Protective Effects of a Diarylheptanoid from Curcuma comosa Against Hydrogen Peroxide-Induced Astroglial Cell Death.

Oxidative stress is one of the major mechanisms causing neuronal and astroglial cell death in various neurological disorders such as Alzheimer's disease, Parkinson's disease, and brain ischemia. Two diarylheptanoids, (3R)-1,7-diphenyl-(4E,6E)-4,6-heptadien-3-ol (ASPP 049) and (3S)-7-(3,4-dihydroxyphenyl)-1-phenyl-(1E)-1-hepten-3-ol (ASPP 092), isolated from Curcuma comosa were investigated for cytoprotective effects on C6 astroglial cells using hydrogen peroxide (H2O2) exposure as a model of oxidative stress. ASPP 092 demonstrated free radical scavenging activity comparable to that of vitamin C, while ASPP 049 showed no antioxidant activity. Treatment with H2O2 at 400 µM for 12 h caused 79 % C6 astroglial cell death which was significantly reduced to 37 % by pretreatment with ASPP 092 (5 µM). In addition, ASPP 092 attenuated the increase in reactive oxygen species production and the decrease in total glutathione level induced by H2O2. The mechanism of ASPP 092 protection against H2O2-induced apoptotic signaling appeared to involve prevention of increase in the level of phosphorylated p53 and the Bax/Bcl-2 ratio as well as cleaved caspase-3. These findings provide new evidence that the diarylheptanoid ASPP 092 from C. comosa possesses antiapoptotic properties and could be further developed as a potential treatment for oxidative stress-related neuronal diseases.

Autophagy inhibition by caffeine increases toxicity of methamphetamine in SH-SY5Y neuroblastoma cell line.

Methamphetamine (METH) is a highly addictive CNS stimulant that its long-term use is associated with the loss of neurons in substantia nigra and development of Parkinson's disease later in life. Common form of METH is Ya-Ba tablet, in which, large portion of caffeine is added to the mass to enhance the stimulatory effect. Previous study demonstrated that caffeine potentiates the toxic effect of METH in association with the production of reactive oxygen species and the induction of apoptosis. Since METH causes induction of autophagy, the question was raised whether this pathway participates in the potentiating effect of caffeine on METH neurotoxicity. We used SH-SY5Y, a neuroblastoma cell line, as an in vitro model to study the effect of METH and caffeine. Co-treatment of non-toxic concentrations of METH, at 0.5 mM, and caffeine, at 1 mM, caused reduction of the cell viability. Reduction of the cell viability was associated with attenuation of autophagy, demonstrated by reduction of LC3-II levels and the number of autophagosome puncta, together with increase of caspase-3 activation. Similar effect was produced by treatment with autophagy inhibitors, 3-MA and wortmanin. Our results suggested that caffeine potentiates METH toxicity through inhibition of autophagy and that autophagy serves as a protective mechanism. In conclusion, we proposed the augmented hazard associated with caffeine and METH combination in Ya-Ba abusers.

Curcumin I mediates neuroprotective effect through attenuation of quinoprotein formation, p-p38 MAPK expression, and caspase-3 activation in 6-hydroxydopamine treated SH-SY5Y cells.

6-Hydroxydopamine (6-OHDA) selectively enters dopaminergic neurons and undergoes auto-oxidation resulting in the generation of reactive oxygen species and dopamine quinones, subsequently leading to apoptosis. This mechanism mimics the pathogenesis of Parkinson's disease and has been used to induce experimental Parkinsonism in both in vitro and in vivo systems. In this study, we investigated the effects of curcumin I (diferuloylmethane) purified from Curcuma longa on quinoprotein production, phosphorylation of p38 MAPK (p-p38), and caspase-3 activation in 6-OHDA-treated SH-SY5Y dopaminergic cells. Pretreatment of SH-SY5Y with curcumin I at concentrations of 1, 5, 10, and 20 µM, significantly decreased the formation of quinoprotein and reduced the levels of p-p38 and cleaved caspase-3 in a dose-dependent manner. Moreover, the levels of the dopaminergic neuron marker, phospho-tyrosine hydroxylase (p-TH), were also dose-dependently increased upon treatment with curcumin I. Our results clearly demonstrated that curcumin I protects neurons against oxidative damage, as shown by attenuation of p-p38 expression, caspase-3-activation, and toxic quinoprotein formation, together with the restoration of p-TH levels. This study provides evidence for the therapeutic potential of curcumin I in the chemoprevention of oxidative stress-related neurodegeneration.

Phytoestrogens mediated anti-inflammatory effect through suppression of IRF-1 and pSTAT1 expressions in lipopolysaccharide-activated microglia.

Microglial activation has been implicated in various neurological disorders, including Alzheimer's disease, Parkinson's disease, multiple sclerosis, and HIV encephalopathy. Phytoestrogens have been shown to be neuroprotective in neurotoxicity models; however, their effect on microglia has not been well established. In the current study, we report that the soy phytoestrogens, genistein, daidzein, and coumestrol, decreased nitric oxide (NO) production induced by lipopolysaccharide (LPS) in the rat microglial cell line (HAPI). The levels of inducible NO synthase (iNOS) mRNA and protein expression were also reduced. Transcription factors known to govern iNOS expression including interferon regulatory factor-1 (IRF-1) and phosphorylated STAT1 were down regulated. These observations explain, at least in part, the inhibitory effect of phytoestrogens on NO production. The levels of monocyte chemoattractant protein-1 and interleukin-6 mRNA, proinflammatory chemokine and cytokine associated with various neurological disorders, were also reduced following LPS stimulation when HAPI cells were pretreated with phytoestrogens. Hence, genistein, daidzein, and coumestrol could serve as anti-inflammatory agents and may have beneficial effects in the treatment of neurodegenerative diseases.

Vitamin E supplement improves erythrocyte membrane fluidity of thalassemia: an ESR spin labeling study.

BACKGROUND: Beta-thalassemia/Hemoglobin E (beta-thal/Hb E) is prevalent in Thailand. The imbalance of globin chains in red blood cells is the primary cause of this anemic disease. The excess alpha-globin in beta-thal/Hb E causes typical damage(s) to membrane of erythroblasts and erythrocytes. By using three paramagnetic labeled compounds (5-, 12-, and 16-spin labeled stearic acids, SLS), the changes of the molecular motion in the lipid bilayer of thalassemic RBCs that have structural modification can be detected. OBJECTIVE: to investigate erythrocyte membrane fluidity and the effect of vitamin E treatment in beta-thalassemia/Hemoglobin E patients by using spin labeling techniques. MATERIAL AND METHOD: The erythrocyte membrane fluidity was investigated in nine splenectomized and five non-splenectomized beta-thalassemia/hemoglobin E (beta-thal/Hb E) patients using EPR spin labeling techniques. To determine the effect of vitamin E on erythrocyte membrane fluidity, only the splenectomized patients were enrolled. Patients were divided into two groups. The first group received 350 mg vitamin E daily for a period of 1 month (n = 5) and the second group received placebo for an equal period (n = 4). Three paramagnetic fatty acid, 5-, 12-, and 16-doxyl stearic acids, (5-, 12- and 16-DS) were used to label phospholipids layer near both the surface (5-DS) and the deeper hydrophobic region of membrane (12-and 16-DS). Lipid peroxidation (TBARs) was measured using a colorimetric method. Vitamin E was measured with high performance liquid chromatography (HPLC). RESULTS: Significantly higher values of erythrocyte membrane fluidity were revealed with 12-, 16-DS in splenectomized patients, as compared with non-splenectomized patients and normal subjects. In 3-thal/Hb E patients, fluidity values, both outer hyperfine splitting (2T(//)) and order parameter (S) of 12-DS showed inverse correlation with serum TBARs. There was no significant difference between the fluidity values measured with 5-DS. After vitamin E supplementation, the erythrocyte membrane fluidity was decreased in almost all patients. In contrast to the vitamin E supplementation group, increased erythrocyte membrane fluidity was demonstrated in the placebo group. Vitamin E supplementation also had effect on other clinical parameters such as increased plasma vitamin E, decreased serum TBARs and no change in hemoglobin. CONCLUSION: The present results suggested the abnormal motion of lipid in the deeper phospholipids region of membrane. In addition, vitamin E supplementation may have a role in the prevention of erythrocyte membrane damage of these patients.

Caffeine potentiates methamphetamine-induced toxicity both in vitro and in vivo.

Ya-Ba, a combination of the two potent psychostimulants methamphetamine (METH) and caffeine (CAF), is commonly used by drug abusers in Thailand and neighboring countries. While the neurotoxic effects of METH are well documented, the toxicity of this combination is mostly unknown. This study aimed to elucidate the effects of this particular drug combination using both in vitro and in vivo models. We found that combined treatment of METH and CAF at individually non-toxic concentrations significantly decreased viability of human neuroblastoma SK-N-SH cells. The reduction in cell survival was accompanied by an increase in reactive oxygen species (ROS) formation and the Bax/Bcl-2 ratio. In vivo data showed that combined administration of METH and CAF increased the mortality rate of rats, with an increase in the level of thiobarbituric acid reactive substances (TBARS), the indicator of oxidative stress, in striatal tissues. The results indicate that caffeine potentiates the toxic effects of methamphetamine, possibly via a mechanism involving an increase in dopamine release and excess ROS generation.

Effects of Curcuma comosa on the expression of atherosclerosis-related cytokine genes in rabbits fed a high-cholesterol diet.

AIM OF THE STUDY: Curcuma comosa has been known to have potential use in cardiovascular diseases, but its immunoregulatory role in atherosclerosis development and liver toxicity has not been well studied. We therefore investigated the effects of Curcuma comosa on the expression of atherosclerosis-related cytokine genes in rabbits fed a high-cholesterol diet. MATERIALS AND METHODS: Twelve male New Zealand White rabbits were treated with 1.0% cholesterol for one month and were subsequently treated with 0.5% cholesterol either alone, or in combination with 5mg/kg/day of simvastatin or with 400mg/kg/day of Curcuma comosa powder for three months. The expression of IL-1, MCP-1, TNF-α, IL-10, and TGF-ß in the isolated abdominal aorta and liver were determined by real-time RT-PCR. Liver toxicity was determined by hepatic enzyme activity. RESULTS: Curcuma comosa significantly decreased the expression of pro-inflammatory cytokines, leading to a stronger reduction in IL-1, MCP-1, and TNF-α expression compared to that was suppressed by simvastatin treatment. However, neither Curcuma comosa nor simvastatin affected the expression of anti-inflammation cytokines. In the liver, Curcuma comosa insignificantly decreased the expression of pro-inflammatory cytokines and significantly increased the expression of the anti-inflammatory cytokine IL-10 without altering the activity of hepatic enzymes. In contrast, simvastatin significantly increased the MCP-1 and TNF-α expressions and serum ALT level, without affecting the expression of anti-inflammatory cytokines. CONCLUSIONS: In this study, we demonstrated that Curcuma comosa exerts anti-inflammatory activity in the aorta and liver without causing liver toxicity, indicating that Curcuma comosa is a potential candidate as an alternative agent in cardiovascular disease therapy.

Curcumin I protects the dopaminergic cell line SH-SY5Y from 6-hydroxydopamine-induced neurotoxicity through attenuation of p53-mediated apoptosis.

Oxidative stress (OS) plays a pivotal role in the pathogenesis of Parkinson's disease (PD). 6-Hydroxydopamine (6-OHDA) is a neurotoxin used to induce oxidative cell death of dopaminergic neurons in experimental models of PD. Curcumin I, or diferuloylmethane is a pure compound isolated from Curcuma longa Linn. that has been reported to have neuroprotective properties. The precise mechanism, however, remains unclear. This study aims to elucidate the mechanisms by which curcumin I exerts its effects, using 6-OHDA-induced neurotoxicity in the human dopaminergic cell line SH-SY5Y. In our experiments, pretreatment with curcumin I improved cell viability, and significantly reduced reactive oxygen species (ROS). Further investigations revealed a reduction of p53 phosphorylation and decrease of the Bax/Bcl-2 ratio, as measured by mRNA expression and protein level. Taken together, these findings indicate that curcumin I protects dopaminergic neurons from 6-OHDA-induced toxicity via the reduction of ROS production, and subsequent attenuation of p53 phosphorylation and reduction of the Bax/Bcl-2 ratio.

Oxidative modification and poor protective activity of HDL on LDL oxidation in thalassemia.

Oxidative modification of low-density lipoprotein (LDL) has been reported in thalassemia, which is a consequence of oxidative stress. However, the levels of oxidized high-density lipoprotein (HDL) in thalassemia have not been evaluated and it is unclear whether HDL oxidation may be linked to LDL oxidation. In this study, the levels of total cholesterol, iron, protein, conjugated diene (CD), lipid hydroperoxide (LOOH), and thiobarbituric acid reactive substances (TBARs) were determined in HDL from healthy volunteers and patients with beta-thalassemia intermedia with hemoglobin E (beta-thal/Hb E). The protective activity of thalassemic HDL on LDL oxidation was also investigated. The iron content of HDL(2) and HDL(3) from beta-thal/HbE patients was higher while the cholesterol content was lower than those in healthy volunteers. Thalassemic HDL(2) and HDL(3) had increased levels of lipid peroxidation markers i.e., conjugated diene, LOOH, and TBARs. Thalassemic HDL had lower peroxidase activity than control HDL and was unable to protect LDL from oxidation induced by CuSO(4). Our findings highlight the oxidative modification and poor protective activity of thalassemic HDL on LDL oxidation which may contribute to cardiovascular complications in thalassemia.

Increased blood oxidative stress in amphetamine users.

Amphetamine derivatives have been shown to be a potential brain neurotoxin based on the production of free radicals that occurs after administration. The purpose of this study was to examine the lipid peroxidation and antioxidant enzymes in the blood of amphetamine users. The plasma lipid peroxidation was determined and reported as thiobarbituric acid reactive substance and was significantly increased (+21%), whereas the activities of the erythrocyte antioxidant enzymes glutathione peroxidase, catalase, and superoxide dismutase were significantly decreased (-32%, -14% and -31%, respectively) in amphetamine users. These results implicated the potential role of oxidative stress in amphetamine-induced neurotoxicity.

Transcriptional regulation of iNOS and COX-2 by a novel compound from Curcuma comosa in lipopolysaccharide-induced microglial activation.

Overproduction of pro-inflammatory mediators resulting from chronic activation of microglia has been implicated in many neurodegenerative disorders, such as Parkinson's disease and Alzheimer's disease. In this study, we investigated the effects of (3R) 1,7-diphenyl-(4E,6E)-4,6-heptadien-3-ol, or compound 049 on the production of pro-inflammatory mediators in lipopolysaccharide (LPS)-treated microglia. Compound 049 is a pure compound fractionated from the hexane extract of Curcuma comosa, an indigenous plant of Thailand traditionally used as an anti-inflammatory agent for the treatment of uterine inflammation. It was found that pretreatment of the highly aggressively proliferating immortalized (HAPI), rat microglial cell line, with compound 049, at the concentrations of 0.1, 0.5 and 1microM significantly decreased LPS-induced NO and PGE(2) production in a concentration-dependent manner. Parallel to the decreases in NO and PGE(2) production was a reduction in the expression of inducible NO synthase (iNOS) and cyclooxygenase 2 (COX-2) as measured by mRNA and protein levels. These results indicate that compound 049 possesses an anti-inflammatory activity and may have a therapeutic potential for the treatment of neurodegenerative diseases related to microglial activation.

Modification of platelet shape change parameter by oxidized lipoprotein from beta-thalassemia/Hemoglobin E.

BACKGROUND: Beta-thalassemia/Hemoglobin E (beta-thal/Hb E) is a congenital hemolytic anemia that is prevalent in Thailand Pulmonary arterial occlusion is the cause of morbidity and mortality in these patients. Abnormality of platelets has been implicated as pathogenesis of this condition. However the blood-borne factors that induce platelet activation are not identified Recently, oxidized low-density lipoproteins (ox-LDLs) had been identified in thalassemic blood. OBJECTIVE: Identify whether oxidized LDL is the blood bone factor that induce platelet activation in beta-thal/Hb E patients. MATERIAL AND METHOD: Platelet activation was measured by monitoring platelet shape change parameter using plasma-free human platelets. The shape change parameter was monitored following exposure to normal LDL, oxidized LDL, and thalassemic LDL. RESULTS: Oxidized LDL, but not the native LDL and thalassemic LDL, showed platelet activation activity. Oxidation of thalassemic LDL with copper give rise to oxidized LDL with platelet activating activity. However less copper was needed by LDL from splenectomized beta-thal/Hb E patients than those from nonsplencectomized beta-thal/Hb E patients. CONCLUSION: LDL from splenectomized beta-thal/Hb E patients is more susceptible for oxidation and gives rise to oxidized-LDL that plays an important role in thrombosis event in these patients.

Modification of platelet shape change parameter by modified phospholipids in oxidized LDL.

BACKGROUND: It is believed that the oxidatively modified lipoproteins play a critical role in activating platelets and is a contributing factor in the etiology of a number of cardiovascular-related diseases. OBJECTIVE: Identify the active component(s) of oxidized LDL that initiated shape change in plasma-free human platelets prepared by a gel filtration method Material and Method: Shape change parameter of platelets was monitored following exposure platelets to LDL, copper sulfate-oxidized LDL, and different types of lipids extracted of the corresponding LDL. RESULTS: Oxidized LDL, but not native LDL, increased the shape-change parameter in a concentration-dependent manner Specifically, phosphatidyl serine from oxidized LDL was responsible for this effect. CONCLUSION: Oxidized phospholipids generated during the oxidative modification of LDL are likely to be the active components responsible for changes in platelet function.

Reactive oxygen species production and MAPK activation are implicated in tetrahydrobiopterin-induced SH-SY5Y cell death.

Tetrahydrobiopterin (BH4), an obligatory cofactor for dopamine (DA) synthesis, has been shown to produce reactive oxygen species (ROS) upon its autoxidation and induce selective dopaminergic cell death in many in vivo and in vitro models of Parkinson's disease (PD). The precise molecular mechanisms underlying neuronal death upon BH4 exposure, however, have not yet been well elucidated. The present study aims to examine the intracellular ROS production and the signal transduction pathways underlying the toxic effects of BH4 on human dopaminergic SH-SY5Y cells. The results show that BH4 treatment at concentrations ranging from 50microM to 400microM induces neuronal death in a dose-dependent manner. In concomitant with the elevation of intracellular ROS formation, BH4-induced activation of MAPK, p38 and ERK1/2 in SH-SY5Y cells is attenuated by pretreatment with MAPK inhibitors, SB203580 or PD98059. These data indicate that MAPK activation and oxidative stress are involved in BH4-induced dopaminergic cell death, possibly through the autoxidation of BH4 and subsequent ROS production.

The signaling cascades of Ganoderma lucidum extracts in stimulating non-amyloidogenic protein secretion in human neuroblastoma SH-SY5Y cell lines.

Ganoderma lucidum (GL) is a medicinal mushroom that possesses various pharmacological properties which are also documented in the ancient reports where GL is praised for its effects on the promotion of health and longevity. In this study, we have investigated the effect of GL mycelia extracts on the non-amyloidogenic protein secretion (sAPPalpha) and the amyloid precursor protein (APP) expression in SH-SY5Y neuroblastoma cells. In order to characterize the signaling pathway which mediates GL-enhanced sAPPalpha secretion, we used inhibitors of nerve growth factor (NGF) signaling pathways, phosphatidylinositol 3 kinase (PI3K), phospholipase Cgamma1 (PLCgamma1), protein kinase C (PKC) and extracellular signal-regulated kinase (ERK1/2), to block GL-mediated sAPPalpha secretion as well as ERK1/2 and PKC activation by using Western blot analysis. Our results provided for the first time evidence that GL mycelia extracts increased APP expression and promoted sAPPalpha secretion. In addition, GL extracts activated ERK1/2 and PKC phosphorylation. The complex signaling cascades of PI3K and ERK may be responsible for GL-mediated sAPPalpha secretion.

Pueraria mirifica, phytoestrogen-induced change in synaptophysin expression via estrogen receptor in rat hippocampal neuron.

OBJECTIVE: To examine Pueraria mirifica (Leguminosae) containing-phytoestrogen effect on synaptic density and involvement of estrogen receptor. MATERIAL AND METHOD: The level of synaptophysin, a presynaptic vesicle protein, was measured using Western blot analysis and immunocytochemistry in hippocampal primary cell cultures at 6 days in vitro. RESULTS: P. mirifica and 17beta-estradiol (0.1 microM) treatment for 4 days, but not for 2 days, significantly increased synaptophysin immunoreactivity and level of synaptophysin. P. mirifica up to 60 microg/ml resulted in a dose related increase in the level of synaptophysin immunoreactivity. The classical estrogen receptor antagonist, ICI 182 780, significantly blocked P. mirifica-induced increase in synaptophysin. CONCLUSION: P. mirifica-containing phytoestrogen affects synaptic density by inducing synaptophysin expression via estrogen receptor.

The in vitro and ex vivo antioxidant properties, hypolipidaemic and antiatherosclerotic activities of water extract of Moringa oleifera Lam. leaves.

UNLABELLED: Moringa oleifera is used in Thai traditional medicine as cardiotonic. Recent studies demonstrated its hypocholesterolaemic effect. However, to be clinically useful, more scientific data are needed. AIM OF THE STUDY: We investigated the antioxidant, hypolipidaemic and antiatherosclerotic activities of Moringa oleifera leaf extract. MATERIALS AND METHODS: Scavenging activity of the extract on 1,1-diphenyl-2-picrylhydrazyl radicals (DPPH), and the inhibitory effect on Cu(2+)-induced low-density lipoprotein (LDL) oxidation were determined in in vitro experiment. The effects of the extract on cholesterol levels, conjugated diene (CD) and thiobarbituric acid reactive substances (TBARS) and plaque formations in cholesterol-fed rabbits were investigated. RESULTS: We found that in scavenging DPPH radicals the extract and Trolox had IC(50) of 78.15+/-0.92 and 2.14+/-0.12microg/ml, respectively. The extract significantly (P<0.05) prolonged the lag-time of CD formation and inhibited TBARS formation in both in vitro and ex vivo experiments in a dose-dependent manner. In hypercholesterol-fed rabbits, at 12 weeks of treatment, it significantly (P<0.05) lowered the cholesterol levels and reduced the atherosclerotic plaque formation to about 50 and 86%, respectively. These effects were at degrees comparable to those of simvastatin. CONCLUSIONS: The results indicate that this plant possesses antioxidant, hypolipidaemic and antiatherosclerotic activities and has therapeutic potential for the prevention of cardiovascular diseases.

A pharmacokinetic study of paracetamol in Thai beta-thalassemia/HbE patients.

BACKGROUND: Thalassemia may alter the pharmacokinetics of several drugs in thalassemic patients. Paracetamol is a commonly used analgesic and antipyretic drug which is extensively metabolized in the liver via glucuronidation. The aim of this study was to compare the pharmacokinetics of paracetamol (PCM) and its metabolites [paracetamol glucuronide (PCM-G), paracetamol sulfate (PCM-S), and paracetamol cysteine (PCM-C)] in 16 patients with 16 normal subjects. METHOD: Following an overnight fast, a single dose of paracetamol (1,000 mg of Tylenol(R)) was given and blood samples were obtained at predose, 0.5, 1, 1.5, 2, 3, 4, 5, 7, and 9 h after dosing for determination of the plasma levels of PCM and its metabolites by high-performance liquid chromatography. RESULTS: There was no significant difference in maximum concentration of PCM between groups. However, a significantly shorter elimination half-life of PCM was observed in the thalassemic subjects (p<0.001). Total apparent clearance of PCM was significantly faster in thalassemic subjects (p<0.01) while the apparent volume of distribution of PCM did not change. The area under the concentration time curve (AUC(0->infinity)) of PCM-G and PCM-S increased in thalassemic subjects (p<0.05) whereas this parameter for PCM-C was slightly lower in the patients. The half-lives of PCM metabolites were significantly shorter (p<0.01) in thalassemic subjects. CONCLUSION: The results indicate that the elimination of PCM and its metabolites in thalassemic subjects is faster than that in normal subjects. Our pharmacokinetic data provide additional evidence that plasma PCM-G is higher in thalassemic patients with hyperbilirubinemia, which could be a casual relationship in regulating the UDP-glucuronosyltransferase expression.

Inhibitory effect of Curcuma comosa on NO production and cytokine expression in LPS-activated microglia.

Curcuma comosa is an indigenous plant of Thailand, which has been traditionally and widely used as an anti-inflammatory agent for the treatment of postpartum uterine bleeding and uterine inflammation. However, the scientific investigation on its anti-inflammatory activity has not been reported. In the present study, we investigated the anti-inflammatory effect of the extract from C. comosa on the responses in microglia stimulated with lipopolysaccharide (LPS). Pretreatment of highly aggressively proliferating immortalized (HAPI) cells, a rat microglial cell line, with the hexane extract of C. comosa rhizome at 10(-9) to 10(-5) g/ml significantly suppressed the levels of NO released from these cells. The attenuation in iNOS protein and mRNA expression was also observed suggesting an interference at transcriptional level. In addition, C. comosa extract inhibited interferon regulatory factor-1 expression which is an essential transcription factor governing the iNOS expression. Moreover, the levels of mRNA expressions of MCP-1 and IL-6 induced by LPS were also prominently decreased in the presence of C. comosa extract. These results suggest that C. comosa extract possesses a strong anti-inflammatory activity and has a potential to be developed as a therapeutic compound for diverse neurological disorders associated with inflammation.

Improvement of vascular function by chronic administration of a cyclo-oxygenase inhibitor in cholesterol-fed rabbits.

1. Atherosclerotic cardio- and cerebrovascular disease is a leading cause of mortality in Western countries. Aspirin-like drugs are widely used to prevent and treat these occlusive cardio- and cerebrovascular diseases. The beneficial effects of these drugs have been largely attributed to inhibition of platelet cyclo-oxygenase activity and thromboxane (TX) A2 production. We investigated the effect of an aspirin-like drug, namely indomethacin, on endothelial function, plaque and platelet aggregation and the formation of vasoactive substances during the development of atherosclerosis in cholesterol-fed rabbits. 2. Rabbits were fed 1% cholesterol (n = 8), 1% cholesterol plus 25 mg/day indomethacin (n = 8) or normal rabbit chow (control group; n = 8) for 12 weeks. Urinary excretion rates of 2,3-dinor-TXB2, 6-keto-prostaglandin (PG) F1alpha, 8-iso-PGF2alpha and nitrate were analysed at the beginning of dietary intervention and at 4 weekly intervals thereafter. At the end of the study period, platelet aggregation, aortic plaque formation and endothelium-dependent and -independent vascular functions of isolated aortic rings ex vivo were assessed. 3. Compared with control, in the cholesterol-fed group, urinary 2,3-dinor-TXB2, 6-keto-PGF1alpha and 8-iso-PGF2alpha excretion and platelet aggregation were significantly increased (P < 0.05), but urinary excretion of nitrate was decreased (P < 0.05). Treatment with indomethacin significantly reduced platelet aggregation, urinary 2,3-dinor-TXB2, 6-keto-PGF1alpha and 8-iso-PGF2alpha excretion (P < 0.05 vs the cholesterol-fed group) and attenuated the reduction in urinary nitrate excretion. 4. Cholesterol feeding progressively increased aortic intimal thickening and impaired endothelium-dependent vasodilator function (P < 0.05 vs control), whereas indomethacin partially prevented aortic plaque formation and restored endothelium-dependent vasodilation (P < 0.05 vs the cholesterol-fed group). 5. The present study demonstrates that indomethacin reduces the progression of atherosclerotic lesions and improves endothelium-mediated vascular responses ex vivo in cholesterol-fed rabbits. The beneficial effects of indomethacin may be due to its ability to prevent the elevation of platelet aggregation, TXA2 (measured as urinary 2,3-dinor-TXB2 excretion) and 8-iso-PGF2alpha formation and to retard the decrease in endogenous nitric oxide synthesis (assessed as urinary excretion of nitrate). Despite indomethacin treatment leading to the suppression of prostacyclin biosynthesis (assessed as urinary 6-keto-PGF1alpha excretion), according to our data, indomethacin appears to preserve endothelial function.