Search

DA-9801, a plant-based drug used for the treatment of diabetic neuropathy, is known to improve angiotensin II (Ang II)-induced vascular endothelial cell dysfunction. However, the underlying mechanism is not fully understood. We aimed to determine whether the protective effect of DA-9801 against Ang II-induced endothelial cell dysfunction was mediated via inhibition of endothelial cell inflammation and apoptosis. Ang II-induced oxidative stress was attenuated by pretreatment of human dermal microvascular endothelial cells (HDMECs) with DA-9801. This prevented the Ang II-induced upregulation of NAD(P)H oxidase (the NOX4 and p22phox subunits) and reactive oxygen species. Further, pretreatment of HDMECs with DA-9801 ameliorated Ang II-mediated nuclear factor kappa B activity via prevention of the upregulation of extracellular signal-regulated kinase and p38 mitogen-activated protein kinase. It also decreased the Ang II-stimulated increase in inducible nitric oxide synthase (NOS) and decreased endothelial NOS protein expression. DA-9801 decreased Ang II-induced upregulation of intercellular adhesion molecule 1, vascular adhesion molecule, and E-selectin in HDMECs. Moreover, TUNEL and annexin V-FITC fluorescence staining for apoptosis and the activities of caspases 9, 7, and 3 decreased in HDMECs pretreated with DA-9801, indicating that the drug enhanced anti-apoptotic pathways. Thus, DA-9801 modulated Ang II-induced endothelial cell dysfunction via inflammatory and apoptotic pathways. more...

Nerve growth factor (NGF) is a neurotrophin that plays a critical role in mammalian learning and memory functions. NGF also regulates neuronal cell differentiation and neurite outgrowth by activating ERK/CREB signaling. This present study examined the effects of a standardized Dioscorea extract (DA-9801), which is composed of Dioscorea japonica Thunb and Dioscorea nipponica Makino on memory function via its NGF-potentiating activities using an in vitro and in vivo paradigm. Cells were incubated with or without different concentrations of DA-9801 (10, 25, and 50 μg/ml) extract for 24 h. The cultured conditioned medium from C6 glioma cells was used for NGF production assay, and neurite length in N2a cells was measured after every 2 h. Mice were orally treated with DA-9801 (10 and 100 mg/kg/day) once daily for 7 days. They were subjected to passive avoidance test to evaluate memory functions. The question of whether DA-9801 induced NGF synthesis was assessed by measuring the levels of NGF in the mouse cortical and hippocampal tissues. Hippocampal cell differentiation and NGF-mediated ERK/CREB signaling were evaluated by performing immunohistochemical analysis using BrdU, ki67, DCX, phosphorylated ERK and CREB in the mouse hippocampus. DA-9801 treatment increased the NGF contents and neurite length, respectively. Mice with DA-9801 administration showed memory enhancement in the passive avoidance test. DA-9801 also increased newborn cell differentiation, neurite length, NGF secretion, and ERK/CREB phosphorylation in the mouse hippocampus. These results suggest that DA-9801 treatment could improve memory function by inducing hippocampal NGF synthesis and ERK/CREB signaling. [ABSTRACT FROM AUTHOR] more...

Neurodegenerative diseases, including Alzheimer’s disease, are prevalent in diabetic patients. DA-9801 is a standardized extract of Dioscorea japonica Thunb. and Dioscorea nipponica Makino, which has been shown to reduce blood glucose levels and exert protective effects against diabetic peripheral neuropathy. Here, we aimed to investigate whether DA-9801 could protect hyperglycemia-induced central neuropathy and to elucidate the underlying mechanisms. DA-9801 protected rat cortical primary neurons against hyperglycemia-induced neurotoxicity by lowering the levels of reactive oxygen species. In addition, we identified the genes whose expression in high glucose-treated primary neurons was significantly altered by DA-9801 treatment. Moreover, the major saponins of DA-9801, dioscin and protodioscin, protected neurons from hyperglycemia-induced toxicity, suggesting that these may be the most important active neuroprotective components of DA-9801. Our results showed that DA-9801 protects neurons derived from the central nervous system and that it may be useful to treat and delay the development of neurodegenerative diseases in diabetic patients. more...

Graphical abstract Highlights • DA-9801 protects rat primary cortical neurons from hyperglycemia-induced cytotoxicity. • DA-9801 reduces reactive oxidative species caused by high glucose treatment. • Microarray analysis reveals that DA-9801 regulates gene expression. • Dioscin and Protodioscin, the major saponins of DA9801, protect cortical neurons. Abstract Neurodegenerative diseases, including Alzheimer's disease, are prevalent in diabetic patients. DA-9801 is a standardized extract of Dioscorea japonica Thunb. and Dioscorea nipponica Makino, which has been shown to reduce blood glucose levels and exert protective effects against diabetic peripheral neuropathy. Here, we aimed to investigate whether DA-9801 could protect hyperglycemia-induced central neuropathy and to elucidate the underlying mechanisms. DA-9801 protected rat cortical primary neurons against hyperglycemia-induced neurotoxicity by lowering the levels of reactive oxygen species. In addition, we identified the genes whose expression in high glucose-treated primary neurons was significantly altered by DA-9801 treatment. Moreover, the major saponins of DA-9801, dioscin and protodioscin, protected neurons from hyperglycemia-induced toxicity, suggesting that these may be the most important active neuroprotective components of DA-9801. Our results showed that DA-9801 protects neurons derived from the central nervous system and that it may be useful to treat and delay the development of neurodegenerative diseases in diabetic patients. [ABSTRACT FROM AUTHOR] more...

Introduction DA-9801, a standardised 50% aqueous ethanolic extract of a mixture of Dioscorea japonica and D. nipponica , is a botanical drug candidate for the treatment of diabetic neuropathy, which finished its US phase II clinical trials recently. An advanced quality control method is needed for further development of DA-9801, considering its high contents of both primary and secondary metabolites. Objective Development of a quality assessment strategy for DA-9801, based on the combination of UHPLC-QTOF/MS, HPLC-ELSD, and 1H-NMR spectroscopy. Methods The method was developed and tested with 15 batch products of DA-9801. The steroidal saponins of DA-9801 were tentatively identified by UHPLC-QTOF/MS and were quantified with the validated HPLC-ELSD method. Primary metabolites of DA-9801 were identified and profiled using 1H-NMR spectrometry. The batch-to-batch equivalence of DA-9801 was tested with the 1H-NMR spectra using spectral binning, correlation analysis, and principal component analysis. Results Six major saponins of DA-9801 were tentatively identified by UHPLC-QTOF/MS. Among them, protodioscin and dioscin were quantified by the validated HPLC-ELSD method. Twenty-six metabolites were identified in 1H-NMR spectra. The similarity between DA-9801 batches could be evaluated with the NMR spectra of DA-9801. The 1H-NMR method also revealed that two Dioscorea species contributed distinct amino acids to the contents of DA-9801. Conclusion This study validates the effectiveness of UHPLC-QTOF/MS, HPLC-ELSD, and 1H NMR-combined method for quality control of DA-9801 and its crude materials. Copyright © 2016 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR] more...

Background Drug transporters play important roles in the absorption, distribution, and elimination of drugs and thereby, modulate drug efficacy and toxicity. With a growing use of poly pharmacy, concurrent administration of herbal extracts that modulate transporter activities with drugs can cause serious adverse reactions. Therefore, prediction and evaluation of drug-drug interaction potential is important in the clinic and in the drug development process. DA-9801, comprising a mixed extract of Dioscoreae rhizoma and Dioscorea nipponica Makino, is a new standardized extract currently being evaluated for diabetic peripheral neuropathy in a phase II clinical study. Method The inhibitory effects of DA-9801 on the transport functions of organic cation transporter (OCT)1, OCT2, organic anion transporter (OAT)1, OAT3, organic anion transporting polypeptide (OATP)1B1, OATP1B3, P-glycoprotein (P-gp), and breast cancer resistance protein (BCRP) were investigated in HEK293 or LLC-PK1 cells. The effects of DA-9801 on the pharmacokinetics of relevant substrate drugs of these transporters were also examined in vivo in rats. Results DA-9801 inhibited the in vitro transport activities of OCT1, OCT2, OAT3, and OATP1B1, with IC50 values of 106, 174, 48.1, and 273 μg/mL, respectively, while the other transporters were not inhibited by 300 μg/mL DA-9801. To investigate whether this inhibitory effect of DA-9801 on OCT1, OCT2, and OAT3 could change the pharmacokinetics of their substrates in vivo, we measured the pharmacokinetics of cimetidine, a substrate for OCT1, OCT2, and OAT3, and of furosemide, a substrate for OAT1 and OAT3, by co-administration of DA-9801 at a single oral dose of 1,000 mg/kg. Pre-dose of DA-9801 5 min or 2 h prior to cimetidine administration decreased the Cmax of cimetidine in rats. However, DA-9801 did not affect the elimination parameters such as half-life, clearance, or amount excreted in the urine, suggesting that it did not inhibit elimination process of cimetidine, which is governed by OCT1, OCT2, and OAT3. Moreover, DA-9801 did not affect the pharmacokinetic characteristics of furosemide, as evidenced by its unchanged pharmacokinetic parameters. Conclusion Inhibitory effects of DA-9801 on OCT1, OCT2, and OAT3 observed in vitro may not necessarily translate into in vivo herb-drug interactions in rats even at its maximum effective dose. [ABSTRACT FROM AUTHOR] more...

DA-9801, the mixture extract of Dioscoreae rhizoma and Dioscorea nipponica Makino, is a new herbal drug currently being evaluated in a phase II clinical study for the treatment of diabetic peripheral neuropathy in Korea. The inhibitory potentials of DA-9801, D. rhizoma extract, D. nipponica Makino extract, and dioscin, an active component of DA-9801, on eight human cytochrome P450 (CYP) enzymes and four UDP-glucuronosyltransferase (UGT) enzymes were investigated in human liver microsomes using liquid chromatography-tandem mass spectrometry. DA-9801 showed slight inhibition of CYP1A2, CYP2C8, UGT1A1, and UGT1A9 enzyme activities with IC values of 396.4, 449.9, 226.0, and 408.8 μg/mL, respectively. D. rhizoma extract showed negligible inhibition of CYP and UGT activities, but D. nipponica extract slightly inhibited CYP1A2, CYP2C8, CYP2C9, UGT1A1, and UGT1A9 activities with IC values of 264.2, 237.1, 206.8, 302.4, and 383.1 μg/mL, respectively. DA-9801 showed volume per dose index values of 0.44-0.88 L for a 200-mg dose, suggesting that they may not cause the inhibition of the metabolism of CYP1A2, CYP2C8, UGT1A1, and UGT1A9-catalyzed drugs in humans. These results suggest that the administration of DA-9801 in human may not cause clinically relevant inhibition of these enzymes. [ABSTRACT FROM AUTHOR] more...

Abstract: Ethnopharmacological relevance: Dioscorea japonica Thunb. has been traditionally used to treat polyuria and diabetes in Korea. Aim of the study: We previously report the effects of Dioscorea japonica Thunb. extract on glucose control, NGF induction, and neuroprotection in a rodent diabetic model. Since the most potent fraction, DA-9801, was identified from a mixture of Dioscorea japonica Thunb. (DJ) and Dioscorea nipponica Makino (DN) following bioactivity-guided fractionation, here, we investigated the potential mechanism of the extract activity against diabetic peripheral neuropathy (DPN). Materials and methods: A 1:3 mixture of DJ and DN was extracted with ethanol (DA-9801) and further fractionated into an ethylacetate-soluble fraction (DA-9801E). Effects of these extracts on neurite outgrowth were measured in PC-12 cells and DRG neurons. Effects on cell viability and TrkA phosphorylation were evaluated in PC-12 cells. NGF induction effect was determined in primary Schwann cells as well as IMS32 cells (immortalized Schwann cells). Results: No cytotoxicity was observed in PC-12 cells at the concentration below 500μg/ml of either DA-9801 or DA-9801E. DA-9801 and DA-9801E at 100μg/ml and 10μg/ml, respectively, showed a significant effect on neurite outgrowth in PC-12 cells and DRG neurons in the presence of or absence a low concentration of NGF (2ng/ml). The Trk-A phosphorylation effect of DA9801 was confirmed in PC-12 cells. An NGF induction effect of these extracts was not detected in either IMS-32 cells, or primary Schwann cells. Conclusions: The NGF agonistic activity of DA-9801 and DA-9801E was demonstrated, which may contribute to their neuroprotective effect against DPN. Studies of the detailed mechanism of these extracts as well as identification of the active components are warranted for the development of an anti-DPN drug from DJ and DN. [Copyright &y& Elsevier] more...

Introduction: DA-9801, a standardised 50% aqueous ethanolic extract of a mixture of Dioscorea japonica and D. nipponica, is a botanical drug candidate for the treatment of diabetic neuropathy, which finished its US phase II clinical trials recently. An advanced quality control method is needed for further development of DA-9801, considering its high contents of both primary and secondary metabolites., Objective: Development of a quality assessment strategy for DA-9801, based on the combination of UHPLC-QTOF/MS, HPLC-ELSD, and 1 H-NMR spectroscopy., Methods: The method was developed and tested with 15 batch products of DA-9801. The steroidal saponins of DA-9801 were tentatively identified by UHPLC-QTOF/MS and were quantified with the validated HPLC-ELSD method. Primary metabolites of DA-9801 were identified and profiled using 1 H-NMR spectrometry. The batch-to-batch equivalence of DA-9801 was tested with the 1 H-NMR spectra using spectral binning, correlation analysis, and principal component analysis., Results: Six major saponins of DA-9801 were tentatively identified by UHPLC-QTOF/MS. Among them, protodioscin and dioscin were quantified by the validated HPLC-ELSD method. Twenty-six metabolites were identified in 1 H-NMR spectra. The similarity between DA-9801 batches could be evaluated with the NMR spectra of DA-9801. The 1 H-NMR method also revealed that two Dioscorea species contributed distinct amino acids to the contents of DA-9801., Conclusion: This study validates the effectiveness of UHPLC-QTOF/MS, HPLC-ELSD, and 1 H NMR-combined method for quality control of DA-9801 and its crude materials. Copyright © 2016 John Wiley & Sons, Ltd., (Copyright © 2016 John Wiley & Sons, Ltd.) more...

Nerve growth factor (NGF)-induced neuronal regeneration has emerged as a strategy to treat neuronal degeneration-associated disorders. However, direct NGF administration is limited by the occurrence of adverse effects at high doses of NGF. Therefore, development of a therapeutic strategy to promote the NGF trophic effect is required. In view of the lack of understanding of the mechanism for potentiating the NGF effect, this study investigated molecular targets of DA-9801, a well-standardized Dioscorea rhizome extract, which has a promoting effect on NGF. An increase in intracellular calcium ion level was induced by DA-9801, and chelation of extracellular calcium ions with ethylene-bis(oxyethylenenitrilo)tetraacetic acid (EGTA) suppressed the potentiating effect of DA-9801 on NGF-induced neurite outgrowth. In addition, EGTA treatment reduced the DA-9801-induced phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2), the major mediators of neurite outgrowth. To find which calcium ion-permeable channel contributes to the calcium ion influx induced by DA-9801, we treated PC12 cells with various inhibitors of calcium ion-permeable channels. NF449, a P2X1 receptor selective antagonist, significantly abolished the potentiating effect of DA-9801 on NGF-induced neurite outgrowth and abrogated the DA-9801-induced ERK1/2 phosphorylation. In addition, transfection with siRNA of P2X1 receptor significantly reduced the DA-9801-enhanced neurite outgrowth. In conclusion, calcium ion influx through P2X1 receptor mediated the promoting effect of DA-9801 on NGF-induced neurite outgrowth via ERK1/2 phosphorylation. more...