Home About us Contact
|
Home About us Contact | ||
|
|
||
Mitochondrial Fraction (mitochondrial + fraction)
Selected AbstractsInhibition of creatine kinase activity by 3-butyl-1-phenyl-2-(phenyltelluro)oct-en-1-one in the cerebral cortex and cerebellum of young ratsJOURNAL OF APPLIED TOXICOLOGY, Issue 6 2010Rodrigo Binkowski de Andrade Abstract In the present study, we investigated the potential in vitro toxicity of the tellurium compound 3-butyl-1-phenyl-2-(phenyltelluro)oct-en-1-one on creatine kinase activity in cerebral cortex and cerebellum of 30-day-old Wistar rats. First, enriched mitochondrial and cytosolic fractions from the two tissues were pre-incubated for 30,min in the presence or absence of 1, 5 or 20,µm of organotellurium and the creatine kinase activity was measured. The organochalcogen reduced creatine kinase activity in a concentration-dependent pattern in the two tissues studied. Furthermore, the enzyme activity was performed after pre-incubation for 30, 60 or 90,min in the presence of 5,µm of the organotellurium. The compound inhibited creatine kinase activity in a time-dependent way in the enriched mitochondrial fraction of both tissues, but not in the cytosolic fraction, indicating different mechanisms for the organochalcogen in the mitochondrial and in the cytosolic creatine kinase. Pre-incubation of tellurium compound with reduced glutathione suggests that creatine kinase activity inhibition might be caused by direct interaction with thiol groups or by oxidative stress. Our findings suggest that creatine kinase inhibition may be one of the mechanisms by which this organotellurium could cause toxicity to the rat brain. Copyright © 2010 John Wiley & Sons, Ltd. [source] (,)Epigallocatechingallate protects the mitochondria against the deleterious effects of lipids, calcium and adenosine triphosphate in isoproterenol induced myocardial infarcted male Wistar ratsJOURNAL OF APPLIED TOXICOLOGY, Issue 8 2008P. T. Devika Abstract The present study was undertaken to evaluate the protective effect of (,)epigallocatechin gallate (EGCG) on mitochondrial lipids, lipid peroxides, Na+/K+ ATPase, calcium and adenosine triphosphate in isoproterenol (ISO) induced myocardial infarction in male Wistar rats. Rats were pretreated with EGCG (30 mg kg,1 body weight) orally using an intragastric tube daily for a period of 21 days. After that, ISO (100 mg kg,1 body weight) was subcutaneously injected to rats at intervals of 24 h for two days. ISO induced rats showed significant increase in the levels of cholesterol, triglycerides and free fatty acids with subsequent decrease in the levels of phospholipids in mitochondrial fraction of the heart. ISO induction also caused significant increase in lipid peroxidation products (thiobarbituric acid reactive substances and lipid hydroperoxides) and significant decrease in the activity of Na+/K+ ATPase in mitochondrial fraction of the heart. A significant increase in the levels of calcium and significant decrease in the levels of adenosine triphosphate were observed in ISO-induced mitochondrial heart. Prior treatment with EGCG (30 mg kg,1) significantly protected these alterations and maintained normal mitochondrial function. Thus, this study confirmed the protective effect of EGCG on mitochondria in experimentally induced cardiotoxicity in Wistar rats. Copyright © 2008 John Wiley & Sons, Ltd. [source] Molecular characterization of mitocalcin, a novel mitochondrial Ca2+ -binding protein with EF-hand and coiled-coil domainsJOURNAL OF NEUROCHEMISTRY, Issue 1 2006Mitsutoshi Tominaga Abstract Here we have identified and characterized a novel mitochondrial Ca2+ -binding protein, mitocalcin. Western blot analysis demonstrated that mitocalcin was widely expressed in mouse tissues. The expression in brain was increased during post-natal to adult development. Further analyses were carried out in newly established neural cell lines. The protein was expressed specifically in neurons but not in glial cells. Double-labeling studies revealed that mitocalcin was colocalized with mitochondria in neurons differentiated from 2Y-3t cells. In addition, mitocalcin was enriched in the mitochondrial fraction purified from the cells. Immunohistochemical studies on mouse cerebellum revealed that the expression pattern of mitocalcin in glomeruli of the internal granular and molecular layers was well overlapped by the distribution pattern of mitochondria. Immunogold electron microscopy showed that mitocalcin was associated with mitochondrial inner membrane. Overexpression of mitocalcin in 2Y-3t cells resulted in neurite extension. Inhibition of the expression in 2Y-3t cells caused suppression of neurite outgrowth and then cell death. These findings suggest that mitocalcin may play roles in neuronal differentiation and function through the control of mitochondrial function. [source] Oxidative modification of mitochondrial proteins and cell death in Parkinson's diseaseJOURNAL OF NEUROCHEMISTRY, Issue 2002W. Maruyama Oxidative stress is one of the cell death mechanisms in neurodegenerative disorders, such as Parkinson's disease (PD) and Alzheimer's disease. Most of reactive oxygen species (ROS) generate in mitochondria through oxidative phosphorylation, and a part of them are not scavenged by antioxidative system and react with bioactive molecules. Recently, alpha-synuclein containing nitrotyrosine, a marker for oxidative modification by peroxynitrite, was identified in Lewy body. In addition, inhibitors of mitochondrial respiratory chain were reported to induce formation of Lewy body-like inclusion in vivo and in vitro. In this paper it was examined whether ROS and reactive nitrogen species (RNS) generated in mitochondria oxidize mitochondrial respiratory enzymes and induce the formation of inclusion body and cell death in PD. Human neuroblastoma SH-SY5Y cells were treated with a peroxynitrite donor, SIN-1, or an inhibitor of complex I, rotenone. After the treatment, proteins modified with toxic aldehydes, 4-hydroxynonenal and acrolein, and containing nitrotyrosine were analyzed by immunoblotting. Particularly in mitochondrial fraction, the oxidized protein was characterized by two-dimensional immunoblotting. Most of the oxidized proteins were detected in subunits proteins of complex I. These results indicate that mitochondrial complex I is a main target of oxidative stress in dopamine neurons and its dysfunction may be involved in the death mechanism in neurodegenerative disorders. [source] Protection against kainate neurotoxicity by ginsenosides: Attenuation of convulsive behavior, mitochondrial dysfunction, and oxidative stressJOURNAL OF NEUROSCIENCE RESEARCH, Issue 3 2009Eun-Joo Shin Abstract We previously demonstrated that kainic acid (KA)-mediated mitochondrial oxidative stress contributed to hippocampal degeneration and that ginsenosides attenuated KA-induced neurotoxicity and neuronal degeneration. Here, we examined whether ginsenosides affected KA-induced mitochondrial dysfunction and oxidative stress in the rat hippocampus. Treatment with ginsenosides attenuated KA-induced convulsive behavior dose-dependently. KA treatment increased lipid peroxidation and protein oxidation and decreased the reduced glutathione/oxidized glutathione (GSH/GSSG) ratio to a greater degree in the mitochondrial fraction than in the hippocampal homogenate. KA treatment resulted in decreased Mn-superoxide dismutase expression anddiminished the mitochondrial membrane potential. Furthermore, KA treatment increased intramitochondrial Ca2+ and promoted ultrastructural degeneration in hippocampal mitochondria. Treatment with ginsenosides dose-dependently attenuated convulsive behavior and the KA-induced mitochondrial effects. Protection appeared to be more evident in mitochondria than in tissue homogenates. Collectively, the results suggest that ginsenosides prevent KA-induced neurotoxicity by attenuating mitochondrial oxidative stress and mitochondrial dysfunction. © 2008 Wiley-Liss, Inc. [source] Pro-inflammatory cytokine production from normal human fibroblasts is induced by Tannerella forsythia detaching factorJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2008N. Tomi Background and Objective:,Tannerella forsythia is a periodontal pathogen. Recently, we have reported that the cytopathic component of T. forsythia contains two distinct factors. One arrests the cell cycle at the G2 phase and the other, named forsythia detaching factor, detaches adhesion-dependent immortalized human cells. In this study, we investigated the biological function of forsythia detaching factor using human normal fibroblasts. Material and Methods:, A recombinant forsythia detaching factor, reported previously, was used. TIG-3 cells, cultured in the absence or presence of forsythia detaching factor, were lysed and the supernatant was analyzed by western blotting with polyclonal forsythia detaching factor antibodies. The cells were subsequently fractionated to isolate the cytoplasmic, mitochondrial and remaining fractions. In order to measure the activity of mitochondria using nicotinamide adenine dinucleotide-linked reductase, the water-soluble tetrazolium method was used. The mitochondrial oxidative membrane potential was estimated by measuring the oxidization-dependent fluorogenic conversion of dihydrotetramethylrosamine using flow cytometry. The concentration of interleukin-8 in the culture supernatant was assayed using a Human IL-8 ELISA kit. Results:, Forsythia detaching factor-treated cells detached from the substratum and aggregated from 3 to 24 h. Then, the detached cells resumed adhesion and proliferated after 48 h. The western blot analysis revealed that most forsythia detaching factor trans -located into the mitochondrial fraction. Forsythia detaching factor suppressed the nicotinamide adenine dinucleotide-linked reductase activity in a dose-dependent manner and consequently increased the mitochondrial oxidative membrane potential. The production of interleukin-8 was reinforced in forsythia detaching factor-treated cells at 72 h through an increase of the mitochondrial oxidative membrane potential. Conclusion:, The forsythia detaching factor might be involved in the virulence of T. forsythia through induction of the pro-inflammatory cytokine interleukin-8. [source] SIRT1 regulation of apoptosis of human chondrocytesARTHRITIS & RHEUMATISM, Issue 9 2009Koji Takayama Objective SIRT1 is known to inhibit apoptosis and to promote survival of various types of cells. However, the roles of SIRT1 in apoptosis of human chondrocytes have never been reported. We undertook this study to investigate the relationship of SIRT1 to apoptosis of human chondrocytes, which is a characteristic feature of osteoarthritis (OA). Methods The expression of SIRT1 in human chondrocytes was examined by reverse transcription,polymerase chain reaction, immunoblotting, and immunohistology of human cartilage samples. The expression of SIRT1 under catabolic, mechanical, and nutritional stresses was investigated by immunoblotting. To examine the effect of SIRT1 on apoptosis, SIRT1 was inhibited by small interfering RNA (siRNA) and activated by resveratrol during nitric oxide (NO),induced apoptosis. TUNEL staining and immunoblotting of cleaved poly(ADP-ribose) polymerase (PARP) were performed to detect apoptosis. To examine the mechanisms of apoptosis, we used immunoblotting to determine the levels of cleaved caspases and mitochondria-related apoptotic signaling proteins, Bax and Bcl-2, in the mitochondrial fraction. Results SIRT1 expression was confirmed in human chondrocytes and human cartilage samples. All catabolic, mechanical, and nutritional stresses inhibited SIRT1 expression. SIRT1 inhibition by siRNA for SIRT1 increased the percentage of TUNEL-positive cells and increased the amounts of cleaved PARP and cleaved caspases 3 and 9 induced by NO. In contrast, treatment with resveratrol decreased the percentage of TUNEL-positive cells and decreased the amounts of cleaved PARP and cleaved caspases 3 and 9 induced by NO. Furthermore, in the mitochondrial fraction, SIRT1 inhibition by siRNA for SIRT1 increased the amount of Bax but reduced the amount of Bcl-2, while resveratrol reduced the amount of Bax but increased the amount of Bcl-2. Conclusion These results indicate that SIRT1 regulates apoptosis in human chondrocytes through the modulation of mitochondria-related apoptotic signals. Further research on SIRT1 might contribute to resolving the pathogenesis of OA. [source] Structural and functional organization of Complex I in the mitochondrial respiratory chainBIOFACTORS, Issue 1-4 2003Cristina Bianchi Abstract Metabolic flux control analysis of NADH oxidation in bovine heart submitochondrial particles revealed high flux control coefficients for both Complex I and Complex III, suggesting that the two enzymes are functionally associated as a single enzyme, with channelling of the common substrate, Coenzyme Q. This is in contrast with the more accepted view of a mobile diffusable Coenzyme Q pool between these enzymes. Dilution with phospholipids of a mitochondrial fraction enriched in Complexes I and III, with consequent increased theoretical distance between complexes, determines adherence to pool behavior for Coenzyme Q, but only at dilution higher than 1:5 (protein:phospholipids), whereas, at lower phospholipid content, the turnover of NADH cytochrome c reductase is higher than expected by the pool equation. [source] Diazoxide acts more as a PKC- , activator, and indirectly activates the mitochondrial KATP channel conferring cardioprotection against hypoxic injuryBRITISH JOURNAL OF PHARMACOLOGY, Issue 8 2006M-Y Kim Background and purpose: Diazoxide, a well-known opener of the mitochondrial ATP-sensitive potassium (mitoKATP) channel, has been demonstrated to exert cardioprotective effect against ischemic injury through the mitoKATP channel and protein kinase C (PKC). We aimed to clarify the role of PKC isoforms and the relationship between the PKC isoforms and the mitoKATP channel in diazoxide-induced cardioprotection. Experimental approach: In H9c2 cells and neonatal rat cardiomyocytes, PKC-, activation was examined by Western blotting and kinase assay. Flavoprotein fluorescence, mitochondrial Ca2+ and mitochondrial membrane potential were measured by confocal microscopy. Cell death was determined by TUNEL assay. Key results: Diazoxide (100 ,M) induced translocation of PKC-, from the cytosolic to the mitochondrial fraction. Specific blockade of PKC-, by either ,V1-2 or dominant negative mutant PKC-, (PKC-, KR) abolished the anti-apoptotic effect of diazoxide. Diazoxide-induced flavoprotein oxidation was inhibited by either ,V1-2 or PKC-, KR transfection. Treatment with 5-hydroxydecanoate (5-HD) did not affect translocation and activation of PKC-, induced by diazoxide. Transfection with wild type PKC-, mimicked the flavoprotein-oxidizing effect of diazoxide, and this effect was completely blocked by ,V1-2 or 5-HD. Diazoxide prevented the increase in mitochondrial Ca2+, mitochondrial depolarization and cytochrome c release induced by hypoxia and all these effects of diazoxide were blocked by ,V1-2 or 5-HD. Conclusions and Implications: Diazoxide induced isoform-specific translocation of PKC-, as an upstream signaling molecule for the mitoKATP channel, rendering cardiomyocytes resistant to hypoxic injury through inhibition of the mitochondrial death pathway. British Journal of Pharmacology (2006) 149, 1059,1070. doi:10.1038/sj.bjp.0706922 [source] Eicosapentaenoic acid and docosahexaenoic acid effects on tumour mitochondrial metabolism, acyl CoA metabolism and cell proliferationCELL BIOCHEMISTRY AND FUNCTION, Issue 2 2001Alison Colquhoun Abstract In order to investigate the effects of high-fat diets rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), Wistar rats bearing subcutaneous implants of the Walker 256 tumour were fed pelleted chow containing low DHA/EPA or high DHA/EPA. The presence of n -3 polyunsaturated fatty acids (PUFAs) led to a marked suppression (35,46%) of tumour growth over a 12 day period. Both the whole tumour homogenate and the Percoll-purified mitochondrial fraction presented significant changes in fatty acid composition. The levels of EPA increased in both n -3 dietary groups while the levels of DHA increased only in the high DHA/EPA group, in comparison with the control chow-fed group. The presence of n -3 PUFAs led to an increase in mitochondrial acyl CoA synthetase activity, but neither the cytoplasmic acyl CoA content nor the n -3 fatty acid composition of the cytoplasmic acyl CoAs was altered by the diet. The content of thiobarbituric acid-reactive substances (TBARS) was increased in the low DHA/EPA group but was unchanged in the high DHA/EPA group. In vitro studies with the Walker 256 cell line showed a 46% decrease in cell growth in the presence of either EPA or DHA which was accompanied by a large decrease in the measured mitochondrial membrane potential. The TBARS content was increased only in the EPA-exposed cells. Cell cycle analysis identified a decrease in G0,G1 phase cells and an increase in G2,M phase cells and apoptotic cells, for both EPA and DHA-exposed cells. The data show that the presence of n -3 PUFAs in the diet is able to significantly after the growth rate of the Walker 256 tumour. The involvement of changes in mitochondrial membrane composition and membrane potential have been indicated for both EPA and DHA, while changes in lipid peroxidation have been identified in the presence of EPA but not of DHA. Copyright © 2001 John Wiley & Sons, Ltd. [source] 3241: Effect of glutaredoxin 2 gene knockout on lens epithelial cells against oxidative stressACTA OPHTHALMOLOGICA, Issue 2010M LOU Purpose The mitochondrial glutaredoxin 2 (Grx2) is known to possess both dethiolase and peroxidase activities, and has shown an ability to protect cells from oxidative stress-induced apoptosis in the human lens epithelial cells. In this study, we further studied the function of Grx2 by using Grx2 knockout mouse lens epithelial (MLE) cells as a model. Methods Primary culture of MLE cells was established from the lenses of wild-type (WT) and Grx2-knockout (Grx2 KO) mice. Cells were probed for ,A-crystallin and Grx2 by Western blot analysis while cell viability was examined by WST-8 assay. Glutathione (GSH) level, Grx2 and Complex I activities, and lactate dehydrogenase (LDH) release were determined by spectrophotometric assays. Reactive oxygen species was detected using DCF-DA fluorescein with a cell sorter. Apoptosis was quantified by flow cytometry. Results Both primary cell cultures were confirmed to be lens epithelial cells by the presence of ,A-crystallin. Western blotting showed normal expression of Grx2 in WT cells but absent in Grx2 KO cells. Both cell types showed similar morphology and growth rate with same level of GSH pool and complex 1 activity in the mitochondrial fraction. However, KO cells were more sensitive to oxidative stress (100 ,M H2O2 for 6 h) and exhibited lower cell viability and more LDH leakage in comparison with the WT cells. In addition, knockdown of Grx2 weakened the cell's ability to detoxify H2O2 and enhanced the H2O2-induced inactivation of complex I in the electron transport chain. Conclusion Grx2 can protect MLE cells from H2O2-induced cell injury, and the mechanism of this protection is likely associated with its ability to detoxify H2O2 and its preservation of complex I activity in the mitochondria. [source] Malonyl-CoA decarboxylase (MCD) is differentially regulated in subcellular compartments by 5,AMP-activated protein kinase (AMPK)FEBS JOURNAL, Issue 13 2004AMPK by adenoviral gene transfer technique, Studies using H9c2 cells overexpressing MCD Malonyl-CoA, a potent inhibitor of carnitine pamitoyl transferase-I (CPT-I), plays a pivotal role in fuel selection in cardiac muscle. Malonyl-CoA decarboxylase (MCD) catalyzes the degradation of malonyl-CoA, removes a potent allosteric inhibition on CPT-I and thereby increases fatty acid oxidation in the heart. Although MCD has several Ser/Thr phosphorylation sites, whether it is regulated by AMP-activated protein kinase (AMPK) has been controversial. We therefore overexpressed MCD (Ad.MCD) and constitutively active AMPK (Ad.CA-AMPK) in H9c2 cells, using an adenoviral gene delivery approach in order to examine if MCD is regulated by AMPK. Cells infected with Ad.CA-AMPK demonstrated a fourfold increase in AMPK activity as compared with control cells expressing green fluorescent protein (Ad.GFP). MCD activity increased 40- to 50-fold in Ad.MCD + Ad.GFP cells when compared with Ad.GFP control. Co-expressing AMPK with MCD further augmented MCD expression and activity in Ad.MCD + Ad.CA-AMPK cells compared with the Ad.MCD + Ad.GFP control. Subcellular fractionation further revealed that 54.7 kDa isoform of MCD expression was significantly higher in cytosolic fractions of Ad.MCD + Ad.CA-AMPK cells than of the Ad.MCD +Ad.GFP control. However, the MCD activities in cytosolic fractions were not different between the two groups. Interestingly, in the mitochondrial fractions, MCD activity significantly increased in Ad.MCD + Ad.CA-AMPK cells when compared with Ad.MCD + Ad.GFP cells. Using phosphoserine and phosphothreonine antibodies, no phosphorylation of MCD by AMPK was observed. The increase in MCD activity in mitochondria-rich fractions of Ad.MCD + Ad.CA-AMPK cells was accompanied by an increase in the level of the 50.7 kDa isoform of MCD protein in the mitochondria. This differential regulation of MCD expression and activity in the mitochondria by AMPK may potentially regulate malonyl-CoA levels at sites nearby CPT-I on the mitochondria. [source] Biochemical and ultrastructural alterations in the rat ventral prostate due to repetitive alcohol drinkingJOURNAL OF APPLIED TOXICOLOGY, Issue 4 2007M. I. Díaz Gómez Abstract Previous studies showed that cytosolic and microsomal fractions from rat ventral prostate are able to biotransform ethanol to acetaldehyde and 1-hydroxyethyl radicals via xanthine oxidase and a non P450 dependent pathway respectively. Sprague Dawley male rats were fed with a Lieber and De Carli diet containing ethanol for 28 days and compared against adequately pair-fed controls. Prostate microsomal fractions were found to exhibit CYP2E1-mediated hydroxylase activity significantly lower than in the liver and it was induced by repetitive ethanol drinking. Ethanol drinking led to an increased susceptibility of prostatic lipids to oxidation, as detected by t-butylhydroperoxide-promoted chemiluminiscence emission and increased levels of lipid hydroperoxides (xylenol orange method). Ultrastructural alterations in the epithelial cells were observed. They consisted of marked condensation of chromatin around the perinuclear membrane, moderate dilatation of the endoplasmic reticulum and an increased number of epithelial cells undergoing apoptosis. The prostatic alcohol dehydrogenase activity of the stock rats was 4.84 times lower than that in the liver and aldehyde dehydrogenase activity in their microsomal, cytosolic and mitochondrial fractions was either not detectable or significantly less intense than in the liver. A single dose of ethanol led to significant acetaldehyde accumulation in the prostate. The results suggest that acetaldehyde accumulation in prostate tissue might result from both acetaldehyde produced in situ but also because of its low aldehyde dehydrogenase activity and its poor ability to metabolize acetaldehyde arriving via the blood. Acetaldehyde, 1-hydroxyethyl radical and the oxidative stress produced may lead to epithelial cell injury. Copyright © 2007 John Wiley & Sons, Ltd. [source] Heptachlor and o-p,DDT effects on protein kinase activities associated with human placenta particulate fractionsJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 3 2009Gladis Magnarelli Abstract Organochlorine pesticides have been detected in placenta. The ability of heptachlor (HC) and 1,1,1-tricholoro-2-(2-chlorophenyl)-2-4-chlorophenyl)ethane (o-p,DDT) to interfere with protein phosphorylation was evaluated. In vitro incubations of cell-free placental villi homogenates with a concentration range 1,100 µM were performed. In particulate fractions, total serine/threonine kinase activity was increased by 10 µM HC and o-p, DDT (59% and 82%, respectively). Maximum eightfold increase was observed with 10 µM o-p, DDT on protein kinase A activity. By contrast, protein kinase C activity was reduced by 10 µM HC and o-p, DDT (40% and 52%, respectively). Endogenous substrate phosphorylation studies demonstrated that slight but significant increase in 24-kDa band labeling was produced in nuclear samples with 1, 10, and 100 µM HC and 100 µM o-p, DDT. Exposition to 100 µM HC increased 85-kDa band labeling. In mitochondrial fractions, 10 µM HC and o-p, DDT increased 24- and 65-kDa bands' labeling. These data indicate that both pesticides affect protein kinase activities in particulate fraction. Nuclear compartmentalization of these compounds, insertion in membranes, and chemical stress production may be associated to the observed effects, thus suggesting deleterious consequences in signaling pathways. © 2009 Wiley Periodicals, Inc. J Biochem Mol Toxicol 23:185,192, 2009; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20277 [source] Xenobiotic response element binding enriched in both nuclear and microsomal fractions of rat cerebellumJOURNAL OF NEUROCHEMISTRY, Issue 1 2003Nobuyuki Kuramoto Abstract Xenobiotic response element (XRE) is a core nucleotide sequence at the upstream of inducible target genes for the transcription factor aryl hydrocarbon receptor (AhR) that is responsible for signal transduction of exogenous environmental pollutants in eukaryotic cells. Immunoblotting analysis revealed the constitutive expression of AhR-related proteins in rat liver and brain, while specific binding of a radiolabelled probe containing XRE was detected in nuclear preparations of both liver and brain on gel retardation electrophoresis. Among discrete rat brain structures examined, cerebellum exhibited the highest XRE binding with less potent binding in hypothalamus, midbrain, medulla-oblongata, hippocampus, cerebral cortex and striatum. In contrast to liver and hippocampus, cerebellum also contained unusually higher XRE binding in microsomal fractions than that in either nuclear or mitochondrial fractions. Limited proteolysis by V8 protease did not markedly affect XRE binding in cerebellar nuclear extracts, with concomitant diminution of that in hepatic and hippocampal nuclear extracts. In primary cultured cerebellar neurons, indigo was effective in significantly increasing XRE binding only when determined immediately after sustained exposure for 120 min in the presence of high potassium chloride. These results suggest the abundance of as-yet unidentified proteins with high affinity for XRE and responsiveness to indigo in both nuclear and microsomal fractions of rat cerebellum. [source] Asiatic acid, a pentacyclic triterpene from Centella asiatica, is neuroprotective in a mouse model of focal cerebral ischemiaJOURNAL OF NEUROSCIENCE RESEARCH, Issue 11 2009Rajanikant G. Krishnamurthy Abstract Asiatic acid, a triterpenoid derivative from Centella asiatica, has shown biological effects such as antioxidant, antiinflammatory, and protection against glutamate- or ,-amyloid-induced neurotoxicity. We investigated the neuroprotective effect of asiatic acid in a mouse model of permanent cerebral ischemia. Various doses of asiatic acid (30, 75, or 165 mg/kg) were administered orally at 1 hr pre- and 3, 10, and 20 hr postischemia, and infarct volume and behavioral deficits were evaluated at day 1 or 7 postischemia. IgG (blood,brain barrier integrity) and cytochrome c (apoptosis) immunostaining was carried out at 24 hr postischemia. The effect of asiatic acid on stress-induced cytochrome c release was examined in isolated mitochondrial fractions. Furthermore, its effects on cell viability and mitochondrial membrane potential were studied in HT-22 cells exposed to oxygen-glucose deprivation. Asiatic acid significantly reduced the infarct volume by 60% at day 1 and by 26% at day 7 postischemia and improved neurological outcome at 24 hr postischemia. Our studies also showed that the neuroprotective properties of asiatic acid might be mediated in part through decreased blood,brain barrier permeability and reduction in mitochondrial injury. The present study suggests that asiatic acid may be useful in the treatment of cerebral ischemia. © 2009 Wiley-Liss, Inc. [source] | ||||||||||||||||