Reactive Oxygen Species Cause Research Articles

Piper crocatum Ameliorates APAP-Induced Hepatotoxicity through Antioxidant and Anti-Inflammatory Mechanisms

Cirrhosis is a serious hepatic disease that occurs worldwide and is caused by progressive fibrosis in the liver. Free radicals are the major cause of reactive oxygen species (ROS) generation, which affects the balance of the liver metabolism. Piper crocatum, commonly called red betel, is widely used in ethnomedicine because it has biological capabilities, including antioxidant and anti-inflammatory properties. The present study was undertaken to examine the possibility of hepaprotective activity of red betel extract on acetaminophen (APAP)-induced HepG2 cells as the cirrhosis in vitro model. Assessment of red betel extract (RBE) was performed using the colorimetric method for the quantification of LDH, AST, and ALT. As preliminary study, a cytotoxicity assay was performed at various RBE concentrations (100.00; 50.00; 25.00; 12.50; 6.25; 3.13 μg/ml) using an MTS assay to decide the safe concentration based on cell viability. The RBE treatment did not affect cell viability, even at a high concentration (100 µg/ml, p > 0.05). RBE at concentrations 25 and 100 µg/ml successfully reduced LDH, AST, and ALT activities in the hepatotoxic model in a dose-dependent manner. In the APAP-induced hepatotoxicity model, RBE positively indicated a hepatoprotective effect and cell amelioration by a decrease in hepatotoxic markers.

Dihydroceramide desaturase regulates the compartmentalization of Rac1 for neuronal oxidative stress.

Disruption of sphingolipid homeostasis is known to cause neurological disorders, but the mechanisms by which specific sphingolipid species modulate pathogenesis remain unclear. The last step of de novo sphingolipid synthesis is the conversion of dihydroceramide to ceramide by dihydroceramide desaturase (human DEGS1; Drosophila Ifc). Loss of ifc leads to dihydroceramide accumulation, oxidative stress, and photoreceptor degeneration, whereas human DEGS1 variants are associated with leukodystrophy and neuropathy. In this work, we demonstrate that DEGS1/ifc regulates Rac1 compartmentalization in neuronal cells and that dihydroceramide alters the association of active Rac1 with organelle-mimicking membranes. We further identify the Rac1-NADPH oxidase (NOX) complex as the major cause of reactive oxygen species (ROS) accumulation in ifc-knockout (ifc-KO) photoreceptors and in SH-SY5Y cells with the leukodystrophy-associated DEGS1H132R variant. Suppression of Rac1-NOX activity rescues degeneration of ifc-KO photoreceptors and ameliorates oxidative stress in DEGS1H132R-carrying cells. Therefore, we conclude that DEGS1/ifc deficiency causes dihydroceramide accumulation, resulting in Rac1 mislocalization and NOX-dependent neurodegeneration.

Ursodeoxycholyl lysophosphatidylethanolamide protects against hepatic ischemia/reperfusion injury via phospholipid metabolism-mediated mitochondrial quality control.

Mitochondrial dysfunction is the leading cause of reactive oxygen species (ROS) burst and apoptosis in hepatic ischemia/reperfusion (I/R) injury. Ursodeoxycholyl lysophosphatidylethanolamide (UDCA-LPE) is a hepatotargeted agent that exerts hepatoprotective roles by regulating lipid metabolism. Our previous studies have shown that UDCA-LPE improves hepatic I/R injury by inhibiting apoptosis and inflammation. However, the role of UDCA-LPE in lipid metabolism and mitochondrial function in hepatic I/R remains unknown. In the present study, we investigated the role of UDCA-LPE in hepatic I/R by focusing on the interface of phospholipid metabolism and mitochondrial homeostasis. Livers from 28-week-old mice, primary hepatocytes and HepG2 cells were subjected to in vivo and in vitro I/R, respectively. Analyses of oxidative stress, imaging, ATP generation, genetics, and lipidomics showed that I/R was associated with mitochondrial dysfunction and a reduction in phospholipids. UDCA-LPE alleviated mitochondria-dependent oxidative stress and apoptosis and prevented the decrease of phospholipid levels. Our study found that cytosolic phospholipase A2 (cPLA2 ), a phospholipase that is activated during I/R, hydrolyzed mitochondrial membrane phospholipids and led to mitochondria-mediated oxidative stress and apoptosis. UDCA-LPE inhibited the interaction between cPLA2 and mitochondria and reduced phospholipid hydrolysis-mediated injury. UDCA-LPE might regulate the crosstalk between the phospholipid metabolism and the mitochondria, restore mitochondrial function and ameliorate I/R injury.

Hypertension Editors' Picks Preeclampsia.

Hypertension Editors' Picks Preeclampsia.

Role of Mitochondrial Dysfunction and Reactive Oxygen Species in Mediating Hypertension in the Reduced Uterine Perfusion Pressure Rat Model of Preeclampsia.

Placental ischemia is believed to be the initial event in the development of preeclampsia. Mitochondrial dysfunction is a cause of reactive oxygen species (ROS) generation and oxidative stress, however, there are not many studies examining the role of mitochondrial ROS in the pathology of preeclampsia. The purpose of this study was to not only examine the effect of placental ischemia on mitochondrial-mediated oxidative stress in reduced uterine perfusion pressure (RUPP) rat model of preeclampsia but to also examine the role of mitochondrial ROS in contributing to hypertension in response to placental ischemia. Female pregnant Sprague Dawley rats were used in this study. On gestational day 14, RUPP surgery was performed. On gestational day 19, blood pressure (mean arterial pressure) was measured, placentas and kidneys were collected from normal pregnant and RUPP rats and processed for mitochondrial respiration, ROS, and oxidative phosphorylation enzyme activities. Renal and placental complex activities, expressions and respiration rates were significantly reduced and mitochondrial ROS was increased in RUPP versus normal pregnant mitochondria. Mean arterial pressure was elevated in RUPP (n=6) compared with normal pregnant rats (n=5; 126±4 versus 103±4 mm Hg; P<0.05) and treatment with mitochondrial-specific antioxidants (MitoQ/MitoTEMPO) significantly reduced mean arterial pressure in RUPPs (n=5-10). Mitochondrial ROS was significantly elevated in endothelial cells incubated with RUPP serum compared from with normal pregnant rats, whereas serum from mito antioxidant-treated RUPP rats attenuated this response. Impaired mitochondrial function and vascular, placental, and renal mitochondrial ROS play an important role in hypertension and reduced fetal weight in response to placental ischemia during pregnancy.

Regulation of Cytochrome C by Phosphorylation: Mitochondrial Respiration and Apoptosis

Mammalian cytochrome c (Cytc) is a small globular protein and functions as a mobile electron carrier in the electron transport chain (ETC), shuttling electrons from bc1 complex to cytochrome c oxidase (COX). In addition, Cytc participates in type II apoptosis, when it is released from the mitochondria. Such a molecule, central to life and death decisions of the cell, should be tightly controlled. We previously showed that Cytc is regulated by cell signaling pathways in a highly tissue-specific manner and that Cytc isolated from heart and liver is phosphorylated on Tyr97 and Tyr48, respectively. Here we add another phosphorylation site and show that Cytc isolated from kidney is phosphorylated on Thr28. This phosphorylation is likely mediated by AMP kinase. Phosphorylated and Thr28Glu phosphomimetic Cytc cause partial inhibition in the reaction with COX supporting our model that under healthy conditions ETC proteins are phosphorylated to limit electron flux in the electron transport chain, which in turn prevents a hyperpolarization of the mitochondrial membrane potential, a known cause of reactive oxygen species (ROS) production and trigger of apoptosis. To test our model we generated Cytc knockout cells expressing Thr28Glu phosphomimetic Cytc. We observed reduction of intact cellular respiration, mitochondrial membrane potential, and ROS. Finally, high resolution crystal structure analyses and molecular dynamics simulations suggest that the Thr28-containing epitope is the most flexible internal part of Cytc, which is in close proximity to key residues on COX and thus suggests a mechanism for the altered enzyme kinetics.

Abstract 031: Placental Ischemia Causes Mitochondrial Dysfunction in Reduced Uterine Perfusion Pressure Rats

Introduction: Placental ischemia is believed to be the initial event in the development of preeclampsia (PE). PE is characterized by new onset hypertension and is associated with reduced fetal weight and placental oxidative stress. Mitochondrial dysfunction is the major cause of reactive oxygen species (ROS) generation. We hypothesize that the placental ischemia causes mitochondrial dysfunction in the reduced uterine perfusion pressure (RUPP) rat model of PE. Thus, the purpose of this study is to examine the effect of placental ischemia on mitochondrial function. Methods: Female Sprague Dawley rats were dived into two groups; normal pregnant (NP) and RUPP rats. On gestational day (GD) 14, RUPP surgery was performed, GD18 carotid catheters were inserted, and GD19 conscious blood pressure (MAP) was measured. GD 19 placentas were collected and mitochondria were isolated for respiration measurements. Respiration measurements included: basal state (isolated mitochondria with no substrates), state 2 (glutamate and malate as complex I substrates), state 3 (ADP stimulated), leak state (oligomycin induced ATP synthase inhibition), and maximal state (FCCP stimulated uncoupled). Cytochrome C oxidase (complex IV) activity was also measured in isolated mitochondria. Results: MAP was elevated in RUPP (n=16) compared to NP rats (n=14) (119±2 vs. 100±2 mmHg, p&lt;0.05). State 3 (149±8 vs 220±12 pmol/sec/mg, p&lt;0.05) and maximal (123±8 vs 159±5 pmol/sec/mg, p&lt;0.05) respiration rates were significantly reduced in RUPP (n=5) vs NP (n=7) rats. However, basal, state 2, and state 4 respiratory rates were not different between RUPP vs NP. Respiratory control ratio (state 3/ state 4) was significantly reduced in RUPP (n=5) vs NP (n=7) (7±1 vs 10±1, p&lt;0.05). Complex IV activity was reduced in RUPP (n=5) vs. NP (n=6) rats (508±14 vs 644±17 nmol e-/min/mg, P&lt;0.05). Conclusion: Reduced mitochondrial respiration and cytochrome c oxidase activity, indicators of mitochondrial dysfunction, occur in response to placental ischemia during pregnancy. Thus, suggesting that mitochondrial dysfunction may contribute to the pathophysiology of oxidative stress and hypertension during PE.

Labisia pumila extract protects skin cells from photoaging caused by UVB irradiation

Labisia pumila (Myrsinaceae), known as "Kacip Fatimah," has been used by many generations of Malay women to induce and facilitate child birth as well as a post partum medicine. However, its topical application on skin has not been reported yet. In this study, we have focused on the anti-photoaging effects of L. pumila. Extract of L. pumila was first analyzed for their antioxidant activities using DPPH (2,2-diphenyl-1-picrylhydrazyl) since UV irradiation is a primary cause of reactive oxygen species (ROS) generation in the skin. The 50% free radical scavenging activity (FSC(50)) of L. pumila extract was determined to be 0.006%, which was equal to that produced by 156 microM ascorbic acid. TNF-alpha and cyclooxygenase (COX-2) play a primary role in the inflammation process upon UV irradiation and are known to be stimulated by UVB. Treatment with L. pumila extract markedly inhibited the TNF-alpha production and the expression of COX-2. Decreased collagen synthesis of human fibroblasts by UVB was restored back to normal level after treatment with L. pumila extract. On the other hand, the enhanced MMP-1 expression upon UVB irradiation was down regulated by L. pumila extract in a dose-dependent manner. Furthermore, treatment of normal keratinocytes with L. pumila extract attenuated UVB-induced MMP-9 expression. These results collectively suggest L. pumila extract has tremendous potential as an anti-photoaging cosmetic ingredient.