Cucurbitacin E Exerts Anti-Proliferative Activity via Promoting p62-Dependent Apoptosis in Human Non-Small-Cell Lung Cancer A549 Cells.

EGFR tyrosine kinase inhibitors (TKIs) are the first-line treatment for advanced EGFR-mutated non-small-cell lung cancer (NSCLC). However, NSCLC patients with wild-type EGFR and KRAS mutation are ineligible for EGFR-TKIs. Therefore, the discovery of new therapeutic agents is urgently needed for NSCLC patients who cannot receive targeted therapies. Natural products possess tremendous chemical diversity and have been extensively investigated for their anticancer activity. In this study, we found that Cucurbitacin E (Cu E), a triterpene of cucurbitacins widely presented in the edible plants of the Cucurbitaceae family, significantly inhibits the viability and proliferation of A549 cells that harbor wild-type EGFR and KRAS mutation. Our results revealed that Cu E increases cell-cycle arrest at G2/M and subG1 phase. Mechanistically, Cu E significantly inhibits the phosphorylation and protein levels of regulatory proteins and hinders G2/M cell-cycle progression. Meanwhile, the treatment of Cu E resulted in DNA damage response and apoptosis. For the first time, we observed that Cu E induces incomplete autophagy as evidenced by increased LC3B-II expression and p62-accumulation. Knockdown of p62 rescued the cells from Cu E-mediated anti-proliferative effect, apoptosis, DNA damage, and ROS production. These findings suggest that Cu E is a promising drug candidate for NSCLC.

E7050 Suppresses the Growth of Multidrug-Resistant Human Uterine Sarcoma by Inhibiting Angiogenesis via Targeting of VEGFR2-Mediated Signaling Pathways.

E7050 is an inhibitor of VEGFR2 with anti-tumor activity; however, its therapeutic mechanism remains incompletely understood. In the present study, we aim to evaluate the anti-angiogenic activity of E7050 in vitro and in vivo and define the underlying molecular mechanism. It was observed that treatment with E7050 markedly inhibited proliferation, migration, and capillary-like tube formation in cultured human umbilical vein endothelial cells (HUVECs). E7050 exposure in the chick embryo chorioallantoic membrane (CAM) also reduced the amount of neovessel formation in chick embryos. To understand the molecular basis, E7050 was found to suppress the phosphorylation of VEGFR2 and its downstream signaling pathway components, including PLCγ1, FAK, Src, Akt, JNK, and p38 MAPK in VEGF-stimulated HUVECs. Moreover, E7050 suppressed the phosphorylation of VEGFR2, FAK, Src, Akt, JNK, and p38 MAPK in HUVECs exposed to MES-SA/Dx5 cells-derived conditioned medium (CM). The multidrug-resistant human uterine sarcoma xenograft study revealed that E7050 significantly attenuated the growth of MES-SA/Dx5 tumor xenografts, which was associated with inhibition of tumor angiogenesis. E7050 treatment also decreased the expression of CD31 and p-VEGFR2 in MES-SA/Dx5 tumor tissue sections in comparison with the vehicle control. Collectively, E7050 may serve as a potential agent for the treatment of cancer and angiogenesis-related disorders.

Phase I Dose-Escalation Study of SCB01A, a Microtubule Inhibitor with Vascular Disrupting Activity, in Patients with Advanced Solid Tumors.

LESSONS LEARNED: SCB01A is a novel microtubule inhibitor with vascular disrupting activity. This first-in-human study demonstrated SCB01A safety, pharmacokinetics, and preliminary antitumor activity. SCB01A is safe and well tolerated in patients with advanced solid malignancies with manageable neurotoxicity. BACKGROUND: SCB01A, a novel microtubule inhibitor, has vascular disrupting activity. METHODS: In this phase I dose-escalation and extension study, patients with advanced solid tumors were administered intravenous SCB01A infusions for 3 hours once every 21 days. Rapid titration and a 3 + 3 design escalated the dose from 2 mg/m2 to the maximum tolerated dose (MTD) based on dose-limiting toxicity (DLT). SCB01A-induced cellular neurotoxicity was evaluated in dorsal root ganglion cells. The primary endpoint was MTD. Safety, pharmacokinetics (PK), and tumor response were secondary endpoints. RESULTS: Treatment-related adverse events included anemia, nausea, vomiting, fatigue, fever, and peripheral sensorimotor neuropathy. DLTs included grade 4 elevated creatine phosphokinase (CPK) in the 4 mg/m2 cohort; grade 3 gastric hemorrhage in the 6.5 mg/m2 cohort; grade 2 thromboembolic event in the 24 mg/m2 cohort; and grade 3 peripheral sensorimotor neuropathy, grade 3 elevated aspartate aminotransferase, and grade 3 hypertension in the 32 mg/m2 cohort. The MTD was 24 mg/m2 , and average half-life was ~2.5 hours. The area under the curve-dose response relationship was linear. Nineteen subjects were stable after two cycles. The longest treatment lasted 24 cycles. SCB01A-induced neurotoxicity was reversible in vitro. CONCLUSION: The MTD of SCB01A was 24 mg/m2 every 21 days; it is safe and tolerable in patients with solid tumors.

Anti-Acne Effects of Cembrene Diterpenoids from the Cultured Soft Coral Sinularia flexibilis.

Acne is a skin disease common in adolescents and increasingly common in the adult population. The major pathologic events of acne vulgaris include increased sebum production, retention hyperkeratosis, carrying commensal skin microbiota, and inflammation. In recent years, more than 10,000 compounds have been isolated and identified from marine organisms. The aim of this study was to discover the potential anti-acne activity of fraction 9 + 10 (SF-E) of Sinularia flexibilis extract and six cembrene diterpenoids. We found that the SF-E significantly reduced Cutibacterium acnes-induced edema in Wistar rat ears. The cembrene diterpenoids including 11-dehydrosinulariolide (SC-2), 3,4:8,11-bisepoxy-7-acetoxycembra-15(17)-en-1,12-olide (SC-7), and sinularin (SC-9) reduced nitric oxide (NO) production with 50% inhibitory concentration of 5.66 ± 0.19, 15.25 ± 0.25, and 3.85 ± 0.25 µM, respectively, and inducible NO synthase expression in RAW 264.7 cells. Moreover, treatment with SC-2, SC-7, and SC-9 significantly suppressed lipopolysaccharide- and heat-killed C. acnes-induced expression of proteins involved in mitogen-activated protein kinase pathway in both RAW 264.7 and HaCaT cells. After treatment with SC-2, SC-7, and SC-9, over-proliferation of HaCaT cells was significantly terminated. In summary, SC-2, SC-7, and SC-9 showed anti-inflammatory effects in RAW 264.7 cells, suggesting that these cembrene diterpenoids obtained from S. flexibilis are natural marine products with potential anti-acne activities.

(+)-Erythro-Δ8'-7S,8R-Dihydroxy-3,3',5'-Trimethoxy-8-O-4'-Neolignan, an Anti-Acne Component in Degreasing Myristica fragrans Houtt.

Acne is a common skin condition observed in adolescents. Nutmeg (Myristica fragrans Houtt) (MF) is a well-known traditional Chinese medicine; its major toxic components, safrole and myristicin, are rich in essential oils. Essential oils of MF (MFO) were extracted by hydrodistillation; the residue was extracted using 50% methanol (MFE-M). The minimum inhibitory concentration (MIC) of MFE-M against Cutibacterium acnes and Staphylococcus aureus was 0.64 mg. Four compounds were obtained from MFE-M: myristicin (1), (+)-erythro-Δ8'-7S,8R- dihydroxy-3,3,5'-trimethoxy-8-O-4'-neolignan (2), (+)-erythro-Δ8'-7-hydroxy-3,4,3',5'-tetramethoxy 8-O-4-neolignan (3), and erythro-Δ8'-7-acetoxy-3,4,3',5'-tetramethoxy-8-O-4'-neolignan (4). Compound 2 exerted the strongest antimicrobial activity, with MICs of 6.25 and 3.12 µg/mL against C. acnes and S. aureus, respectively. Moreover, 2 inhibited NO, PGE2, iNOS, and COX-2 levels in RAW 264.7 cells induced by LPS or heat-killed C. acnes; NO production at 50% inhibitory concentrations (IC50) was 11.07 and 11.53 µg/mL, respectively. Myristicin and safrole content was higher in MFO than in MFE-M. MFO and MFE-M caused no skin irritation after a single topical application in Wistar rats. MFE-M, with low safrole and myristicin content, did not cause skin irritation and exhibited an anti-acne effect; moreover, 2 was identified as the active substance. Therefore, MFE-M could be employed to develop anti-acne compounds for use in cosmetics.

Stability of Principal Hydrolysable Tannins from Trapa taiwanensis Hulls.

The fruit and hulls of the water caltrop (Trapa taiwanensis Nakai) are used as hepatoprotective herbal tea ingredients in Taiwan. The stability of hydrolysable tannins in herbal drinks has rarely been reported. In the present study, two hydrolysable tannins, tellimagrandin II (TGII) and 1,2,3,4,6-pentagalloylglucopyranose (PGG), were isolated from water caltrop hulls. The stability of the two compounds was evaluated by treatment with various pH buffer solutions, simulated gastric fluid and intestinal fluid, different temperatures, and photo-irradiation at 352 nm in different solvents. Results showed that TGII and PGG were more stable in a pH 2.0 buffer solution (with 91.88% remaining) and in a water solution with 352 nm irradiation (with 95% remaining). TGII and PGG were more stable in methanol or ethanol solutions (with >93.69% remaining) than in an aqueous solution (with <43.52% remaining) at 100 °C. In simulated gastric fluid, more than 96% of the hydrolysable tannins remained after incubation at 37 °C for 4 h. However, these hydrolysable tannins were unstable in simulated intestinal fluid, as after incubation at 37 °C for 9 h, the content of TGII had decreased to 31.40% and of PGG to 12.46%. The synthetic antioxidants, butyl hydroxy anisole (BHA), di-butyl hydroxy toluene (BHT), and propyl gallate, did not exhibit photoprotective effects on these hydrolysable tannins. However, catechin, a natural antioxidant, displayed a weak photoprotective effect. Ascorbic acid had a short-term thermal-protective effect but not a long-term protective effect. The different stability properties of hydrolysable tannins in solutions can be used in the development of related herbal teas in the future.

The natural compound 2,3,5,4'-tetrahydroxystilbene-2-O-ß-d glucoside protects against adriamycin-induced nephropathy through activating the Nrf2-Keap1 antioxidant pathway.

2,3,5,4'-Tetrahydroxystilbene-2-O-ß-d-glucoside (THSG) is an active compound extracted from Polygonum multiflorum Thunb. This herb and radix Polygoni Multiflori preparata have been used to treat arteriosclerosis, hyperlipidemia, hypercholesterolemia, and diabetes for thousands of years. This study aimed to investigate the protective effects of THSG in an Adriamycin (AD)-induced focal segmental glomerulosclerosis (FSGS) mouse model and the underlying mechanisms in an in vitro system. Mice were treated with THSG (2.5 and 10 mg/kg, oral gavage) for 24 consecutive days. On the third day, mice were intravenously given a single dose of AD (10 mg/kg). At the end of the experiment, plasma and kidney samples were harvested to evaluate the therapeutic effects of THSG. The potential mechanisms of THSG in protecting against AD-induced cytotoxicity were examined using a real-time polymerase chain reaction, immunoblots, lactate dehydrogenase assay, and a cellular oxidized-thiol detection system in a mouse mesangial cell line. In this study, THSG showed concentration-dependent protective effects in ameliorating the progression of AD-induced FSGS. THSG suppressed albuminuria and hypercholesterolemia and reduced the status of lipid peroxidation in urine, plasma, and kidney tissue samples. Furthermore, THSG protected against podocyte damage, reduced renal fibrotic gene expressions, and alleviated the severity of glomerulosclerosis. Treatment of mouse mesangial cells with THSG induced nuclear factor erythroid-derived 2-like 2 (Nrf2) nuclear translocation, increased heme oxygenase-1 and NAD(P)H:quinone oxidoreductase (NQO)-1 gene expressions, and reduced cellular thiol oxidation and resistance to AD-induced cytotoxicity. Silencing Nrf2 and its repressor protein, Kelch-like ECH-associated protein 1 (Keap1), abolished these protective effects of THSG. In conclusion, THSG can play a protective role in ameliorating the progression of FSGS in a mouse model through activation of the Nrf2-Keap1 antioxidant pathway. Although a well-designed therapeutic study is needed, THSG may be applied to manage chronic kidney disease.

In Vitro and In Vivo Anti-Osteoarthritis Effects of 2,3,5,4'-Tetrahydroxystilbene-2-O-ß-d-Glucoside from Polygonum Multiflorum.

Polygonum multiflorum Thunb. is a traditional herbal medicine that is rich in polyphenols. The major compound, 2,3,5,4'-tetrahydroxystilbene-2-O-ß-d-glucoside (THSG) has many pharmacological activities, such as antioxidative and free radical-scavenging properties, and the abilities to reduce hyperlipidemia, prevent lipid peroxidation, and protect the cardiovascular system. In this study, the anti-osteoarthritis (OA) effects of THSG were explored using in vitro and in vivo models. THSG inhibited nitric oxide (NO) and prostaglandin E2 (PGE2) production and inducible NO synthase (iNOS) and cyclooxygenase-2 expressions by lipopolysaccharide-stimulated RAW 264.7 cells. On the other hand, THSG inhibited PGE2 production and iNOS and matrix metalloproteinase-13 expressions by interleukin-1ß-stimulated primary rat chondrocytes. Through a mono-iodoacetate-induced rat OA model assay, THSG reduced paw edema and improved the weight-bearing distribution. Therefore, THSG has anti-inflammatory activity and could be applied as a lead compound for the development as an OA drug.

Intermedin A, a New Labdane Diterpene Isolated from Alpinia intermedia, Prolonged the Survival Time of P-388D1 Tumor-Bearing CDF1 Mice.

Eight ethanolic extracts of indigenous Taiwanese plants of the genus Alpinia were tested for tumor cytotoxicity against AGS, Hep G2, HeLa, KB, and HL-60 cells. Among the 50 % and 95 % EtOH extracts of eight Alpinia species, the cytotoxic effects of Alpinia intermedia leaves were the strongest. When the leaf extract of A. intermedia was partitioned using n-hexane and aqueous solvents, the n-hexane layer showed a greater cytotoxic effect and could prolong the survival time of P-388D1 tumor-bearing CDF1 mice. Two new labdane diterpene derivatives, intermedin A (1) and intermedin B (2), and coronarin E (3) were isolated from the n-hexane layer of A. intermedia. Intermedin A induced apoptosis in HL-60 cells at 30 µg/mL and significantly prolonged the survival time of P-388D1 tumor-bearing CDF1 mice by 48.7 % at 20 mg/kg of body weight. We suggest that intermedin A is a major compound of A. intermedia and has a cytotoxic effect on HL-60 and P-388D1 cells.

Multiple Activities of Punica granatum Linne against Acne Vulgaris.

Acne is a common skin condition with sebum overproduction, hyperkeratosis, Propionibacterium acnes (P. acnes) and Staphylococcus aureus, and inflammation. Punica granatum (pomegranate) is well-known for its anti-inflammatory effects; however, few studies have discussed the anti-acne effects of pomegranate. In this study, we found that pomegranate extract (PG-E) significantly reduced P. acnes-induced edema in Wistar rat ears. Therefore, an evaluation platform using multiple pathogenic mechanisms of acne was established to explore the anti-acne effects of pomegranate. Results showed that PG-E inhibited bacterial growth and lipase activity. Through a bioguided-fractionation-isolation system, four hydrolysable tannins, punicalagin (1), punicalin (2), strictinin A (3), and granatin B (4), were isolated. Compounds 1 and 2 had greater anti-bacterial activities and anti-testosterone-induced HaCaT proliferative effects than the others. Compounds 1, 3, and 4 displayed lipase inhibitory effects. Compound 4 decreased cyclooxygenase-2 expression and downregulated prostaglandin E2 production in heat-killed P. acnes-treated RAW 246.7 cells. In conclusion, PG-E is abundant in hydrolysable tannins that display multiple anti-acne capacities, including anti-bacterial, anti-lipase, anti-keratinocyte proliferation, and anti-inflammatory actions. Hence, PG-E has great potential in the application of anti-acne and skin-care products, and punicalagin (1), the most effective component in PG-E, can be employed as a quality control marker.

Imperatorin exerts antiallergic effects in Th2-mediated allergic asthma via induction of IL-10-producing regulatory T cells by modulating the function of dendritic cells.

Imperatorin is a furanocoumarin compound which exists in many medicinal herbs and possesses various biological activities. Herein, we investigated the antiallergic effects of imperatorin in asthmatic mice and explored the immunomodulatory actions of imperatorin on immune cells. We used a murine model of ovalbumin (OVA)-induced asthma to evaluate the therapeutic potential of imperatorin. Additionally, bone marrow-derived dendritic cells (DCs; BMDCs) were used to clarify whether imperatorin exerts an antiallergic effect through altering the ability of DCs to regulate T cells. Oral administration of imperatorin to OVA-sensitized and -challenged mice decreased serum OVA-specific immunoglobulin E (IgE) production, attenuated the airway hyperresponsiveness (AHR), and alleviated airway inflammation in a dose-dependent manner. Notably, secretions of Th2 cytokines and chemokines were reduced, and numbers of interleukin (IL)-10-producing regulatory T cells (Tregs) increased in imperatorin-treated mice. Imperatorin inhibited proinflammatory cytokines and IL-12 production but enhanced IL-10 secretion by lipopolysaccharide (LPS)-stimulated BMDCs. Compared to fully mature DCs, imperatorin-treated DCs expressed high levels of the inducible costimulatory ligand (ICOSL) and Jagged1 molecules, and had the regulatory capacity to promote the generation of IL-10-producing CD4(+) T cells in vitro. Additionally, imperatorin directly suppressed activated CD4(+) T-cell proliferation and cytokine production. Imperatorin may possess therapeutic potential against Th2-mediated allergic asthma not only via stimulating DC induction of Tregs but also via direct inhibition of Th2 cell activation. These findings provide new insights into how imperatorin affects the Th2 immune response and the development of imperatorin as a Treg-type immunomodulatory agent to treat allergic asthma.

The clinical and biochemical characteristics associated with insulin resistance in non-obese young women.

AIM: The aim of this study was to investigate the biomarkers of insulin resistance in non-obese women. DESIGN: This was a retrospective study. PATIENTS: A total 229 non-obese women (Body mass index: BMI < 25) were evaluated. MAIN OUTCOME MEASURE(S): Serum levels of various androgens, cardiovascular risk and metabolic components. RESULTS: There were no significant differences in the prevalence of polycystic ovary syndrome (PCOS), hyperprolactinemia, or premature ovarian failure (POF) between the non-obese women with and without insulin resistance. Non-obese women with insulin had significantly higher serum thyroid stimulation hormone (TSH) levels and resistin and lower serum adiponectin levels than non-obese women without insulin resistance; however, the inflammatory biomarkers and serum androgen levels did not differ between the two groups. Furthermore, using step-wise multivariate regression analysis applied by the risk factors listed above, TSH was the only predictive factor for insulin resistance in non-obese reproductive-aged women. CONCLUSIONS: Thyroid function should play an important role in developing insulin resistance for non-obese women. Serum androgens and inflammation might not contribute to insulin resistance in these women.

2,3,5,4'-Tetrahydroxystilbene-2-O-ß-glucoside Isolated from Polygoni Multiflori Ameliorates the Development of Periodontitis.

Periodontitis, a chronic infection by periodontopathic bacteria, induces uncontrolled inflammation, which leads to periodontal tissue destruction. 2,3,5,4'-Tetrahydroxystilbene-2-O-beta-glucoside (THSG), a polyphenol extracted from Polygoni Multiflori, reportedly has anti-inflammatory properties. In this study, we investigated the mechanisms of THSG on the Porphyromonas gingivalis-induced inflammatory responses in human gingival fibroblasts and animal modeling of ligature-induced periodontitis. Human gingival fibroblast cells were treated with lipopolysaccharide (LPS) extracted from P. gingivalis in the presence of resveratrol or THSG to analyze the expression of TNF-α, IL-1ß, and IL-6 genes. Increased AMP-activated protein kinase (AMPK) activation and SirT1 expression were induced by THSG. Treatment of THSG decreased the expression of LPS-induced inflammatory cytokines, enhanced AMPK activation, and increased the expression of SirT1. In addition, it suppressed the activation of NF-κB when cells were stimulated with P. gingivalis LPS. The anti-inflammatory effect of THSG and P. Multiflori crude extracts was reproduced in ligature-induced periodontitis animal modeling. In conclusion, THSG inhibited the inflammatory responses of P. gingivalis-stimulated human gingival fibroblasts and ameliorated ligature-induced periodontitis in animal model.

Antinociceptive and Anti-Inflammatory Effects of Zerumbone against Mono-Iodoacetate-Induced Arthritis.

The fresh rhizome of Zingiber zerumbet Smith (Zingiberaceae) is used as a food flavoring and also serves as a folk medicine as an antipyretic and for analgesics in Taiwan. Zerumbone, a monocyclic sesquiterpene was isolated from the rhizome of Z. zerumbet and is the major active compound. In this study, the anti-inflammatory and antinociceptive effects of zerumbone on arthritis were explored using in vitro and in vivo models. Results showed that zerumbone inhibited inducible nitric oxide (NO) synthase (iNOS), cyclooxygenase (COX)-2 expressions, and NO and prostaglandin E2 (PGE2) production, but induced heme oxygenase (HO)-1 expression in a dose-dependent manner in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. When zerumbone was co-treated with an HO-1 inhibitor (tin protoporphyrin (SnPP)), the NO inhibitory effects of zerumbone were recovered. The above results suggest that zerumbone inhibited iNOS and COX-2 through induction of the HO-1 pathway. Moreover, matrix metalloproteinase (MMP)-13 and COX-2 expressions of interleukin (IL)-1ß-stimulated primary rat chondrocytes were inhibited by zerumbone. In an in vivo assay, an acetic acid-induced writhing response in mice was significantly reduced by treatment with zerumbone. Furthermore, zerumbone reduced paw edema and the pain response in a mono-iodoacetate (MIA)-induced rat osteoarthritis model. Therefore, we suggest that zerumbone possesses anti-inflammatory and antinociceptive effects which indicate zerumbone could be a potential candidate for osteoarthritis treatment.

Adjunctive traditional Chinese medicine therapy improves survival in patients with advanced breast cancer: a population-based study.

BACKGROUND: Traditional Chinese medicine (TCM) is one of the most common complementary and alternative medicines used in the treatment of patients with breast cancer. However, the clinical effect of TCM on survival, which is a major concern in these individuals, lacks evidence from large-scale clinical studies. METHODS: The authors used the Taiwan National Health Insurance Research Database to conduct a retrospective population-based cohort study of patients with advanced breast cancer between 2001 and 2010. The patients were separated into TCM users and nonusers, and Cox regression models were applied to determine the association between the use of TCM and patient survival. RESULTS: A total of 729 patients with advanced breast cancer receiving taxanes were included in the current study. Of this cohort, the mean age was 52.0 years; 115 patients were TCM users (15.8%) and 614 patients were TCM nonusers. The mean follow-up was 2.8 years, with 277 deaths reported to occur during the 10-year period. Multivariate analysis demonstrated that, compared with nonusers, the use of TCM was associated with a significantly decreased risk of all-cause mortality (adjusted hazards ratio [HR], 0.55 [95% confidence interval, 0.33-0.90] for TCM use of 30-180 days; adjusted HR, 0.46 [95% confidence interval, 0.27-0.78] for TCM use of >180 days). Among the frequently used TCMs, those found to be most effective (lowest HRs) in reducing mortality were Bai Hua She She Cao, Ban Zhi Lian, and Huang Qi. CONCLUSIONS: The results of the current observational study suggest that adjunctive TCM therapy may lower the risk of death in patients with advanced breast cancer. Future randomized controlled trials are required to validate these findings.

Pharmacological effects of "jutsu" (Atractylodis rhizome and Atractylodis lanceae rhizome) on 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI)-induced head twitch response in mice (I).

Hallucinations are a common non-motor symptom of Parkinson's disease and various forms of dementias. Yokukansan and Yokukansankachimpihange have attracted attention due to their effectiveness in the treatment of hallucinations of dementia. To clarify which component in these formulas contribute to the effects, at first, we focused on their differences in compositions to examine the pharmacological effects on the selective 5-HT2A/2C agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI)-induced head-twitch response (HTR) in mice that has been used as animal hallucination model. Results indicated that water extract of Byaku-jutsu (Atractylodes japonica) showed a stronger inhibitory effect on DOI-induced HTR than that of So-jutsu (A. lancea) corresponding to their major constituents of atractylenolide III and ß-eudesmol, and suggested that the major constituents should be active constituents contributing to the antihallucination effects of Byaku- and So-jutsu. Besides, the part B-C ring (butenolide) in atractylenolide III was found to be similar to the structure of serotonin and suggested that the B-C ring may partially play role in antagonistic activity against serotonin receptors. Thus, a novel, rational design of butenolide-related compounds may as potential lead compounds for new drug development. Analysis of the chemical components of Byaku- and So-jutsu and further study on their structure-activity relationships are currently in progress.

Chronopharmacology of diuresis via metabolic profiling and key biomarker discovery of the traditional Chinese prescription Ji-Ming-San using tandem mass spectrometry in rat models.

BACKGROUND: Ji-Ming-Shan (JMS) is a traditional prescription used for patients with rheumatism, tendons swelling, relief of foot pain, athlete's foot, diuresis, gout. Although many studies have investigated the active compounds in each herb, the functional mechanism behind its therapeutic effect remains unclear. STUDY DESIGN: Metabolic cages for sample collection. The serum components obtained from the experimental animals were analyzed using LC-MS/MS. Furthermore, cross-analysis using the software MetaboAnalyst and Venn diagrams were used to investigate chronopharmacology of JMS in the animal models. PURPOSE: The aim of this study is to analyze the diuretic effects of JMS and to explore their chronopharmacology involved in organ regulation through four-quarter periods from serum samples of rat models. METHODS: Metabolic cages were used for collecting the urine samples and PocketChem UA PU-4010, Fuji DRI-CHEM 800 were used to examine the urine biochemical parameters. The serum components were identified through ultra-performance liquid chromatography-quadrupole time-of-flight (UPLC-Q-TOF) with a new developed method. Cross analysis, Venn diagram, MetaboAnalyst were used to investigate the key biomarker and major metabolism route with the oral administration of the drug. RESULT: JMS significantly changed the 6 h urine volume with no observed kidney toxicity. Urine pH value ranges from 7.0 to 7.5. The chronopharmacology of JMS diuresis activity were 0-6 and 6-12 groups. UPLC-Q-TOF analyses identified 243 metabolites which were determined in positive mode and negative mode respectively. With cross analysis in the Venn diagram, one key biomarker naringenin-7-O-glucoside has been identified. Major metabolic pathways such as 1: Glycerophospholipid metabolism, 2: Primary bile acid biosynthesis, 3: Sphingolipid metabolism, 4: Riboflavin metabolism, 5: Linoleic acid metabolism, 6: Butanoate metabolism. CONCLUSION: JMS significantly changed the urine output of animals in the 0-6 and 6-12 groups. No change in urine pH was observed and also kidney toxicity. A new UPLC-Q-TOF method was developed for the detection of the metabolites of JMS after oral administration. The cross analysis with Venn diagram and identified the key biomarker of JMS namely naringenin-7-O-glucoside. The results showed that six major pathways are involved in the gastrointestinal system and the liver. This study demonstrated the capability of JMS prescription in the regulation of diuresis and identified a key biomarker that is responsible for its therapeutic effect.

Anti-osteoporotic effects of Yi Mai Jian on bone metabolism of ovariectomized rats.

Yi Mai Jian herbal formula (YMJ) is formulated with Eucommiae Folium, Astragali Radix, Ligustri Lucidi Fructus, and Elaeagnus Fructus to improve bone function in traditional Chinese medicine. The anti-osteoporotic effects of YMJ in bone metabolism were evaluated in ovariectomized (OVX) rats. The skeletal structure of the femur and vertebrae was analyzed after treating OVX rats with YMJ for 114 days. The results showed that YMJ significantly increased the bone mineral density (BMD) and trabecular number (Tb. N) of the femur and 5th lumbar vertebrae and reduced trabecular separation (Tb. Sp). Moreover, trabecular bone volume/total tissue volume (BV/TV), bone stiffness, and maximum femur load were significantly increased. The serum concentrations of NTX1 and PYD were significantly decreased. According to these results, YMJ could ameliorate osteoporosis in ovariectomized rats. Eucommiae Folium and Elaeagnus Fructus inhibited osteoclast differentiation, Ligustri Lucidi Fructus inhibited calcium reabsorption, Astragali Radix stimulated osteoblast proliferation, and Astragali Radix and Eucommiae Folium stimulated mineralization. Therefore, the combination of the four herbs into one formula, YMJ, could alleviate bone remodeling caused by low estrogen levels. We suggest that YMJ could be a healthy food candidate for preventing post-menopausal osteoporosis.

Immunomodulatory effects of Hedysarum polybotrys extract in mice macrophages, splenocytes and leucopenia.

Astragali Radix (Huang-Qi) is a popular herbal medicine commonly used as a constituent in tonic herbal preparations. Hedysarum polybotrys Handel-Mazzetti is one species used of Astragali Radix. In this study, the immunomodulatory properties of H. polybotrys were explored by LPS-activated and SNP-treated RAW 264.7 cells and splenocytes and, daunoblastina-induced leucopenia BALB/c mice. Formononetin was used as the bioactive marker to monitor the quality of the H. polybotrys extracts. H. polybotrys was extracted with hot-water and methanol, and MeOH extract partitioned with H2O (M-H) and ethyl acetate (M-EA) to yield four different fractions. M-EA had the highest formononetin and total proanthocyanidin content and showed stronger inhibitory effects on the production and expression of NO, PGE2, iNOS and COX-2 in LPS-activated RAW 264.7 cells and splenocytes than the other fractions. In addition, M-EA significantly stimulated the proliferation of LPS-activated RAW 264.7 cells and splenocytes, enhanced NO radicals scavenging and attenuated NO-induced cytotoxicity.  Furthermore, M-EA also significantly increased the rate of recovery of white blood cells level in daunoblastina-induced leucopenia mice. These evidences suggest that this traditional Qi-tonifying herb has potential effects in clinical conditions when immune-enhancing and anti-inflammatory effect is desired.

Cinnamic aldehyde, an anti-inflammatory component in Du-Huo-Ji-Sheng-Tang, ameliorates arthritis in II collagenase and monosodium iodoacetate induced osteoarthritis rat models.

Background and aim: Du-Huo-Ji-Sheng-Tang (DHJST) is a Chinese herbal formula used for arthralgia and arthritis treatment clinically. This study aims to evaluate the joint-protecting efficacy of DHJST and to identify the active constituents as the evaluation marker. Experimental procedure: DHJST can be categorized into three recipes: Blood-tonifying-herbs Si-Wu-Tang (SWT), Wind-dampness-dispelling-herbs (WDH) and Qi-tonifying-herbs (TH). All formulas were used to explore the joint-protecting efficacies. Results and conclusion: s: Firstly, DHJST could decrease the arthritis progression in the monosodium-iodoacetate-induced rat and cure arthritis in the type II collagenase-induced rat. Further, in lipopolysaccharide-stimulated RAW 264.7 cells, DHJST, TH and Cinnamomum cassia (CC), an ingredient in TH, were the most potent nitric oxide (NO) and prostaglandin E2 (PGE2) inhibitors. The major components, cinnamic aldehyde, showed the strongest NO and PGE2 inhibition. Up-regulated inducible NO synthase (iNOS) and cyclooxygenase-2 were inhibited by DHJST, TH, CC, and cinnamic aldehyde. In interleukin-1ß-stimulated primary chondrocytes, upregulated iNOS was inhibited by DHJST, TH, Cinnamomum cassia, and cinnamic aldehyde. Upregulated matrix metalloprotease-13 was only inhibited by DHJST and TH and Eucommia ulmoides (EU) extract. Results suggest that DHJST presented joint-protective and cure arthritis effects. TH presented equal joint-protective effects as DHJST. The major anti-inflammatory ingredient in TH was Cinnamomum cassia in TH. And cinnamic aldehyde was the potent anti-inflammatory active compound in Cinnamomum cassia. Therefore, this study may facilitate the modern use of DHJST with TH as a simplified version but equally effective anti-osteoarthritic agents with cinnamic aldehyde as a quality control marker of DHJST and TH in osteoarthritis prevention or treatment.