Schisanhenol Attenuates OxLDL-Induced Endothelial Dysfunction via an AMPK-Dependent Mechanism.

Atherosclerotic cardiovascular diseases, commonly known as the formation of fibrofatty lesions in the artery wall, are the leading causes of death globally. Oxidized low-density lipoprotein (oxLDL) is one of the major components of atherosclerotic plaques. It is evident that dietary supplementation containing sources of antioxidants can prevent atherogenic diseases. Schisanhenol (SAL), a dibenzocyclooctene lignin, has been shown to attenuate oxLDL-induced apoptosis and the generation of reactive oxygen species (ROS) in endothelial cells. However, the underlying molecular mechanisms are still largely unknown. In this study, human umbilical vein endothelial cells (HUVECs) were pre-treated with SAL and oxLDL. Our results showed that adenosine monophosphate-activated protein kinase (AMPK) phosphorylation was enhanced in cells pre-treated with SAL in time-dependent and dose-dependent manners. Subsequently, oxLDL-induced AMPK dephosphorylation and protein kinase C (PKC) phosphorylation were significantly reversed in the presence of SAL. In addition, SAL treatment led to an inhibiting effect on the oxLDL-induced membrane assembly of NADPH oxidase subunits, and a similar effect was observed in ROS generation. This effect was further confirmed using knockdown AMPK with small interfering RNA (siRNA) and pharmaceutical reagents, such as the AMPK activator (AICAR), PKC inhibitor (Gö 6983), and ROS inhibitor (DPI). Furthermore, the oxLDL-induced intracellular calcium rise and the potential collapse of the mitochondrial membrane reduced the Bcl-2/Bax ratio, and released cytochrome c from the mitochondria, leading to the subsequent activation of caspase-3 in HUVECs, which were also markedly suppressed by SAL pretreatment. The results mentioned above may provide additional insights into the possible molecular mechanisms underlying the cardiovascular protective effects of SAL.

Polyunsaturated fatty acids and inflammatory markers in major depressive episodes during pregnancy.

INTRODUCTION: Prenatal depression (PND) is a common psychiatric disorder in pregnant women and leads to psychosocial dysfunction, high suicidal rate, and adverse childcare. Patients with PND have omega-3 polyunsaturated fatty acid (omega-3 or n-3 PUFAs) deficits, which might link to chronic low-grade inflammatory process and the pathophysiological mechanisms of depression. In this case-control study, we examined the levels of PUFAs and inflammatory cytokines in PND. METHOD: Blood samples were obtained and analyzed from 16 healthy controls and 17 depressed cases (PND group) diagnosed with Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV). Independent sample t-test and correlation analysis were performed with Statistical Package for the Social Sciences (SPSS) logistics correlation analysis. RESULTS: PND group had significantly lower levels of total n-3 (p=0.026), docosahexaenoic acid (DHA) (p=0.020) and eicosapentaenoic (EPA) (p=0.019) but a higher omega-6 (n-6)/n-3 PUFAs ratio (p=0.007) and tumor necrosis factor alpha (TNF-α) (p=0.016) level. Moreover, the duration of current PND episodes were also significantly correlated with DHA, EPA, n-3 PUFAs, n-6/n-3 ratio and TNF-α. In terms of PUFAs and cytokine levels, only DHA was inversely correlated with TNF-α. CONCLUSION: PND is significantly associated with lower DHA, EPA, and total n-3 PUFAs levels and an increased n-6/n-3 PUFAs ratio, while the duration of PND is associated with lower levels of n-3 PUFAs, including DHA and EPA. The correlation of PUFAs levels with depression and TNF-α level grant further investigation into the inflammatory process underlying PND, mediated by PUFAs.

Diallyl trisulfide (DATS) inhibits mouse colon tumor in mouse CT-26 cells allograft model in vivo.

Our earlier studies showed that DATS induced apoptosis in human colon cancer HT29 and colo 205 cell lines in vitro. However, there is no report to show that DATS induced apoptosis in vitro and inhibited CT26 cancer cells in vivo on a murine allograft animal model. In vitro studies, the results indicated that DATS induced morphological changes and induction of apoptosis in CT26 cells. In vivo studies, CT26 cancer cells were implanted into BALB/c mice and groups of mice were treated with vehicle, DATS (10 and 50 mg/kg of body weight). DATS were injected once per four days intraperitoneally (i.p.), with treatment starting 4 weeks prior to cells inoculation. Treatment with vehicle or with 10 and 50 mg/kg of DATS resulted in a reduction in tumor volume and weight. Tumor volume and total hemoglobin in allograft mice treated with 50 mg/kg DATS were significantly smaller than that in the control group. These findings indicated that DATS inhibits tumor growth in an allograft animal model. Thus, DATS may represent a colon cancer preventive agent and can be used in the future.

The roles of AIF and Endo G in the apoptotic effects of benzyl isothiocyanate on DU 145 human prostate cancer cells via the mitochondrial signaling pathway.

It is well known that the response of cancer cells to chemotherapeutic drugs involves the activation of apoptotic pathways. Benzyl isothiocyanate (BITC) is an important compound found in plant food and has been shown to have anti-cancer effects on human cancer cells, but its effect on prostate cancer cells in vitro remains unknown. The aim of the present study was to investigate the effects of BITC on DU 145 human prostate cancer cells in order to clarify whether a time/concentration range for optimal BITC-induced apoptosis exists and to find the associated signaling pathway. Cell morphological changes, percentage of cell viability, DNA damage and apoptosis in DU 145 cells were examined by phase-contrast microscopy, flow cytometric assay, 4',6-diamidine-20-phenylindole dihydrochloride staining, comet assay and Western blotting analysis. The results indicate that BITC induces cell morphological changes, decreases the percentage of viable cells (induction of cell cytotoxicity), and induces DNA damage and apoptosis in DU 145 cells in a time- and dose-dependent manner. Flow cytometric assays indicated that BITC promoted reactive oxygen species and Ca2+ productions and decreased the levels of mitochondrial membrane potential (ΤYm), while the pre-treatment with N-acetylcysteine caused an increase in the percentage of viable cells. BITC also promoted caspase-3, -8 and -9 activities. Furthermore, when cells were pre-treated with the caspase-3 inhibitor and then treated with BITC, this led to an increase in the percentage of viable cells. Confocal laser microscopy examination indicated that BITC promoted the expression of AIF and Endo G, which were released from the mitochondria in DU 145 cells. In conclusion, BITC induces apoptosis in DU 145 cells through the release of AIF and Endo G from the mitochondria and also promotes caspase-3 activation.

Effects of acupuncture on post-cesarean section pain.

BACKGROUND: Post-operation pain is a very subjective phenomenon. The aim of this study was to find out the effects of acupuncture or electro-acupuncture on post-cesarean pain. METHODS: Sixty women, who had had spinal anesthesia during cesarean section at the Department of Obstetrics of China Medical University Hospital, were randomly assigned to the control group, the acupuncture group, and the electro-acupuncture group. After the operation, we applied subjects with acupuncture or electro-acupuncture on the bilateral acupuncture point, San Yin Jiao (Sp6), and the patient controlled analgesia (PCA). The first time of requesting morphine, the frequency of PCA demands in 24 hours, and the doses of PCA used were recorded double blindly. In addition, monitoring the subjects' vital signs, the opioid-related side effects, and the pain scores was done. RESULTS: The results showed that the acupuncture group and the electro-acupuncture group could delay the time of requesting morphine up to 10 - 11 minutes when compared with the control group. The total dose of PCA used within the first 24 hours was 30% - 35% less in the acupuncture group and the electro-acupuncture group when compared with the control group, which was indicated in statistical significance. However, there was no significant difference between the acupuncture group and the electro-acupuncture group. The electro-acupuncture group's and the acupuncture group's pain scores were lower than the control group's within the first 2 hours. Both were statistically significant. However, two hours later, there were no significant differences of the visual analogue scale (VAS) scores between either of the treatment groups and the control group. Finally, the incidence of opioid-related side effects, such as dizziness, was less in the acupuncture group and electro-acupuncture group than in the control group. CONCLUSIONS: This study shows that the application of acupuncture and electro-acupuncture could definitely delay the time of requesting pain relief medication after cesarean section and decrease the PCA doses used within the first 24 hours.

Crude extracts of Solanum lyratum protect endothelial cells against oxidized low-density lipoprotein-induced injury by direct antioxidant action.

BACKGROUND: Oxidized low-density lipoprotein (oxLDL) is a proatherogenic molecule that accumulates in the vascular wall and contributes to the pathogenesis of vascular dysfunction early in the development of atherosclerosis. The whole plant of Solanum lyratum is a traditional Chinese medicine that has been used for centuries to treat cancer, tumors, and herpes. However, the cellular and molecular mechanisms of its antioxidant effects are still largely unknown. This study tested the hypothesis that Solanum lyratum Thunberg extract (SLE) could block oxLDL-induced endothelial dysfunction in cultured human umbilical vein endothelial cells (HUVECs). Possible mechanisms were explored. METHODS: Antioxidative activities of SLE were assayed by measuring the scavenging of 1,1-diphenyl-2-picryl-hydrazyl (DPPH) free radical and the inhibition of copper-mediated or cell-mediated LDL oxidation. Production of reactive oxygen species (ROS) and the expression of adhesion molecules were evaluated in HUVECs after exposure to oxLDL and treatment with SLE. Several apoptotic signaling pathways were investigated. RESULTS: SLE scavenged DPPH and also delayed the kinetics of LDL oxidation in a dose-dependent manner. SLE attenuated the level of oxLDL-induced ROS generation, diminished the expression of endothelial NO synthase (eNOS), and enhanced the expression of adhesion molecules (vascular cellular adhesion molecule-1, E-selectin, and monocyte chemotactic protein-1) and the adherence of monocytic THP-1 cells to HUVECs. OxLDL increased the concentration of intracellular calcium, disturbed the balance of the Bcl-2 protein family, destabilized the mitochondrial membrane potential, increased the amount of cytochrome c released into the cytosol, and increased the activation of caspase 3. These detrimental effects were ameliorated dose-dependently by SLE (P < .05). CONCLUSION: Crude extracts of Solanum lyratum protect against oxLDL-induced injury in endothelial cells by direct antioxidant action. CLINICAL RELEVANCE: Atherosclerosis is a chronic inflammatory disease characterized by lipid-laden lesions within arterialblood vessel walls. Inhibiting the oxidation of low-density lipoprotein may be an effective way to prevent or delay theprogression of atherosclerosis. This study underscores the potential clinical benefits and application of Solanum lyratumextract in controlling oxidized low-density lipoprotein-associated vascular injury and cardiovascular disease.

Ginkgo biloba extract attenuates oxLDL-induced oxidative functional damages in endothelial cells.

Atherosclerosis is a chronic inflammatory process with increased oxidative stress in vascular endothelium. Ginkgo biloba extract (GbE), extracted from Ginkgo biloba leaves, has commonly been used as a therapeutic agent for cardiovascular and neurological disorders. The aim of this study was to investigate how GbE protects vascular endothelial cells against the proatherosclerotic stressor oxidized low-density lipoprotein (oxLDL) in vitro. Human umbilical vein endothelial cells (HUVECs) were incubated with GbE (12.5-100 microg/ml) for 2 h and then incubated with oxLDL (150 microg/ml) for an additional 24 h. Subsequently, reactive oxygen species (ROS) generation, antioxidant enzyme activities, adhesion to monocytes, cell morphology, viability, and several apoptotic indexes were assessed. Our data show that ROS generation is an upstream signal in oxLDL-treated HUVECs. Cu,Zn-SOD, but not Mn-SOD, was inactivated by oxLDL. In addition, oxLDL diminished expression of endothelial NO synthase and enhanced expression of adhesion molecules (ICAM, VCAM, and E-selectin) and the adherence of monocytic THP-1 cells to HUVECs. Furthermore, oxLDL increased intracellular calcium, disturbed the balance of Bcl-2 family proteins, destabilized mitochondrial membrane potential, and triggered subsequent cytochrome c release into the cytosol and activation of caspase-3. These detrimental effects were ameliorated dose dependently by GbE (P < 0.05). Results from this study may provide insight into a possible molecular mechanism underlying GbE suppression of the oxLDL-mediated vascular endothelial dysfunction.

Gypenosides induced G0/G1 arrest via inhibition of cyclin E and induction of apoptosis via activation of caspases-3 and -9 in human lung cancer A-549 cells.

Gynostemma pentaphyllum Makino is known in Asia for its effect on the treatment of hepatitis and cardiovascular diseases. Gypenosides (Gyp) are the major components extracted from Gynostemma pentaphyllum Makino. However, the molecular mechanism underlying the Gyp-induced cell cycle arrest and apoptotic process is unclear. In this study, the chemopreventive role of Gyp in human lung cancer (A549) cells in vitro was evaluated by studying the regulation of the cell cycle and apoptosis. Gyp induced GO/G1 arrest and apoptosis in the human lung cancer A549 cells. Investigation of the cyclin-dependent protein kinase inhibitors by Western blotting showed that p16, p21, p27 and p53 proteins were increased with the increasing time of incubation with Gyp in the A549 cells. This increase may be the major factor by which Gyp caused GO/G1 arrest in the examined cells. Flow cytometric assay and gel electrophoresis of DNA fragmentation also confirmed that Gyp induced apoptosis in the A549 cells. Our data demonstrated that Gyp-induced apoptotic cell death was accompanied by up-regulation of Bax, caspase-3 and caspase-9, but down-regulation of the Bcl-2 levels. Taken together, Gyp appears to exert its anticancer properties by inducing GO/GI-phase arrest and apoptosis via activation of caspase-3 in human lung A549 cancer cells.

Omega-3 fatty acids for major depressive disorder during pregnancy: results from a randomized, double-blind, placebo-controlled trial.

BACKGROUND: Perinatal depression is common, and treatment remains challenging. Depression has been reported to be associated with the abnormality of omega-3 polyunsaturated fatty acids (PUFAs). A profound decrease of omega-3 PUFAs in the mother during pregnancy is associated with the higher demand of fetal development and might precipitate the occurrence of depression. In this study, we examined the efficacy of omega-3 PUFA monotherapy for the treatment of depression during pregnancy. METHOD: From June 2004 to June 2006, we conducted an 8-week, double-blind, placebo-controlled trial comparing omega-3 PUFAs (3.4 g/d) with placebo in pregnant women with major depressive disorder (DSM-IV criteria). No psychotropic agent was given 1 month prior to or during the study period. The Hamilton Rating Scale for Depression (HAM-D) was scored every other week as the primary measurement of efficacy, while the Edinburgh Postnatal Depression Scale (EPDS) and Beck Depression Inventory (BDI) were secondary measures. RESULTS: Thirty-six subjects were randomly assigned to either omega-3 PUFAs or placebo, and 33 among them were evaluated in more than 2 visits. A total of 24 subjects completed the study. As compared to the placebo group, subjects in the omega-3 group had significantly lower HAM-D scores at weeks 6 (p = .001) and 8 (p = .019), a significantly higher response rate (62% vs. 27%, p = .03), and a higher remission rate, although the latter did not reach statistical significance (38% vs. 18%, p = .28). At the study end point, subjects in the omega-3 group also had significantly lower depressive symptom ratings on the EPDS and BDI. The omega-3 PUFAs were well tolerated and there were no adverse effects on the subjects and newborns. CONCLUSIONS: Omega-3 PUFAs may have therapeutic benefits in depression during pregnancy. In regard to the safety issue and psychotherapeutic effect, as well as health promotion to mothers and their newborns, it is worthy to conduct replication studies in a larger sample with a broad regimen of omega-3 PUFAs in pregnant women with depression. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00618865.

Effects of curcumin on N-bis(2-hydroxypropyl) nitrosamine (DHPN)-induced lung and liver tumorigenesis in BALB/c mice in vivo.

Curcumin (diferuloylmethane), a phenolic compound from the plant Curcuma longa (Linn.) has been shown to exhibit antitumor activity and apoptosis in many human cancer cell lines including that of lung and liver cancer. In this study, curcumin was evaluated in BALB/c mice for its ability to inhibit pulmonary and liver adenoma formation and growth after they were orally treated with N-bis(2-hydroxypropyl)nitrosamine (DHPN). Animals were treated with DHPN in water for approximately 14 days before multiple doses of curcumin were given intraperitoneally. It was found that 200 microM curcumin reduced lung and liver tumor multiplicity by 37% (p<0.05) and 30% (p<0.05) respectively. The results indicated that curcumin significantly inhibited pulmonary and liver adenoma formation and growth in BALB/c mice. The precise mechanism by which curcumin inhibits lung and liver tumorigenesis remains to be elucidated. Thus, curcumin appears to be a promising new chemotherapeutic and preventive agent for lung and liver cancer induced by DHPN.

Gypenosides inhibited N-acetylation of 2-aminofluorene, N-acetyltransferase gene expression and DNA adduct formation in human cervix epithelioid carcinoma cells (HeLa).

N-acetylation plays an important role in the metabolism of arylamine drugs and carcinogens and is catalyzed by cytosolic N-acetyltransferase (NAT). Gypenosides are the major components of Gynostemma pentaphyllum Makino which had been used as a natural folk medicine in the Chinese populations. Gypenosides were selected for examining the inhibition on the N-acetylation of 2-aminofluorene (AF), DNA-AF adduct formation and NAT gene expression in the human cervix epithelioid carcinoma cell line (HeLa). Various concentrations of gypenosides were individually added to the culture medium of human cervix epithelioid carcinoma cells (HeLa). The N-acetylation of AF was determined by high performance liquid chromatography (HPLC) assaying for the amounts of acetylated 2-aminofluorene (AAF) and nonacetylated 2-aminofluorene (AF). The N-acetylation of AF in the human HeLa cancer cells was suppressed by gypenosides in a dose-dependent manner. The data also demonstrated that gene expression (NAT1 mRNA) of NAT in human cervix epithelioid carcinoma cells (HeLa) was inhibited and decreased by gypenosides. After the incubation of HeLa cells with 30 or 60 microM AF and with or without 350 microg/ml gypenosides cotreatment, DNA was isolated and hydrolyzed to nucleotides, adducted nucleotides were extracted into butanol and analyzed DNA-AF adducts by HPLC. The data demonstrated that gypenosides decrease the levels of DNA-AF adduct formation in HeLa cells.

N-acetyltransferase is involved in gypenosides-induced N-acetylation of 2-aminofluorene and DNA adduct formation in human cervix epidermoid carcinoma cells (Ca Ski).

The effects of gypenosides on the inhibition of N-acetyltransferase (NAT) activity, AF-DNA adduct formation and NAT gene expression in a human cervix cancer cell line (Ca Ski) were studied. Various concentrations of gypenosides were added to the cytosols or individually to the culture medium of human cervix cancer cells. The NAT activity was determined by high performance liquid chromatography, assaying for the amounts of acetylated 2-aminofluorene (AAF) and non-acetylated 2-aminofluorene (AF). The NAT activity in the human cervix intact cancer cells and cytosols was suppressed by gypenosides in a dose-dependent manner. The results also demonstrated that gene expression (NAT1 mRNA) in human cervix cancer cells was decreased by gypenosides in a dose-dependent manner. The apparent values of Km and Vmax of NAT of human cervix cancer cells were also decreased by gypenosides in cytosols. Gypenosides may act as noncompetitive inhibitors. After the incubation of human cervix cancer cells with 30 or 60 microM AF and with or without 350 micrograms/ml gypenosides co-treatment, the cells were recovered, DNA was prepared and hydrolyzed to nucleotides; adducted nucleotides were extracted in butanol and AF-DNA adducts were analyzed by HPLC. The results demonstrated that gypenosides decreased the levels of AF-DNA adduct formation in these cells. The NAT PCR and cDNA microarray also demonstrated that gypenosides inhibited NAT mRNA expression in human cervix cancer cells.