Differentiation of Cyanthillium cinereum, a smoking cessation herb, from its adulterant Emilia sonchifolia using macroscopic and microscopic examination, HPTLC profiles and DNA barcodes.

Cyanthillium cinereum (L.) H.Rob. is one of the most popular herbal smoking cessation aids currently used in Thailand, and its adulteration with Emilia sonchifolia (L.) DC. is often found in the herbal market. Therefore, the quality of the raw material must be considered. This work aimed to integrate macro- and microscopic, chemical and genetic authentication strategies to differentiate C. cinereum raw material from its adulterant. Different morphological features between C. cinereum and E. sonchifolia were simply recognized at the leaf base. For microscopic characteristics, trichome and pappus features were different between the two plants. HPTLC profiles showed a distinct band that could be used to unambiguously differentiate C. cinereum from E. sonchifolia. Four triterpenoid compounds, ß-amyrin, taraxasterol, lupeol, and betulin, were identified from the distinct HPTLC band of C. cinereum. The use of core DNA barcode regions; rbcL, matK, ITS and psbA-trnH provided species-level resolution to differentiate the two plants. Taken together, the integration of macroscopic and microscopic characterization, phytochemical analysis by HPTLC and DNA barcoding distinguished C. cinereum from E. sonchifolia. The signatures of C. cinereum obtained here can help manufacturers to increase the quality control of C. cinereum raw material in commercialized smoking cessation products.

Short-Term Efficacy and Safety of Adding Ezetimibe to Current Regimen of Lipid-Lowering Drugs in Human Immunodeficiency Virus-Infected Thai Patients Treated with Protease Inhibitors.

Long-term complications of protease inhibitor (PI) treatment includes increased cardiovascular risks due to dyslipidemia in patients infected with human immunodeficiency virus (HIV). Ezetimibe reduces low-density lipoprotein cholesterol (LDL-C) without drug interactions with PIs and statins. Furthermore, the addition of ezetimibe to statins is an optional treatment in HIV-infected patients with uncontrolled dyslipidemia. The objective of this study was to determine the short-term efficacy and safety of adding ezetimibe to the currently administered statin regimen. Thirty-two patients received ezetimibe (10 mg daily) in addition to their ongoing lipid-lowering therapy for 18 weeks. Serum LDL-C, total cholesterol (TC), triglycerides (TGs), TC/high-density lipoprotein cholesterol (HDL-C) ratio, and HDL-C were measured at baseline, and weeks 6, 12, and 18. Safety parameters were assessed by adverse event reports and laboratory assessments throughout the study. The mean percent change from baseline to endpoint in LDL-C, TC, TGs, and TC/HDL-C ratio were -23.3% (p<0.001), -15.0% (p=0.001), -22.1% (p=0.004), and -16.2% (p=0.018), respectively. No adverse event or other abnormal laboratory results occurred. Addition of ezetimibe to currently administered lipid-lowering drugs in HIV-infected patients receiving PIs with uncontrolled dyslipidemia demonstrated significantly improved efficacy in reducing their LDL-C, TC, TGs, and TC/HDL-C ratio levels. Moreover, this therapy was safe and well-tolerated.

Prevalence of CTR1 and ERCC1 Polymorphisms and Response of Biliary Tract Cancer to Gemcitabine-Platinum Chemotherapy

Purpose: Biliary tract cancer (BTC)is an aggressive disease with a poor prognosis. Most patients are diagnosed at an advanced stage for which curative surgery is not possible and gemcitabine-platinum chemotherapy is the treatment of choice for advanced cases. Several studies had focused on biomarkers to predict response from platinum drugs in lung cancer, but information is limited for BTC. In this study, two single nucleotide polymorphisms (SNPs) in the copper transporter (CTR1) and excision repair cross-complementary group 1 (ERCC1) genes were investigated as predictive biomarkers of objective response to gemcitabine-platinum. Methods: This cohort study aimed to assess any associations of genetic polymorphisms of these proteins active in drug pathway with treatment response in advanced BTC patients. Twenty six patients were enrolled. DNA was extracted from peripheral blood and genetic polymorphisms were assessed by Taqman allelic discrimination assay. Response was evaluated according to RECIST version 1.1. Results: For the CTR1 polymorphism, GT was the most common genotype (61.5%) followed by GG (34.6%), and TT (3.8%). For the ERCC1 polymorphism, only 2 genotypes were found, CC and CT at 57.7% and 42.3%, respectively. Genetic polymorphisms were not found to be singly associated with response. However, when the 2 genetic polymorphisms were combined, GG/CC showed a higher response rate than the others (p=0.018, Fisher's Exact Test). Conclusion: This is the first study to show an association between CTR1 and ERCC1 polymorphisms and response to gemcitabine-platinum in advanced BTC patients. These polymorphisms might be used as biomarkers to predict response in such cases in the future.

Influence of ABCC2 and ABCC4 polymorphisms on tenofovir plasma concentrations in Thai HIV-infected patients.

Tenofovir (TFV) is eliminated by renal excretion, which is mediated through multidrug-resistant protein 2 (MRP2) and MRP4, encoded by ABCC2 and ABCC4, respectively. Genetic polymorphisms of these transporters may affect the plasma concentrations of tenofovir. Therefore, the aim of this study was to investigate the influence of genetic and nongenetic factors on tenofovir plasma concentrations. A cross-sectional study was performed in Thai HIV-infected patients aged ≥18 years who had been receiving tenofovir disoproxil fumarate at 300 mg once daily for at least 6 months. A middose tenofovir plasma concentration was obtained. Multivariate analysis was performed to investigate whether there was an association between tenofovir plasma concentrations and demographic data, including age, sex, body weight, estimated glomerular filtration rate (eGFR), hepatitis B virus coinfection, hepatitis C virus coinfection, duration of tenofovir treatment, concomitant use of ritonavir-boosted protease inhibitors, and polymorphisms of ABCC2 and ABCC4. A total of 150 Thai HIV-infected patients were included. The mean age of the patients was 43.9 ± 7.2 years. The mean tenofovir plasma concentration was 100.3 ± 52.7 ng/ml. In multivariate analysis, a low body weight, a low eGFR, the concomitant use of ritonavir-boosted protease inhibitors, and the ABCC4 4131T → G variation (genotype TG or GG) were independently associated with higher tenofovir plasma concentrations. After adjusting for weight, eGFR, and the concomitant use of ritonavir-boosted protease inhibitors, a 30% increase in the mean tenofovir plasma concentration was observed in patients having the ABCC4 4131 TG or GG genotype. Both genetic and nongenetic factors affect tenofovir plasma concentrations. These factors should be considered when adjusting tenofovir dosage regimens to ensure the efficacy and safety of a drug. (This study has been registered at ClinicalTrials.gov under registration no. NCT01138241.).

Long-term cisplatin exposure impairs autophagy and causes cisplatin resistance in human lung cancer cells.

Cisplatin-based chemotherapy frequently resulted in acquired resistance of cancer cells. The underlying mechanism of such resistance is not fully understood especially the involvement of autophagy and autophagic cell death. This study thus investigated whether an alteration in autophagy could be responsible for cisplatin resistance in the long-term exposure lung carcinoma cells. The cisplatin resistant clone (H460/cis) of H460 cells was established by exposing the cells with gradually increasing concentrations of cisplatin until chemoresistance acquisition was elucidated by MTT, Hoechst 33342 staining and comet assays. Degree of autophagosome formation and level of LC3 marker were evaluated by acridine orange and western blot analysis, respectively. H460/cis cells exhibited irregular shape with ~3-fold resistant to cisplatin-induced apoptosis compared with H460 cells. Proteins analysis for LC3 indicated that the levels of LC3 in resistant cells were significantly lower than those in H460 cells. Moreover, autophagosome formation detected by acridine orange staining was dramatically reduced in the resistant cells, suggesting the role of autophagy in attenuating of cisplatin-induced cell death. Further, co-treatment of cisplatin with autophagy inducer, trifluorperazine, could resensitize H460/cis cells to cisplatin-induced cell death. Our findings reveal the novel mechanisms causing cisplatin resistance in lung carcinoma cells after long-term drug exposure regarding autophagy.

Protective effect of quercetin on hydrogen peroxide-induced tight junction disruption.

Tight junction is a crucial structure in the control of paracellular transport across epithelial/endothelial barriers. This study investigated the protective effect of quercetin against hydrogen peroxide (H(2)O(2))-induced tight junction disruption and hyperpermeability in ECV304 monolayers. Nonlethal concentration of H(2)O(2) (100 micromol/L; 4 hours) decreased expression of the tight junction proteins zonular occudens (ZO)-1 and occludin as well as disrupted the junction structure at the cell border. Concurrently, the increased activities of extracellular signal-regulated kinase (ERK)1/2 and p38 mitogen-activated protein kinase (MAPK) were observed. Pretreatment with quercetin (10 micromol/L; 30 minutes) prior to H(2)O(2) prevented the loss of ZO-1 and occludin. In addition, the level of phosphorylated p38 MAPK, but not that of the phosphorylated ERK1/2, decreased in quercetin-pretreated group. These findings suggested that the protective effect of quercetin involved the inhibition of phosphorylated p38 MAP activity. Furthermore, quercetin could also preserve the functional integrity of ECV304 monolayers from H(2)O(2) exposure.

Canarosine: a new guanidine alkaloid from Canavalia rosea with inhibitory activity on dopamine D1 receptors.

A new acyclic guanidine alkaloid, canarosine (1), together with five known compounds, beta-sitosterol (2), stigmasterol (3), daucosterol (4), epi-inositol 6-O-methyl ether (5), and rutin (6), were isolated from the aerial parts of Canavalia rosea. Their structures were established on the basis of their spectroscopic data. In the radioligand receptor binding assay, canarosine (1), at a concentration of 100 microg/ml, caused 91% inhibition of the dopamine D1 receptor binding with an IC50 value of 39.4 +/- 5.8 microM.

Satratoxin H generates reactive oxygen species and lipid peroxides in PC12 cells.

Satratoxin H, a mycotoxin, is thought to induce apoptosis of PC12 cells through the activation of p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) in a glutathione (GSH)-sensitive manner. The present study was undertaken to further elucidate the mechanism by which satratoxin H induces cell death in PC12 cells. Satratoxin H caused apoptosis of PC12 cells within 24-h, as determined by DNA fragmentation and flow cytometric analysis. Satratoxin H increased reactive oxygen species (ROS) production and lipid peroxidation, as determined by malondialdehyde formation. These effects were attenuated by incubation of cells with GSH, suggesting that satratoxin H-induced increase in apoptosis of serum-deprived PC12 cells may be partially mediated through the generation of ROS.

Cytotoxic C-benzylated chalcone and other constituents of Ellipeiopsis cherrevensis.

A new natural C-benzylated chalcone, 2',4'-dihydroxy-3'-(2-hydroxybenzyl)-6'-methoxychalcone (2), along with two other flavonoids, tiliroside and kaempferol 3-O-rutinoside, and an oxoaporphine alkaloid, lanuginosine were isolated from the aerial parts of Ellipeiopsis cherrevensis (Annonaceae). Two known polyoxygenated cyclohexene derivatives, ferrudiol and zeylenol, and a new analog, ellipeiopsol D, were also isolated. The chalcone 2 exhibited cytotoxic activity against human small-cell lung-cancer (NCI-H187), epidermoid carcinoma (KB) and breast cancer (BC) cell lines with IC50 values of 1.40, 5.31 and 13.92 microg/mL, respectively. This compound also showed antimalarial activity against Plasmodium falciparum with an IC50 value of 7.1 microg/mL as well as antimicrobacterial activity against Mycobacterium tuberculosis with a MIC of 25 mg/mL.

Involvement of reactive oxygen species and stress-activated MAPKs in satratoxin H-induced apoptosis.

Satratoxins, members of the trichothecene mycotoxin family, have been known to be harmful to health. However, the mechanisms underlying the toxicity still remain unclear. The present study is undertaken to elucidate the mechanisms of the satratoxin H-induced cytotoxicity in PC12 cells. Satratoxin H caused cytotoxicity, which was reflected from apoptosis determined by chromatin staining and flow cytometry. Satratoxin H stimulated the phosphorylation of extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK). Pre-incubation with SB203580, a p38 MAPK inhibitor, or SP600125, a JNK inhibitor, but not PD98059, an ERK inhibitor, reduced satratoxin-induced cytotoxicity. Co-incubation of cells with glutathione, N-acetyl-L-cysteine or glutathione reductase inhibited cytotoxicity and the phosphorylation of p38 MAPK induced by satratoxin H. Our data suggest that satratoxin H-induced apoptosis in PC12 cells is dependent on the activation of p38 MAPK/JNK and the increase in reactive oxygen species.