activity in Small Mesenteric Arteries Is Downregulated by Angiotensin II but Not by Hypertension

Many studies reported reduced antioxidant capacity in the vasculature under hypertensive conditions. However, little is known about the effects of antihypertensive treatments on the regulation of vascular antioxidant enzymes. Thus, we hypothesized that antihypertensive treatments prevent the reduction of antioxidant enzyme activity and expression in the small vessels of angiotensin II-induced hypertensive rats (ANG). We observed the small mesenteric arteries and small renal vessels of normotensive rats (NORM), ANG, and ANG treated with a triple antihypertensive therapy of reserpine, hydrochlorothiazide, and hydralazine (ANG + TTx). Systolic blood pressure was increased in ANG, which was attenuated by 2 weeks of triple therapy (127, 191, and 143 mmHg for NORM, ANG, and ANG + TTx, respectively; p < 0.05). Total superoxide dismutase (SOD) activity in the small mesenteric arteries of ANG was lower than that of NORM. The protein expression of SOD1 was lower in ANG than in NORM, whereas SOD2 and SOD3 expression was not different between the groups. Reduced SOD activity and SOD1 expression in ANG was not restored in ANG + TTx. Both SOD activity and SOD isoform expression in the small renal vessels of ANG were not different from those of NORM. Interestingly, SOD activity in the small renal vessels was reduced by TTx. Between groups, there was no difference in catalase activity or expression in both the small mesenteric arteries and small renal vessels. In conclusion, SOD activity in the small mesenteric arteries decreased by angiotensin II administration, but not by hypertension, which is caused by decreased SOD1 expression.

Abstract 046: Cardiac Mitochondria are Impaired After Transient Neonatal High Oxygen Exposure in a Rat Model of Prematurity-Related Condition

The heart relies on adequate mitochondrial (mito) ATP production to match myocardial demand, mostly through oxidative phosphorylation. Preterm birth (PT) results in ex utero development of an immature myocardium and PT-born individuals represent a newly recognized group at high risk of cardiovascular diseases. Our group has shown that newborn rats exposed to high oxygen (O 2 ), mimicking PT-related conditions, develop O 2 -induced cardiomyopathy (OIC) and dysfunction later in life. However, whether mito impairments prevail in the programmed left ventricle (LV) changes associated with PT is unknown. We aimed to determine whether OIC is associated with altered LV mito characteristics in 4 wks-old rats. Male rat pups were kept with their mother in 80% O 2 (OIC) or room air (Ctrl) from days 3 to 10 of life. Results are mean±SEM; OIC vs. Ctrl are compared using t-test (n=4/group.P&lt;0.05). At 4 wks, extracellular flux analysis of isolated LV cardiomyocytes revealed significantly decreased O 2 consumption rate in OIC (12.14±5.74 vs. 67.39±13.61 pmoles/min/ug). OIC rats show reduced LV mito copy number determined by the ratio between mito and genomic DNA (26.7±6.30 vs. 52.7±7.32), biogenesis markers Pgc1α (0.59±0.14 vs. 1.18±0.25), and citrate synthase (0.20±0.16 vs. 1.60±0.56) mRNA, and reduced Complex IV (0.66±0.15 vs. 1.26±0.09) protein expression. Gene expression (RT-PCR) of key glycolytic enzymes, hexokinase (3.34±0.59 vs. 0.88±0.39) and glucose-6-phosphate dehydrogenase (2.85±0.48 vs. 0.58±0.08) is significantly increased, indicating a shift toward glycolysis. OIC also were found to have higher LV mtROS production (7581±51 vs. 5191±99, fluorescence arbitrary units) and decreased antioxidant SOD2 protein expression (0.84±0.09 vs. 1.04±0.05). Taken together our results indicate that neonatal hyperoxia exposure leads to decreased LV mito biogenesis and function with impaired oxidative phosphorylation capacity in juvenile animals. Further studies are needed to determine the role of long-term mito dysfunction in the increased susceptibility to heart failure observed after deleterious neonatal conditions.

Abstract 515: Bariatric Surgery Reverses Vascular Pathology in Mice With Morbid Obesity and Type 2 Diabetes

Objective: The mechanism of improved insulin sensitivity and vascular function following bariatric surgery remains unclear. We assessed the role of bariatric surgery in coronary arteriolar function in murine type 2 diabetic mice. Approach and Results: The animals were randomly allocated into normal control (Lean) sham, Lean surgery, db/db sham, and db/db surgery (at 5, 10, 20 and 30 days post-operatively). Impaired vasodilation to acetylcholine (ACh) in db/db mice was partially restored by bariatric surgery. Ghrelin administration restored endothelium-dependent dilation in db/db mice, but surgery-induced restoration was disappeared by ghrelin antagonist. Additionally, the incubation of coronary arterioles with IL-6 prevented the restoration of ACh-induced vasodilation. Bariatric surgery attenuated plasma concentration and protein expression of IL-6 in db/db. Bariatric surgery increased protein expression of eNOS and SOD2 in db/db mice. Conclusions: Decreased IL-6 expression after bariatric surgery induce up-regulation of eNOS and SOD2, and down-regulation of iNOS and O 2 .− , leading to restoration of endothelial function in coronary arterioles in type 2 diabetes.

Shock waves increase pulmonary vascular leakage, inflammation, oxidative stress, and apoptosis in a mouse model.

Severe lung damage is a major cause of death in blast victims, but the mechanisms of pulmonary blast injury are not well understood. Therefore, it is important to study the injury mechanism of pulmonary blast injury. A model of lung injury induced by blast exposure was established by using a simulation blast device. The effectiveness and reproducibility of the device were investigated. Eighty mice were randomly divided into eight groups: control group and 3 h, 6 h, 12 h, 24 h, 48 h, 7 days and 14 days post blast. The explosive device induced an explosion injury model of a single lung injury in mice. The success rate of the model was as high as 90%, and the degree of lung injury was basically the same under the same pressure. Under the same conditions, the thickness of the aluminum film can be from 0.8 mm to 1.6 mm, and the peak pressure could be from 95.85 ± 15.61 PSI to 423.32 ± 11.64 PSI. There is no statistical difference in intragroup comparison. A follow-up lung injury experiment using an aluminum film thickness of 1.4 mm showed a pressure of 337.46 ± 18.30 PSI induced a mortality rate of approximately 23.2%. Compared with the control group (372 ± 23 times/min, 85.9 ± 9.4 mmHg, 4.34 ± 0.09), blast exposed mice had decreased heart rate (283 ± 21 times/min) and blood pressure (73.6 ± 3.6 mmHg), and increased lung wet/dry weight ratio(2.67 ± 0.11), marked edematous lung tissue, ruptured blood vessels, infiltrating inflammatory cells, increased NF-κB (4.13 ± 0.01), TNF-α (4.13 ± 0.01), IL-1β (2.43 ± 0.01) and IL-6 (4.65 ± 0.01) mRNA and protein, decreased IL-10(0.18 ± 0.02) mRNA and protein ( P < 0.05). The formation of ROS and the expression of MDA5 (4.46 ± 0.01) and IREα (3.43 ± 0.00) mRNA and protein were increased and the expression of SOD-1 (0.28 ± 0.02) mRNA and protein was decreased ( P < 0.05). Increased expression of Bax (3.54 ± 0.00) and caspase 3 (4.18 ± 0.01) mRNA and protein inhibited the expression of Bcl-2 (0.39 ± 0.02) mRNA and protein. The changes of pulmonary edema, inflammatory cell infiltration, and cell damage factor expression increased gradually with time, and reached the peak at 12-24 h after the outbreak, and returned to normal at 7-14 days. Detonation injury can lead to edema of lung tissue, pulmonary hemorrhage, rupture of pulmonary vessels, induction of early inflammatory responses accompanied by increased oxidative stress in lung tissue cells and increased apoptosis in mice experiencing blast injury. The above results are consistent with those reported in other literatures. It is showed that the mouse lung blast injury model is successfully modeled, and the device can be used for the study of pulmonary blast injury. Impact statement The number of patients with explosive injury has increased year by year, but there is no better treatment. However, the research on detonation injury is difficult to carry out. One of the factors is the difficulty in making the model of blast injury. The laboratory successfully developed and produced a simulation device of explosive knocking through a large amount of literature data and preliminary experiments, and verified the preparation of the simulation device through various experimental techniques. The results showed that the device could simulate the shock wave-induced acute lung injury generated, which was similar to the actual knocking injury. The experimental process was controlled. Under the same condition, there was no statistical difference between the groups. It is possible to realize miniaturization and precision of an explosive knocking simulation device, which is a good experimental tool for further research on the mechanism of organ damage caused by detonation and the development of protective drugs.

L-Carnitine and Resveratrol Action on Oxidative Response and Differentiation in H9c2 Rat Cardiomyocytes

Diabetic heart disease (DHD) results in structural and functional deterioration due to progressive loss of cardiomyocytes and cardiac stem cells, leading to a premature aging of the diabetic heart. The causes of DHD development are not completely clarified, but it is evident that higher oxidative stress of diabetes induces cardiac structural alterations. Recently, the use of stem cells or cardiac progenitors have inspired the most promising therapeutic strategies but these approaches are limited in diabetic patients. Herein we investigate L-Carnitine (LC) and Resveratrol (RSV) actions on oxidative stress and differentiation process in H9c2 rat cardiomyocytes. H9c2, during proliferation or differentiation condition, were treated with LC (0.5 or 5mM) or RSV (0.1 or 25μM). LC significantly enhanced anti-oxidative response by increasing (48 and 72h) mitochondrial SOD2 protein expression and STAT3 activation. LC did not induce p21 translocation at the nuclear level, demonstrating not to have an inhibitory effect on the cell cycle. LC decreased protein synthesis of CaMKII, whose activation is associated with hypertrophic process. In addition, the protective LC effect on H9c2 cardiomyocytes in stress condition was evaluated treating cells with LC 48h before exposure to H2O2: LC significantly reduced intracellular ROS concentration in a dose-dependent manner. RSV 0.1μM while increasing CaMKII content, did not show any effect on the proliferative potential of H9c2. RSV 25μM induced a significant proliferation decrease in favor of H9c2 differentiation, as shown by ERKs pathway activation and α-actinin protein synthesis promotion. RSV 25μM significantly reduced ROS generation induced by rotenone in neo cardiomyotubes (72h). The ability to improve the microenvironment damaged by oxidative stress and to stimulate cardiac progenitor differentiation could recognize LC/RSV as valid adjuvant therapies to preserve cardiomyocytes survival and ameliorate heart regeneration. Disclosure I. Terruzzi: None. P. Senesi: None. A. Montesano: None. F. Vacante: None. L. Luzi: Speaker's Bureau; Self; A. Menarini Diagnostics, AstraZeneca, Eli Lilly and Company. Research Support; Self; Gelesis. Consultant; Self; McKinsey &amp; Company. Speaker's Bureau; Self; Menarini Group, Merck Sharp &amp; Dohme Corp.. Research Support; Self; Novartis AG. Speaker's Bureau; Self; Novo Nordisk A/S. Research Support; Self; Sunstar Foundation. Speaker's Bureau; Self; Smith &amp; Nephew.

Sulforaphane effects on oxidative stress parameters in culture of adult cardiomyocytes

The aim of this study was to analyse the effect of sulforaphane (SFN) in cultures of adult cardiomyocytes, evaluating oxidative stress at different times. Cells were isolated, cultured, and divided into 4 groups: Control, SFN (5μM), H2O2 (5μM), and SFN+H2O2 (5μM both), and subdivided into groups undergoing 1 or 24 h of SFN incubation. After 1 h of incubation, reactive oxygen species production was 40% lower in the SFN group than the Control, and lipid peroxidation was 63% higher in the H2O2 group than the Control, and it was reduced in both of the SFN groups. The SOD activity was 59% higher in groups incubated for 24 h than in those incubated for 1 h. Protein expression of SOD-1 and SOD-2 was higher in the 24-h groups compared to the 1-h groups (55% and 24%, respectively). The Nrf2 protein expression in the 1-h groups was 17% higher than in the 24-h groups, and the SFN + H2O2 group had 40% more Nrf2 than the Control in the 1-h groups. Unlike Nrf2, the PGC-1α expression was 69% higher in the 24-h groups in relation to the 1-h groups. Regarding the 24-h groups, the SFN and SFN+H2O2 groups were higher than the Control (32% and 33%, respectively), and the SFN+H2O2 group was increased (21%) compared to H2O2. SFN had a protective action against oxidative damage, but had no effect on the antioxidant enzymes analyzed. The different responses in the expression of Nrf2 and PGC-1α in relation to the incubation times, draws attention to the importance of establishing a timeline of the action of SFN, since there appears to be a temporal difference in its mechanism in adult cardiomyocytes.

Effect of pyrrolidine dithiocarbamate on oxidative stress and mitochondrial function of lung tissue in mice with acute lung injury

To investigate the effects of pyrrolidine dithiocarbamate (PDTC) on oxidative stress and mitochondrial function of lung tissue in mice with acute lung injury (ALI) induced by lipopolysaccharide (LPS). Forty female Balb/c mice were randomly divided into normal saline (NS) control group, LPS model group, PDTC group, and PDTC+LPS group, with 10 mice in each group. The model of mice with ALI was reproduced by intraperitoneal injection of 15 mg/kg LPS. PDTC was administered intraperitoneally with 50 mg/kg PDTC 1 hour before LPS treatment in the PDTC+LPS group. The mice in NS control group was given intraperitoneal injection of 0.1 mL NS only, and those in PDTC group was given intraperitoneal injection of 50 mg/kg PDTC only. The mice were sacrificed at 24 hours after model reproduction, and the lung tissues were harvested. The total antioxidant capacity (T-AOC) of lung tissue was measured by spectrophotometric kits. The content of malondialdehyde (MDA) was determined by thiobarbituric acid reactive substances assay. The protein expressions of superoxide dismutases (SOD1, SOD2) and catalase (CAT) in lung tissue were determined by Western Blot. Mitochondria from mouse lungs were isolated, and adenosine triphosphate (ATP) synthesis was measured with a luciferase/luciferin-based approach. The mitochondrial membrane potential (ΔΨm) was estimated by using Rhodamine. The mRNA expressions of mitochondrial uncoupling proteins (UCPs) were determined by reverse transcription-polymerase chain reaction (RT-PCR). LPS stimulation could significantly increase oxidative stress in lung tissue of mice and lead to mitochondrial dysfunction. The results showed that the protein expressions of T-AOC and SOD1 were decreased, the level of MDA was increased, the ATP synthesis was decreased in the mitochondrial, the ΔΨm was decreased, and the mRNA expression of UCP2 was decreased. However, there was no significant change in the expressions of SOD2, CAT in lung tissue and UCP1, UCP3 in the mitochondria. Pretreatment with PDTC could obviously alleviate the increase in LPS-induced oxidative stress in lung tissue and mitigate mitochondrial dysfunction. Compared with the LPS model group, T-AOC in lung tissue of PDTC+LPS group was significantly increased (U/g: 0.35±0.08 vs. 0.31±0.07), the level of MDA was significantly decreased (μmol/mg: 13.29±1.13 vs. 17.54±1.72), the protein expression of SOD1 was significantly upregulated (SOD1 protein: 1.13±0.11 vs. 0.71±0.09), ATP synthesis was significantly increased in the mitochondrial (μmol/mg: 49.23±5.42 vs. 36.92±2.21), ΔΨm was significantly increased (mV: 226.03±11.69 vs. 194.86±7.79), and the mRNA expression of UCP2 was significantly increased (2-ΔΔCt: 0.88±0.06 vs. 0.73±0.04). The differences were statistically significant (all P < 0.05). In lung tissue of normal mice, PDTC treatment also had the effect of anti-oxidizing, reducing oxidative stress and promoting ATP synthesis in the mitochondrial. Compared with the NS control group, the level of T-AOC (U/g: 0.49±0.09 vs. 0.43±0.06) and the protein expressions of SOD2 and CAT (SOD2 protein: 1.33±0.08 vs. 1.00±0.11, CAT protein: 1.39±0.08 vs. 1.00±0.11), and ATP synthesis in the mitochondrial of PDTC group was significantly increased (μmol/mg: 61.53±4.92 vs. 53.33±3.20), MDA was significantly decreased (μmol/mg: 10.27±1.25 vs. 12.27±1.36), with statistical differences, but had no effect on the protein expression of SOD1 in lung tissue and ΔΨm and UCPs mRNA expressions in mitochondrion. LPS can induce ALI in mice, increased oxidative stress in lung tissue, and induce mitochondrial dysfunction by inhibiting ATP synthesis. PDTC pretreatment has anti-oxidative effect on LPS-induced ALI, and can mitigate mitochondrial dysfunction.

Strong SOD2 expression and HPV-16/18 positivity are independent events in cervical cancer.

It is well known that persistent infection with high-risk HPV (hr-HPV), mostly HPV-16 and 18, is the main cause of cervical cancer development. Manganese superoxide dismutase (MnSOD or SOD2) are highly expressed in different neoplasia. The present study investigated SOD2 protein expression and the presence of hr-HPV types in 297 cervical samples including non-neoplastic tissue, cervical intraepithelial neoplasia grade 3 (CIN3), squamous cell carcinoma (SCC) and adenocarcinoma (ADC). Strong SOD2 expression was significantly higher in ADC (82%) than CIN3 (52%) or SCC (64%). There was no association between SOD2 expression and HPV 16 and/or 18 detection for every lesion analyzed. Binary Logist Regression revealed that strong SOD2 expression (OR: 27.50, 6.16-122.81) and HPV 16 and/or HPV 18 (OR: 12.67, 4.04-39.74) were independently more associated with CIN3 than non-neoplastic cervix. Strong SOD2 expression (OR: 3.30, 1.23-8.86) and HPV 16 and/or HPV 18 (OR: 3.51, 1.03-11.87) were independently more associated with ADC than SCC. Similar findings for SOD2 expression were observed by the Cochran Mantel-Haenszel test, controlling for HPV-16 and/or HPV 18. In conclusion, the expression of SOD2 was increased in CIN3 and SCC, and more increased in cervical ADC than in SCC. Strong SOD2 expression was statistically independent of the presence of HPV 16 and/or 18. These findings suggest that the mitochondrial antioxidant system and HPV infection could follow independent pathways in the carcinogenesis of cervical epithelium and in the differentiation to SCC or ADC of the cervix.

Effect of GAPDS overexpression on high glucose-induced oxidative damage

The occurrence of infertility in diabetic patients is attributed to oxidative damage of peroxidized products. High glucose-induced mitochondrial oxidative stress and glycolytic enzyme inactivation is considered to be an important mediator for sperm dysfunction. In this study, we successfully constructed TM3-GAPDS stable strain and investigated the role of sperm specific glyceraldehyde-3-phosphate dehydrogenase (GAPDS) on high glucose-induced apoptosis in TM3 cells. High glucose decreased the protein expression of SOD2 and catalase, while the level of intracellular ROS and the apoptosis - related protein increased in TM3 cells. Furthermore, high glucose-induced oxidative stress and apoptosis were reversed by GAPDS overexpression or antioxidant treatment. In conclusion, our data suggest that GAPDS overexpression antagonize high glucose-induced apoptosis by controlling ROS accumulation in TM3 cells.

Scutellarin protects against vascular endothelial dysfunction and prevents atherosclerosis via antioxidation

BackgroundScutellarin is the major constituent responsible for the clinical benefits of Erigeron breviscapus (Vant.) Hand.-Mazz which finds a long history of ethnopharmacological use in Traditional Chinese Medicine. Scutellarin as a pure compound is now under investigation for its protections against various tissue injuries. PurposeThis study aims to examine the effects of scutellarin on oxidative stress-induced vascular endothelial dysfunction and endothelial cell damage, and then to evaluate the therapeutic efficacy of scutellarin in preventing atherosclerosis in rats. MethodsRadical scavenging ability of scutellarin was determined in vitro. Impact of scutellarin on endothelium-dependent relaxation (EDR) of rabbit thoracic aortic rings upon 1, 1-diphenyl-2-picrylhydrazyl (DPPH) challenge was measured. Influences of scutellarin pre-treatment on the levels of reactive oxygen species (ROS), activities of antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase and catalase, and the expression of SOD1 and NADPH oxidase 4 (Nox4) in human umbilical vein endothelial cells (HUVECs) injured by H2O2 were examined. Anti-atherosclerotic effect of scutellarin was evaluated in rats fed with high fat diet (HFD). ResultsScutellarin showed potent antioxidant activity in vitro. Pretreatment of scutellarin retained the EDR of rabbit thoracic aortic rings damaged by DPPH. In H2O2 injured-HUVECs the deleterious alterations in ROS levels and antioxidant enzymes activity were reversed by scutellarin and the mRNA and protein expression of SOD1 and Nox4 were restored also. Oral administration of scutellarin dose-dependently ameliorated hyperlipidemia in HFD-fed rats and alleviated oxidative stress in rat serum, mimicking the effects of reference drug atorvastatin. ConclusionScutellarin protects against oxidative stress-induced vascular endothelial dysfunction and endothelial cell damage in vitro and prevents atherosclerosis in vivo through antioxidation. The results rationalize further investigation into the clinical use of scutellarin in cardiovascular diseases.

Cardiac and placental mitochondrial characterization in a rabbit model of intrauterine growth restriction

BackgroundIntrauterine growth restriction (IUGR) is associated with cardiovascular remodeling persisting into adulthood. Mitochondrial bioenergetics, essential for embryonic development and cardiovascular function, are regulated by nuclear effectors as sirtuins. A rabbit model of IUGR and cardiovascular remodeling was generated, in which heart mitochondrial alterations were observed by microscopic and transcriptomic analysis. We aimed to evaluate if such alterations are translated at a functional mitochondrial level to establish the etiopathology and potential therapeutic targets for this obstetric complication. MethodsHearts and placentas from 16 IUGR-offspring and 14 controls were included to characterize mitochondrial function. ResultsEnzymatic activities of complexes II, IV and II + III in IUGR-hearts (−11.96 ± 3.16%; −15.58 ± 5.32%; −14.73 ± 4.37%; p < 0.05) and II and II + III in IUGR-placentas (−17.22 ± 3.46%; p < 0.005 and −29.64 ± 4.43%; p < 0.001) significantly decreased. This was accompanied by a not significant reduction in CI-stimulated oxygen consumption and significantly decreased complex II SDHB subunit expression in placenta (−44.12 ± 5.88%; p < 0.001). Levels of mitochondrial content, Coenzyme Q and cellular ATP were conserved. Lipid peroxidation significantly decreased in IUGR-hearts (−39.02 ± 4.35%; p < 0.001), but not significantly increased in IUGR-placentas. Sirtuin3 protein expression significantly increased in IUGR-hearts (84.21 ± 31.58%; p < 0.05) despite conserved anti-oxidant SOD2 protein expression and activity in both tissues. ConclusionsIUGR is associated with cardiac and placental mitochondrial CII dysfunction. Up-regulated expression of Sirtuin3 may explain attenuation of cardiac oxidative damage and preserved ATP levels under CII deficiency. General significanceThese findings may allow the design of dietary interventions to modulate Sirtuin3 expression and consequent regulation of mitochondrial imbalance associated with IUGR and derived cardiovascular remodeling.

Effects of 17β-Estradiol on Activity, Gene and Protein Expression of Superoxide Dismutases in Primary Cultured Human Lens Epithelial Cells

ABSTRACTPurpose: Protective effects of estradiol against H2O2-induced oxidative stress have been demonstrated in lens epithelial cells. The purpose of this study was to investigate the effects of 17β-estradiol (E2) on the different superoxide dismutase (SOD) isoenzymes, SOD-1, SOD-2, and SOD-3, as well as estrogen receptors (ERs), ERα and ERβ, in primary cultured human lens epithelial cells (HLECs).Materials and methods: HLECs were exposed to 0.1 µM or 1 µM E2 for 1.5 h and 24 h after which the effects were studied. Protein expression and immunolocalization of SOD-1, SOD-2, ERα, and ERβ were studied with Western blot and immunocytochemistry. Total SOD activity was measured, and gene expression analyses were performed for SOD1, SOD2, and SOD3.Results: Increased SOD activity was seen after 1.5 h exposure to both 0.1 µM and 1 µM E2. There were no significant changes in protein or gene expression of the different SODs. Immunolabeling of SOD-1 was evident in the cytosol and nucleus; whereas, SOD-2 was localized in the mitochondria. Both ERα and ERβ were immunolocalized to the nucleus, and mitochondrial localization of ERβ was evident by colocalization with MitoTracker. Both ERα and ERβ showed altered protein expression levels after exposure to E2.Conclusions: The observed increase in SOD activity after exposure to E2 without accompanying increase in gene or protein expression supports a role for E2 in protection against oxidative stress mediated through non-genomic mechanisms.

Superoxide Anion Production and Bioenergetic Profile in Young and Elderly Human Primary Myoblasts.

Sarcopenia is the age-related loss of skeletal muscle mass, strength, and function. It is associated with regenerative difficulties by satellite cells, adult muscle stem cells, and alteration of oxidative management, mainly the increase in superoxide anions (O2 •−). We aimed to investigate the relation between regenerative deficit in elderly and increase in O2 •− production along with mitochondrial alterations. Myoblasts and myotubes from skeletal muscle of young and elderly healthy subjects (27.8 ± 6 and 72.4 ± 6.5 years old) were measured: (1) superoxide dismutase activity and protein content, (2) mitochondrial O2 •− production levels, (3) O2 •− production variability, and (4) mitochondrial bioenergetic profile. Compared to young myoblasts, elderly myoblasts displayed decreased SOD2 protein expression, elevated mitochondrial O2 •− baseline levels, and decreased oxidative phosphorylation and glycolysis. Additionally, elderly versus young myotubes showed elevated mitochondrial O2 •− levels when stressed with N-acetyl cysteine or high glucose and higher glycolysis despite showing comparable oxidative phosphorylation levels. Altogether, the elderly may have less metabolic plasticity due to the impaired mitochondrial function caused by O2 •−. However, the increased energy demand related to the differentiation process appears to activate compensatory mechanisms for the partial mitochondrial dysfunction.

Fumonisin B1 induces oxidative stress in oesophageal (SNO) cancer cells

Fumonisin B1 (FB1) is a ubiquitous contaminant of maize that is epidemiologically linked to oesophageal cancer (OC) in South Africa. FB1-induced oxidative stress mediates toxicity in animals and human cell lines, but the effects relating to OC are limited. Given the species-specific effects of FB1, this study investigated FB1-mediated toxicity and oxidative stress in spindle-shaped N-cadherin (+) CD45 (−) osteoblastic (SNO) cells. Following exposure to FB1 (0–20 μM) for 48 h, mitochondrial membrane potential and intracellular reactive oxygen species (iROS) were measured (flow cytometry). Malondialdehyde concentration (lipid peroxidation) was determined spectrophotometrically. ATP and reduced glutathione (GSH) concentrations were quantified using luminometry, gene expression of SOD2 by qPCR and protein expression of SOD2, GPx1, Nrf2 and HSP70 by western blotting. Mitochondrial depolarization increased at 10 μM and 20 μM FB1, with a concomitant reduction in ATP, iROS and GSH at both concentrations. Lipid peroxidation increased at 10 μM FB1 exposure. While transcript levels of SOD2 were significantly increased, protein levels decreased. Protein expression of GPx1, Nrf2 and HSP70 increased. In contrast to the 10 μM and 20 μM FB1 treatment, mitochondrial depolarization decreased at 1.25 μM FB1. Intracellular ROS and ATP were decreased and lipid peroxidation increased. Decreased GSH was accompanied by a decrease in GPx1 protein levels, and increased HSP70 and Nrf2. SOD2 expression and protein levels were significantly increased. Overall these results indicate that FB1 caused increased ROS that were counteracted by engaging the antioxidant defense. Furthermore, the peculiar response at 1.25 μM FB1 is noteworthy, as compared to the other two concentrations tested.

Skeletal muscle and performance adaptations to high-intensity training in elite male soccer players: speed endurance runs versus small-sided game training

PurposeTo examine the skeletal muscle and performance responses across two different exercise training modalities which are highly applied in soccer training.MethodsUsing an RCT design, 39 well-trained male soccer players were randomized into either a speed endurance training (SET; n = 21) or a small-sided game group (SSG; n = 18). Over 4 weeks, thrice weekly, SET performed 6–10 × 30-s all-out runs with 3-min recovery, while SSG completed 2 × 7–9-min small-sided games with 2-min recovery. Muscle biopsies were obtained from m. vastus lateralis pre and post intervention and were subsequently analysed for metabolic enzyme activity and muscle protein expression. Moreover, the Yo–Yo Intermittent Recovery level 2 test (Yo–Yo IR2) was performed.ResultsMuscle CS maximal activity increased (P < 0.05) by 18% in SET only, demonstrating larger (P < 0.05) improvement than SSG, while HAD activity increased (P < 0.05) by 24% in both groups. Na+–K+ ATPase α1 subunit protein expression increased (P < 0.05) in SET and SSG (19 and 37%, respectively), while MCT4 protein expression rose (P < 0.05) by 30 and 61% in SET and SSG, respectively. SOD2 protein expression increased (P < 0.05) by 28 and 37% in SET and SSG, respectively, while GLUT-4 protein expression increased (P < 0.05) by 40% in SSG only. Finally, SET displayed 39% greater improvement (P < 0.05) in Yo–Yo IR2 performance than SSG.ConclusionSpeed endurance training improved muscle oxidative capacity and exercise performance more pronouncedly than small-sided game training, but comparable responses were in muscle ion transporters and antioxidative capacity in well-trained male soccer players.

The additive effects of atorvastatin and insulin on renal function and renal organic anion transporter 3 function in diabetic rats

Hyperglycemia-induced oxidative stress is usually found in diabetic condition. 3-hydroxy-3-methylglutaryl coenzyme-A (HMG-CoA) reductase inhibitors, statins, are widely used as cholesterol-lowering medication with several “pleiotropic” effects in diabetic patients. This study aims to evaluate whether the protective effects of atorvastatin and insulin on renal function and renal organic anion transporter 3 (Oat3) function involve the modulation of oxidative stress and pancreatic function in type 1 diabetic rats. Type 1 diabetes was induced by intraperitoneal injection of streptozotocin (50 mg/kg BW). Atorvastatin and insulin as single or combined treatment were given for 4 weeks after diabetic condition had been confirmed. Diabetic rats demonstrated renal function and renal Oat3 function impairment with an increased MDA level and decreased SOD protein expression concomitant with stimulation of renal Nrf2 and HO-1 protein expression. Insulin plus atorvastatin (combined) treatment effectively restored renal function as well as renal Oat3 function which correlated with the decrease in hyperglycemia and oxidative stress. Moreover, pancreatic inflammation and apoptosis in diabetic rats were ameliorated by the combined drugs treatment. Therefore, atorvastatin plus insulin seems to exert the additive effect in improving renal functionby alleviating hyperglycemiaand the modulation of oxidative stress, inflammation and apoptosis.

Expression of Hypoxia-Inducible Factor-1α and Association with Cu/Zn Superoxide Dismutase in the Reproductive Tract Tissues of Adult Male Roe Deer (Capreolus capreolus) During the Reproduction Season

Abstract This study aimed to determine the mRNA and protein expressions of HIF-1α and SOD-1 in testes and epididymis of adult male roe deer during the periods of reproductive season. The mRNA expression levels of investigated factors were examined using RT-PCR, while protein expression levels were evaluated using Western blot analysis. Differences between tissues within this period and between periods were evaluated. Testicular tissue exhibited lower HIF-1α and SOD-1 mRNA expression levels during all periods as compared to epididymal tissues. Among the epididymal tissues, cauda epididymis showed higher HIF-1α mRNA expression levels in comparison with caput and corpus tissues in the pre-rut period, and not much lower SOD-1 mRNA expression levels in comparison with caput in all periods. Protein expression of HIF-1α was higher in the epididymis than in the testicular tissues in the pre-rut and rut periods. Its expression was the highest in caput epididymal tissue compared with either the testicular, corpus and cauda epididymal tissue during rut and higher than corpus and testis in pre- and post-rut periods. Testicular tissue was characterized by lower SOD-1 protein expression levels than in epididymis in pre-rut and rut periods. This study indicates that the reproductive tract of male roe deer possesses a tissue-specific defense system which protects the tissues from oxygen deficiency damage, which is important during cyclic changes occurring in the reproductive tissues during the reproductive season.

Indonesian herbal medicine prevents hypertension-induced left ventricular hypertrophy by diminishing NADPH oxidase-dependent oxidative stress.

Indonesian herbal medicine Centella asiatica, Justicia gendarussa and Imperata cylindrica decoction (CJID) are known to be efficacious for hypertension. Oxidative stress plays an important role in hypertension-induced left ventricular hypertrophy (H-LVH). This study evaluated whether CJID inhibit cardiac remodeling in spontaneously hypertensive rats (SHRs) through mechanism of oxidative stress-related cardiac-NADPH oxidase (NOXs) pathway: NOX1, NOX2 and NOX4. Forty-weeks-old SHRs and normotensive-WKY rats, were both randomly divided into 2 groups: CJID and control. All rats were treated for 5 weeks. Systolic blood pressure (SBP) and heart rate (HR) were measured. LV morphology, function and performance were assessed by histological staining and echocardiography. Serum and cardiac superoxide dismutase (SOD) activity and malondialdehyde (MDA) content were assessed. Cardiac superoxide and hydrogen peroxide (H2O2) productions, protein expressions of SOD2, SOD3, NOX1, NOX2 and NOX4 were also determined. We found that SBP and HR were significantly decreased in SHRs-treated group. Echocardiography showed that CJID significantly improved LV morphometry and function. CJID decreased MDA level, but increased SOD activity. Cardiac superoxide and H2O2 generation were decreased in SHRs-treated group. CJID caused cardiac SODs expressions to be increased but NOXs expressions to be suppressed. In conclusion, CJID prevents H-LVH by reducing reactive oxygen species production via the NOXs-dependent pathway.