A system for reporting and evaluating adverse drug reactions of herbal medicine in Taiwan from 1998 to 2016.

The Taiwan Adverse Drug Reaction Reporting System for Herbal Medicine (TADRRS-HM) has systematically documented suspected adverse events from adverse drug reaction (ADR) reports from 1998 (prior to its formal establishment in 2001) and evaluates safety profiles of herbal medicines. This article describes findings from 2079 ADR reports filed between 1998 and 2016: 941 reports involved single herbs and 87 involved folk herbals; 842 were generated from clinical trials, while 209 ADR reports involving foods, health foods, dietary supplement foods and herbal cuisine were grouped as Other. Severity assessments using the Modified Hartwig and Siegel scale classified 72.4% of ADRs as mild, 17.4% as moderate and 6.5% as severe. System Organ Class classification of the ADRs identified gastrointestinal system disorders as the most common (33.4%), followed by skin and subcutaneous tissue disorders (21.2%). The TADRRS-HM records indicate that herbal medicines may cause a wide range of ADRs. Aconiti Radix, Xiao-Qing-Long-Tang, and Datura suaveolens were the most commonly reported single herb, herbal formula, and folk herbal, respectively. The data indicate that herbal medicines may cause a wide range of ADRs. This system will confer long-term benefits for the development of Taiwan's herbal medicines adverse reaction database and facilitate epidemiological analysis.

Systematic probabilistic risk assessment of pesticide residues in tea leaves.

Multiple pesticide residues are frequently present in tea leaves and while the majority of residues satisfy Taiwan's current health regulations, there are potential health effects from pesticide exposure that are of great concern for tea drinkers. We undertook a systematic probabilistic risk assessment of 59 pesticides in tea leaves from 1629 tea leaf samples obtained by Taiwan's Food and Drug Administration in two monitoring surveys in 2015. Bayesian statistics used a Markov Chain Monte Carlo approach to estimate posterior distributions of pesticide residues in tea leaves, lifetime average daily doses and hazard quotients (HQs) of evaluated pesticides. We classified 95th percentile values of HQs into three categories: 0 < HQ < 0.5, 0.5 ≤ HQ ≤ 1 and 1 < HQ. The 95th percentiles of HQs for triazophos (3.39), carbofuran (2.04) and endosulfan (1.80) exceeded 1 in the adult population; the HQ for 3-OH carbofuran was 0.97 and was less than 0.5 for the remaining 55 pesticides. The health risk posed by pesticide residues for tea drinkers is negligible, if triazophos, carbofuran, endosulfan, and 3-OH carbofuran residues satisfy regulatory standards. However, five legacy pesticides, DDT, methomyl, carbofuran, dicofol and endosulfan, were identified. To reduce uncertainties, this study combined Bayesian statistics with a mode of action approach for systematic risk assessment of co-exposure to multiple pesticide residues in tea leaf samples. Measuring pesticide transfer rates will improve the quality of future risk assessments concerning residues in tea leaves. Appropriate management of pesticides in Taiwanese tea farms and monitoring of pesticide residues in imported tea is warranted to protect Taiwan's tea drinkers.

Exploration of anti-cancer effects and mechanisms of Zuo-Jin-Wan and its alkaloid components in vitro and in orthotopic HepG2 xenograft immunocompetent mice.

BACKGROUND: Zuo-Jin-Wan (ZJW), a two-herb formula consisting of Coptis chinensis (CC) and Evodia rutaecarpa (ER), is commonly used in traditional Chinese medicine for the treatment of cancers. However, the efficacies and mechanisms of ZJW and its alkaloid components on cancers are still unclear. METHODS: Here we investigated the anti-cancer effects and mechanisms of ZJW, CC, ER, berberine, and evodiamine in cells and in intrahepatic xenograft mice. RESULTS: Treatment of HepG2 cells with ZJW, CC, ER, berberine, and evodiamine significantly displayed cytotoxic effects in a dose- and time-dependent manner. Hierarchical cluster analysis of gene expression profiles showed that CC and ZJW shared a similar mechanism for the cytotoxic effects, suggesting that CC was the active ingredient of ZJW for anti-cancer activity. Network analysis further showed that c-myc was the likely key molecule involved in the regulation of ZJW-affected gene expression. A human hepatoma xenograft model was established by intrahepatic injection of HepG2 cells containing nuclear factor-κB-driven luciferase genes in immunocompetent mice. In vivo bioluminescence imaging showed that cells had been successfully transplanted in mouse liver. Oral administration of ZJW for 28 consecutive days led to a significant decrease in the accumulation of ascites, the ratio of tumor-to-liver, and the number of transplanted cells in livers. CONCLUSIONS: In conclusion, our findings suggested for the first time that ZJW significantly suppressed human cancer cell growth in orthotopic HepG2 xenograft-bearing immunocompetent mice. Moreover, c-myc might play a potent role in the cytotoxic mechanisms of ZJW, CC, ER, berberine, and evodiamine.

Anti-tumor effect of Radix Paeoniae Rubra extract on mice bladder tumors using intravesical therapy.

Radix Paeoniae Rubra (RPR) is the dried root of Paeonia lactiflora Pallas and Paeonia veitchii Lynch, and is a herbal medicine that is widely used in traditional Chinese medicine for the treatment of blood-heat and blood-stasis syndrome, similarly to Cortex Moutan. The present study identified the same three components in RPR and Cortex Moutan extracts. In addition, it has been reported that RPR has an anti-cancer effect. Bladder cancer is the seventh most common type of cancer worldwide. Due to the high recurrence rate, identifying novel drugs for bladder cancer therapy is essential. In the present study, RPR extract was evaluated as a bladder cancer therapy in vitro and in vivo. The present results revealed that RPR extract reduced the cell viability of bladder cancer cells with a half maximal inhibitory concentration of 1-3 mg/ml, and had an extremely low cytotoxic effect on normal urothelial cells. Additionally, RPR decreased certain cell cycle populations, predominantly cells in the G1 phase, and caused a clear sub-G increase. In a mouse orthotopic bladder tumor model, intravesical application of RPR extract decreased the bladder tumor size without altering the blood biochemical parameters of the mice. In summary, the present results demonstrate the anti-proliferative properties of RPR extract on bladder cancer cells, and its anti-bladder tumor effect in vivo. Compared to Cortex Moutan extract, RPR extract may provide a more effective alternative therapeutic strategy for the intravesical therapy of superficial bladder cancer.

Ethanol Extract of Hedyotis diffusa Willd Affects Immune Responses in Normal Balb/c Mice In Vivo.

Numerous clinical anticancer drugs are obtained from natural plants and Hedyotis diffusa Willd (EEHDW) has been used as a major component in Traditional Chinese medicine formulas since a long time. Ethanol extracts of EEHDW have been shown to possess various biological activities including anticancer function in vitro. Our earlier studies have shown that EEHDW affects immune responses in WEHI-3-generated leukemia mice, but EEHDW has not been reported to affect immune responses in a normal mouse model. Herein, we investigated whether EEHDW could affect immune responses on normal murine cells in vivo. Normal BALB/c mice were orally treated with or without EEHDW at 0, 16, 32, and 64 mg/kg or 32 mg/kg by i.p. for 3 weeks, then were weighed, and blood, liver and spleen samples were collected for further experiments. Results indicated that EEHDW did not significantly affect body and liver weight but significantly increased the spleen weight by i.p. treatment when compared to control groups. Flow cytometric assays indicated that EEHDW promoted CD11b levels at 16, 32 and 64 mg/kg oral treatment, CD19 levels at 16, 32, 64 mg/kg oral treatment and i.p. treatment, and Mac-3 levels at 16, 32 and 64 mg/kg oral treatment, however, it did not significantly affect the levels of CD3. Oral treatment with 16 and 32 mg/kg of EEHDW significantly decreased macrophage phagocytosis from PBMC; 32 mg/kg of EEHDW by i.p. treatment significantly increased phagocytosis activity of macrophages obtain from the peritoneal cavity. EEHDW at 32 mg/kg by i.p. treatment led to an increase of NK cell activities compared to oil control groups. EEHDW at 32 mg/kg of EEHDW by i.p. treatment increased B- and T-cell proliferation. Based on these observations, EEHDW seems to have promoted immune responses in this murine model.

Ethanol extract of Hedyotis diffusa willd upregulates G0/G1 phase arrest and induces apoptosis in human leukemia cells by modulating caspase cascade signaling and altering associated genes expression was assayed by cDNA microarray.

The authors' previous study has shown that water extract of Hedyotis diffusa Willd (HDW) promoted immune response and exhibited anti-leukemic activity in BALB/c leukemic mice in vivo. In this study, the anti-proliferation effects of ethanol extract of H. diffusa Willd (EEHDW) on lung cancer cell lines (A549, H1355, and LLC), leukemia cell lines (HL-60, WEHI-3), and a mouse melanoma cell line (B16F10) in vitro were investigated. The results demonstrated that EEHDW suppressed the cell proliferation of A549, H1355, HL-60, WEHI-3, and B16F10 cells as well as reduced cell viability in a concentration-dependent manner. We found that EEHDW inhibited the cell proliferation of HL-60 cells in concentration-dependent manner. In addition, EEHDW triggered an arrest of HL-60 cells at G0/G1 phase and sub-G1 population (apoptotic cells). EEHDW provoked DNA condensation and DNA damage in HL-60 cells. The activities of caspase-3, caspase-8, and caspase-9 were elevated in EEHDW-treated HL-60 cells. DNA microarray to investigate and display the gene levels related to cell growth, signal transduction, apoptosis, cell adhesion, cell cycle, DNA damage and repair, transcription and translation was also used. These findings suggest that EEHDW may be a potential herbal medicine and therapeutic agent for the treatment of leukemia.

Cortex Moutan Induces Bladder Cancer Cell Death via Apoptosis and Retards Tumor Growth in Mouse Bladders.

Cortex Moutan is the root bark of Paeonia suffruticosa Andr. It is the herbal medicine widely used in Traditional Chinese Medicine for the treatment of blood-heat and blood-stasis syndrome. Furthermore, it has been reported that Cortex Moutan has anticancer effect. In this study, the Cortex Moutan extract was evaluated in bladder cancer therapy in vitro and in vivo. Cortex Moutan extract reduces cell viability with IC50 between 1~2 mg/ml in bladder cancer cells, and it has lower cytotoxicity in normal urotheliums. It arrests cells in G1 and S phase and causes phosphatidylserine expression in the outside of cell membrane. It induces caspase-8 and caspase-3 activation and poly(ADP-ribose) polymerase degradation. The pan caspase inhibitor z-VAD-fmk reverses Cortex Moutan-induced cell death. Cortex Moutan also inhibits cell invasion activity in 5637 cells. In mouse orthotopic bladder cancer model, intravesical application of Cortex Moutan decreases the bladder tumor size without altering the blood biochemical parameters. In summary, these results demonstrate the antiproliferation and anti-invasion properties of Cortex Moutan in bladder cancer cells and its antibladder tumor effect in vivo. Cortex Moutan may provide an alternative therapeutic strategy for the intravesical therapy of superficial bladder cancer.

Analysis of urinary aristolactams by on-line solid-phase extraction coupled with liquid chromatography-tandem mass spectrometry.

Aristolochic acids (AAs), nephrotoxicants and known human carcinogens, are a mixture of structurally related derivatives of nitrophenanthrene carboxylic acids with the major components being aristolochic acid I and aristolochic acid II. People may ingest small amounts of AAs from its natural presence in medicinal plants and herbs of the family Aristolochiaceae, including the genera Aristolochia and Asarum, which have been used worldwide in folk medicine for centuries. In order to assess AA intake, an on-line solid-phase extraction coupled with liquid chromatography-tandem mass spectrometry (on-line SPE-LC/MS/MS) method was developed to analyze their most abundant corresponding metabolites, aristolactams (ALs), in urine to serve as biomarkers. The limits of quantitation were 0.006 ng for aristolactam I (AL-I), and 0.024 ng for aristolactam II (AL-II) on column. Recovery varied from 98.0% to 99.5%, and matrix effects were within 75.3-75.4%. This method was applied to analyze ALs in the urine samples collected on days 1, 2, 4, and 7 from mice treated with 30 mg/kg or 50mg/kg AAs. Their half lives were estimated to be 3.55 h and 4.00 for AL-I, and 4.04 and 4.83 h for AL-II, depending on AAs doses. These results demonstrated that the first simple on-line SPE-LC/MS/MS method was successfully developed to analyze urinary ALs with excellent sensitivity and specificity to serve as biomarkers to assess current AA intake from AAs-containing Chinese herbs.

Emodin, aloe-emodin and rhein inhibit migration and invasion in human tongue cancer SCC-4 cells through the inhibition of gene expression of matrix metalloproteinase-9.

Emodin, aloe-emodin and rhein are major compounds in rhubarb (Rheum palmatum L.), used in Chinese herbal medicine, and found to have antitumor properties including cell cycle arrest and apoptosis in many human cancer cells. Our previous studies also showed that emodin, aloe-emodin and rhein induced apoptosis in human tongue cancer SCC-4 cells. However, the detail regarding emodin, aloe-emodin and rhein affecting migration and invasion in SCC-4 cells are not clear. In the present study, we investigated whether or not emodin, aloe-emodin and rhein inhibited migration and invasion of SCC-4 cells. Herein, we demonstrate that emodin, aloe-emodin and rhein inhibit the protein levels and activities of matrix metalloproteinase-2 (MMP-2) but did not affect gene expression of MMP-2, however, they inhibited the gene expression of MMP-9 and all also inhibited the migration and invasion of human tongue cancer SCC-4 cells. MMP-9 (gelatinase-B) plays an important role and is the most associated with tumor migration, invasion and metastasis in various human cancers. Results from zymography and Western blotting showed that emodin, aloe-emodin and rhein treatment decrease the levels of MMP-2, urokinase plasminogen activator (u-PA) in a concentration-dependent manner. The order of inhibition of associated protein levels and gene expression of migration and invasion in SCC-4 cells are emodin >aloe-emodin >rhein. Our results provide new insight into the mechanisms by which emodin, aloe-emodin and rhein inhibit tongue cancers. In conclusion, these findings suggest that molecular targeting of MMP-9 mRNA expression by emodin, aloe-emodin and rhein might be a useful strategy for chemo-prevention and/or chemo-therapeutics of tongue cancers.

Gastrodia elata modulated activator protein 1 via c-Jun N-terminal kinase signaling pathway in kainic acid-induced epilepsy in rats.

Gastrodia elata (Orchidaceae) is a Chinese herb. Our previous study showed that Gastrodia elata is able to reduce epileptic seizures, oxygen free radicals, microglia activation, and apoptosis in kainic acid (KA)-treated rats. Activator protein 1 (AP-1) is involved in modulating the neuronal plasticity and apoptosis. Therefore, the aim of this study was to investigate the role of AP-1 in antiepileptic effect of Gastrodia elata. Gastrodia elata (0.5, 1.0g/kg) or valproic acid (VA, 250mg/kg) was administered orally in Sprague-Dawley rats for 1 week before and 2 weeks after intraperitoneal injection of KA. Protein levels of AP-1 were determined by measuring c-Jun and c-Fos proteins, and the mitogen-activated protein (MAP) kinases activations were determined by measuring the phosphorylations of extracellular signal-regulated kinases, p38, and c-Jun N-terminal kinases (JNKs) in the frontal cortex and the hippocampus of rat brain using Western blotting. These results indicated that pre-treatment with Gastrodia elata or VA activated JNK signal pathway and c-Jun expression, while post-treatment with Gastrodia elata or VA suppressed both the JNK signaling pathway and the c-Jun expression induced by KA. These findings suggested that Gastrodia elata regulated the AP-1 expression via the JNK signaling pathway in KA-induced epilepsy.

Inhibitory effect of Magnolia officinalis and lovastatin on aortic oxidative stress and apoptosis in hyperlipidemic rabbits.

Oxidative stress and apoptosis are 2 major characteristics of the progression of atherosclerosis. Both lovastatin and Magnolia officinalis are hypocholesterolemic agents. Therefore, we investigated the effect of M. officinalis extract on the prevention of atherosclerosis in comparison with lovastatin. Twenty hyperlipidemic rabbits were served one of the following diets: a high-fat and cholesterol diet (cholesterol group, 10% corn oil and 0.5% cholesterol), a high fat and cholesterol diet supplemented with M. officinalis extract (300 mg/kg) or lovastatin (6 mg/kg). The plasma lipids, oxidative stress (measured by free radical, malondialdehyde, and oxidative DNA damage), and arterial lesions significantly decreased in the M. officinalis and lovastatin groups when compared with the cholesterol group. Moreover, the expressions of Fas ligand, caspase 8, and caspase 9 in the aortic arches were also markedly lowered after M. officinalis and lovastatin supplements. Therefore, the results indicate that the antiatherogenic effect of M. officinalis is involved with a suppression of oxidative stress and with the down-regulation of apoptosis-related gene expression in hyperlipidemic rabbits.

Paeonol reduced cerebral infarction involving the superoxide anion and microglia activation in ischemia-reperfusion injured rats.

Both Moutan cortex of Paeonia suffruticosa Andrews (MC) and the root of Paeonia lactiflora Pall (PL) are important Traditional Chinese herbs used commonly to treat inflammatory and pyretic disorders. Paeonol, a common component of MC causes anti-platelet aggregation and scavenges free radicals. Therefore, the aim of the present study is to investigate the effects of Paeonol on cerebral infarct. A total of 60 male Sprague-Dawley (SD) rats were studied. An animal model of cerebral infarct was established by occluding both common carotid arteries and the right middle cerebral artery for 90 min, followed by a 24 h period of reperfusion. The percentage of cerebral infarction area to total brain area in each piece of brain tissue, and neuro-deficit score were measured. Superoxide anion was determined by the number of lucigenin-chemiluminescence (CL) counts. ED1 (mouse anti rat CD68) and interleukin-1beta (IL-1beta) immunostaining in the cerebral infarction region were also investigated for activation of microglia. The results indicated that Paeonol 15 and 20 mg/kg pretreatment and 20 mg posttreatment reduced the cerebral infarction area; Paeonol 15 and 20 mg/kg pretreatment reduced the neuro-deficit score. In addition, Paeonol 20 mg/kg pretreatment reduced the lucigenin-CL counts at 2 h period of reperfusion. The number of ED1 and IL-1beta immunoreactive cells also reduced in the cerebral infarction region; there were no significant changes in blood sugar levels. The results show that Paeonol reduced cerebral infarct and neuro-deficit in rat, suggesting Paeonol might play a similar role in reducing cerebral infarction in humans. Paeonol suppresses and scavenges superoxide anion, and inhibit microglia activation and IL-1beta in ischemia-reperfusion injured rats.

Gastrodia elata BL mediates the suppression of nNOS and microglia activation to protect against neuronal damage in kainic acid-treated rats.

Our previous studies showed that Gastrodia elata (GE), an herb used in traditional Chinese medicine, has both anti-convulsive and free radical-scavenging activities in kainic acid (KA)-treated rats. The aim of the present study was to further investigate possible physiological mechanisms of GE against activities of neuronal nitric oxide synthase (nNOS) and microglia in KA-treated rats; 0.5 g/kg and 1.0 g/kg of GE extract were administered orally, whereas 20 mg/kg of N-nitro-L-arginine methyl ester (L-NAME) was administered intraperitoneally (ip), both at 30 minutes prior to KA (2 microg/2 microl) being injected into the right hippocampus region of rats. ED1-staining, apoptotic, inducible nitric oxide synthase (iNOS), and nNOS-staining cells were observed in the hippocampus region. The results indicated that 1.0 g/kg of GE and 20 mg/kg of L-NAME reduced the counts of ED1-stained cells, and 0.5 g/kg and 1.0 g/kg of GE, and 20 mg/kg of L-NAME reduced the numbers of apoptotic cells and nNOS-staining cells. In addition, 20 mg/kg of L-NAME also reduced the numbers of iNOS-staining cells, but 0.5 g/kg and 1.0 g/kg of GE did not. This study demonstrated that GE was able to reduce nNOS, microglia activation and apoptosis, suggesting that GE has a protective effect against neuronal damage in KA-treated rats.

Effects of Ding-Chuan-Tang on bronchoconstriction and airway leucocyte infiltration in sensitized guinea pigs.

Ding-Chuan-Tang (DCT), a traditional Chinese medicine, has been used in treatment of the bronchial asthma for several centuries. However, the therapeutic mechanism of these Chinese medicine are still far from clear. To understand the mechanism of antiasthmatic property of DCT. A guinea pig model of allergic asthma was used to investigate the effects of DCT on ovalbumin-induced early and late asthmatic responses and airway inflammation, particularly the extent of eosinophil infiltration, and examine it direct beta2-adrenoceptor agonist activity in guinea-pig isolated trachea. We had used three different protocals in ovalbumin sensitized guinea pigs by administrating 10 g/kg of DCT extracts to sensitized guinea pigs 30 min before antigen challenge (group I), 5 hr after antigen challenge (group II) and 2.5 g/kg once daily from the day of sensitization to the day of challenge. Our result showed that administration of DCT singificantly inhibited the antigen induced immediate asthmatic responses (IAR) in group I and inhibited both IRA and late asthmatic responses (LAR) in actively sensitized guinea pig in group III. DCT caused concentration-dependent relaxations in strips of guinea pig trachea contracted with carbachol, however ICI-118551, a selective beta2-adrenoceptor antagonist, didn't significantly competitively inhibit the relaxations caused by DCT. Furthermore, examination of bronchoalveolar lavage fluid (BALF) revealed that DCT significantly inhibited the increase in percent of eosinophils in the airway after antigen challenge in three group. Histopathologic examination showed DCT suppressed the eosinophil infiltration into lung tissue. These results suggest that the antiasthmatic effect of DCT is mainly due to its bronchodilatation effect and its ability to inhibit the eosinophil into the airway and there is prophylactic effect of DCT on allergen-induced airway inflammation.

Formation of N-(2-hydroxyethyl)valine in human hemoglobin-effect of lifestyle factors.

The formation of N-(2-hydroxyethyl)valine (HEV) in hemoglobin has been considered as a biomarker to assess exogenous and endogenous exposures to ethylene oxide (EO) and/or ethylene (ET). Factors associated with daily exposures to such compounds might significantly affect the formation of HEV. Tobacco smoke containing EO elicited a significant increase in the levels of HEV amongst smokers, although other factors related to lifestyles may warrant further studies. The objective of this study was to specifically analyze HEV using a modified Edman degradation technique in order to study the association between lifestyle related factors (smoking, second-hand smoke exposure, tea and alcohol consumption) and HEV formation in vivo. Total of 148 Taiwanese volunteers with no history of occupational exposure to either EO or ET were recruited in this study. The HEV levels for smokers (204 +/- 151 pmol HEV/g globin, n = 70 ) were greater than those for non-smokers (57 +/- 46 pmol HEV/g globin, n = 78), HEV level increasing with the number of cigarettes smoked by subjects per day with a rate of 8.8 pmol HEV/g globin per cigarettes per day. Further analysis revealed that the rate of HEV formation in our study subjects was significantly associated with the number of daily cigarettes smoked (P < 0.001), but was not associated with tea or alcohol consumption, second-hand smoke exposure, subject age, or subject gender. These results suggest that the significantly higher levels of HEV for smokers than for non-smokers were mainly due to subject exposure to EO contained in cigarette smoke.