Antioxidant and Inhibitory Activities of Filipendula glaberrima Leaf Constituents against HMG-CoA Reductase and Macrophage Foam Cell Formation.

In our search for bioactive components, various chromatographic separations of the organic fractions from Filipendula glaberrima leaves led to the isolation of a new ellagitannin and a triterpenoid, along with 26 known compounds. The structures of the isolates were determined based on their spectroscopic properties and chemical evidence, which were then evaluated for their antioxidant activities, inhibitory activities on 3-hydroxy-3-methylglutaryl-coenzyme A reductase, and foam cell formation in THP-1 cells to prevent atherosclerosis. Rugosin B methyl ester (1) showed the best HMG-CoA reductase inhibition and significantly reduced ox-low-density lipoprotein-induced THP-1 macrophage-derived foam cell formation at 25 µM. In addition, no cytotoxicity was observed in THP-1 cells at 50 µg/mL of all extracts in the macrophage foam cell formation assay. Therefore, F. glaberrima extract containing 1 is promising in the development of dietary supplements due to its potential behavior as a novel source of nutrients for preventing and treating atherosclerosis.

Hydrangenol, an active constituent of Hydrangea serrata (Thunb.) Ser., ameliorates colitis through suppression of macrophage-mediated inflammation in dextran sulfate sodium-treated mice.

Ulcerative colitis (UC) is a chronic disease of the colon characterized by mucosal damage and relapsing gastrointestinal inflammation. Hydrangea serrata (Thunb.) Ser. and its bioactive compound, hydrangenol, are reported to have anti-inflammatory effects, but few studies have investigated the effects of hydrangenol in colitis. In the present study, we evaluated for the first time the anti-colitic effects and molecular mechanisms of hydrangenol in a dextran sodium sulfate (DSS)-induced mouse colitis model. To investigate the anti-colitic effects of hydrangenol, DSS-induced colitis mice, HT-29 colonic epithelial cells treated with supernatant from LPS-inflamed THP-1 macrophages, and LPS-induced RAW264.7 macrophages were used. In addition, to clarify the molecular mechanisms of this study, quantitative real time-PCR, western blot analysis, TUNEL assay, and annexin V-FITC/PI double staining analysis were conducted. Oral administration of hydrangenol (15 or 30 mg kg-1) significantly alleviated DSS-induced colitis by preventing DAI scores, shortening colon length, and colonic structural damage. F4/80+ macrophage numbers in mesenteric lymph nodes and macrophage infiltration in colonic tissues were significantly suppressed following hydrangenol treatment in DSS-exposed mice. Hydrangenol significantly attenuated DSS-induced destruction of the colonic epithelial cell layer through regulation of pro-caspase-3, occludin, and claudin-1 protein expression. Moreover, hydrangenol ameliorated abnormal tight junction protein expression and apoptosis in HT-29 colonic epithelial cells treated with supernatant from LPS-inflamed THP-1 macrophages. Hydrangenol suppressed the expression of pro-inflammatory mediators, such as iNOS, COX-2, TNF-α, IL-6, and IL-1ß through NF-κB, AP-1, and STAT1/3 inactivation in DSS-induced colon tissue and LPS-induced RAW264.7 macrophages. Taken together, our findings suggest that hydrangenol recovers the tight junction proteins and down-regulates the expression of the pro-inflammatory mediators by interfering with the macrophage infiltration in DSS-induced colitis. Our study provides compelling evidence that hydrangenol may be a candidate for inflammatory bowel disease therapy.

Aster glehni F. Schmidt Extract Modulates the Activities of HMG-CoA Reductase and Fatty Acid Synthase.

Aster glehni F. Schmidt (AG), is a natural product known to have anti-obesity effects, but the mechanism underlying these effects is not well documented. We hypothesized that AG may have inhibitory effects on enzymes related to lipid accumulation. Herein, AG fractions were tested against HMG-CoA reductase (HMGR) and fatty acid synthase (FAS), two important enzymes involved in cholesterol and fatty acid synthesis, respectively. We found that dicaffeoylquinic acid (DCQA) methyl esters present in AG are largely responsible for the inhibition of HMGR and FAS. Since free DCQA is a major form present in AG, we demonstrated that a simple methylation of the AG extract could increase the overall inhibitory effects against those enzymes. Through this simple process, we were able to increase the inhibitory effect by 150%. We believe that our processed AG effectively modulates the HMGR and FAS activities, providing promising therapeutic potential for cholesterol- and lipid-lowering effects.

Hydrangenol Isolated from the Leaves of Hydrangea serrata Attenuates Wrinkle Formation and Repairs Skin Moisture in UVB-Irradiated Hairless Mice.

Our previous study showed that hydrangenol isolated from Hydrangea serrata leaves exerts antiphotoaging activity in vitro. In this study, we determined its antiphotoaging effect in UVB-irradiated HR-1 hairless mice. We evaluated wrinkle formation, skin thickness, histological characteristics, and mRNA and protein expression using qRT-PCR and Western blot analysis in dorsal skins. Hydrangenol mitigated wrinkle formation, dorsal thickness, dehydration, and collagen degradation. Hydrangenol increased the expression of involucrin, filaggrin, and aquaporin-3 (AQP3) as well as hyaluronic acid (HA) production via hyaluronidase (HYAL)-1/-2 downregulation. Consistent with the recovery of collagen composition, the expression of Pro-COL1A1 was increased by hydrangenol. Matrix metalloproteinase (MMP)-1/-3, cyclooxygenase-2 (COX-2), and interleukin-6 (IL-6) expression was reduced by hydrangenol. Hydrangenol attenuated the phosphorylation of mitogen-activated protein kinases (MAPKs) including ERK and p38, activator protein 1 (AP-1) subunit, and signal transduction and activation of transcription 1 (STAT1). Hydrangenol upregulated the expression of nuclear factor-E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), NAD(P)H quinone dehydrogenase 1 (NQO-1), glutamate cysteine ligase modifier subunit (GCLM), and glutamate cysteine ligase catalysis subunit (GCLC). Taken together, our data suggest that hydrangenol can prevent wrinkle formation by reducing MMP and inflammatory cytokine levels and increasing the expression of moisturizing factors and antioxidant genes.

Aster glehni Extract Containing Caffeoylquinic Compounds Protects Human Keratinocytes through the TRPV4-PPARδ-AMPK Pathway.

Aster glehni (AG) has been used in cooking and as a medicine to treat various diseases for over hundreds of years in Korea. To speculate the protective effects of AG on skin barrier, we estimated the protein levels of biomarkers related to skin barrier protection in human keratinocytes, HaCaT cells treated with sodium dodecyl sulfate (SDS), or 2,4-dinitrochlorobenzene (DNCB). The protein levels for keratin, involucrin, defensin, tumor necrosis factor alpha (TNFα), peroxisome proliferator-activated receptor delta (PPARδ), 5' adenosine monophosphate-activated protein kinase (AMPK), serine palmitoyltransferase long chain base subunit 2 (SPTLC2), and transient receptor potential cation channel subfamily V member 4 (TRPV4) were evaluated using western blotting or immunocytochemistry in HaCaT cells. AG extract increased the protein levels of PPARδ, phosphorylated AMPK, SPTLC2, keratin, involucrin, and defensin compared to the SDS or DNCB control group. However, TNFα expression increased by SDS or DNCB was decreased with AG extract. The order of action of each regulatory biomarker in AG pathway was identified TRPV4→PPARδ→AMPK from antagonist and siRNA treatment studies. AG can ameliorate the injury of keratinocytes caused by SDS or DNCB through the sequential regulation of TRPV4→PPARδ→AMPK pathway.

Caffeoylquinic Acid-Rich Extract of Aster glehni F. Schmidt Ameliorates Nonalcoholic Fatty Liver through the Regulation of PPARδ and Adiponectin in ApoE KO Mice.

Aster glehni is well known for its therapeutic properties. This study was performed to investigate the effects of A. glehni on nonalcoholic fatty liver disease (NAFLD) in atherosclerotic condition, by determining the levels of biomarkers related to lipid metabolism and inflammation in serum, liver, and adipose tissue. Body and abdominal adipose tissue weights and serum triglyceride level decreased in all groups treated with A. glehni. Serum adiponectin concentration and protein levels of peroxisome proliferator-activated receptor δ, 5' adenosine monophosphate-activated protein kinase, acetyl-CoA carboxylase, superoxide dismutase, and PPARγ coactivator 1-alpha in liver tissues increased in the groups treated with A. glehni. Conversely, protein levels of ATP citrate lyase, fatty acid synthase, tumor necrosis factor α, and 3-hydroxy-3-methylglutaryl-CoA reductase and the concentrations of interleukin 6 and reactive oxygen species decreased upon A. glehni. Triglyceride concentration in the liver was lower in mice treated with A. glehni than in control mice. Lipid accumulation in HepG2 and 3T3-L1 cells decreased upon A. glehni treatment; this effect was suppressed in the presence of the PPARδ antagonist, GSK0660. Our findings suggest that A. glehni extracts may ameliorate NAFLD through regulation of PPARδ, adiponectin, and the related subgenes.

Inhibitory effect of moschamine isolated from Carthamus tinctorius on LPS-induced inflammatory mediators via AP-1 and STAT1/3 inactivation in RAW 264.7 macrophages.

Seeds of Carthamus tinctorius L. (Compositae) have been used in Korean traditional medicines for the treatment of cardiovascular and bone diseases. In this study, we investigated the anti-inflammatory effects of known serotonin derivatives (1-9) isolated from the ethyl acetate (EtOAc) soluble fraction from the seeds of C. tinctorius. Compound 2, identified as moschamine, most potently inhibited lipopolysaccharide (LPS)-induced production of prostaglandin E2 (PGE2) and nitric oxide (NO) in RAW 264.7 macrophages. Moschamine concentration-dependently inhibited LPS-induced PGE2 and NO production in RAW 264.7 macrophages. Consistent with these findings, moschamine suppressed the protein and mRNA levels of cyclooxygenase-2 (COX-2), microsomal prostaglandin E2 synthase (mPGES)-1, and inducible NO synthase (iNOS), interleukin (IL)-6, and IL-1ß. In addition, pretreatment of moschamine significantly inhibited LPS-stimulated the transcriptional activity of activator protein-1 (AP-1) and the phosphorylation of signal transducer and activator of transcription (STAT)1/3 in RAW 264.7 macrophages. Moreover, moschamine inhibited LPS-induced the phosphorylation of p38 mitogen-activated protein kinase (p38) and extracellular signal-regulated kinase (ERK), but it had no effect on c-Jun N-terminal kinase (JNK). These results suggest that the mechanism of anti-inflammatory activity of moschamine is associated with the downregulation of COX-2, mPGES-1, iNOS, IL-6, and IL-1ß expression through the suppression of AP-1 and STAT1/3 activation in LPS-induced RAW 264.7 macrophages.

Synthesis and biological evaluation of fluoro-substituted 3,4-dihydroquinazoline derivatives for cytotoxic and analgesic effects.

As a bioisosteric strategy to overcome the poor metabolic stability of lead compound KYS05090S, a series of new fluoro-substituted 3,4-dihydroquinazoline derivatives was prepared and evaluated for T-type calcium channel (Cav3.2) block, cytotoxic effects and liver microsomal stability. Among them, compound 8h (KCP10068F) containing 4-fluorobenzyl amide and 4-cyclohexylphenyl ring potently blocked Cav3.2 currents (>90% inhibition) at 10µM concentration and exhibited cytotoxic effect (IC50=5.9µM) in A549 non-small cell lung cancer cells that was comparable to KYS05090S. Furthermore, 8h showed approximately a 2-fold increase in liver metabolic stability in rat and human species compared to KYS05090S. Based on these overall results, 8h (KCP10068F) may therefore represent a good backup compound for KYS05090S for further biological investigations as novel cytotoxic agent. In addition, compound 8g (KCP10067F) was found to partially protect from inflammatory pain via a blockade of Cav3.2 channels.

6'-O-Caffeoyldihydrosyringin isolated from Aster glehni suppresses lipopolysaccharide-induced iNOS, COX-2, TNF-α, IL-1ß and IL-6 expression via NF-κB and AP-1 inactivation in RAW 264.7 macrophages.

Previously, we found that ethyl acetate extract fraction of Aster glehni exhibited anti-hyperuricemic effects in animal models and also five new caffeoylglucoside derivatives were isolated from this fraction. In this work, we evaluated the anti-inflammatory effects of these caffeoylglucoside derivatives and found that 6'-O-caffeoyldihydrosyringin (2, CDS) most potently inhibited the LPS-induced production of nitric oxide (NO) and prostaglandin E2 (PGE2) in RAW 264.7 macrophages. In addition, CDS was found to concentration-dependently reduce the production of NO, PGE2, and the pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1ß (IL-1ß) induced by LPS in macrophages. Consistent with these observations, CDS concentration-dependently inhibited LPS-induced inducible nitric oxide synthase (iNOS) and cyclooxidase-2 (COX-2) expression at the protein level and also iNOS, COX-2, TNF-α, and IL-6, IL-1ß expression at the mRNA level. Furthermore, CDS suppressed the LPS-induced transcriptional activities of nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) as well as the phosphorylation of p65 and c-Fos. Taken together, these results suggest that the anti-inflammatory effect of CDS is associated with the downregulation of iNOS, COX-2, TNF-α, IL-1ß, and IL-6 expression via the negative regulation of NF-κB and AP-1 activation in LPS-induced RAW 264.7 macrophages.

Opuntia ficus-indica seed attenuates hepatic steatosis and promotes M2 macrophage polarization in high-fat diet-fed mice.

Opuntia ficus-indica (L.) is a popular edible plant that possesses considerable nutritional value and exhibits diverse biological actions including anti-inflammatory and antidiabetic activities. In this study, we hypothesized that DWJ504, an extract of O ficus-indica seed, would ameliorate hepatic steatosis and inflammation by regulating hepatic de novo lipogenesis and macrophage polarization against experimental nonalcoholic steatohepatitis. Mice were fed a normal diet or a high-fat diet (HFD) for 10 weeks. DWJ504 (250, 500, and 1000 mg/kg) or vehicle (0.5% carboxymethyl cellulose) were orally administered for the last 4 weeks of the 10-week HFD feeding period. DWJ504 treatment remarkably attenuated HFD-induced increases in hepatic lipid content and hepatocellular damage. DWJ504 attenuated increases in sterol regulatory element-binding protein 1 and carbohydrate-responsive element-binding protein expression and a decrease in carnitine palmitoyltransferase 1A. Although DWJ504 augmented peroxisome proliferator-activated receptor α protein expression, it attenuated peroxisome proliferator-activated receptor γ expression. Moreover, DWJ504 promoted hepatic M2 macrophage polarization as indicated by attenuation of the M1 marker genes and enhancement of M2 marker genes. Finally, DWJ504 attenuated expression of toll-like receptor 4, nuclear factor κB, tumor necrosis factor α, interleukin 6, TIR-domain-containing adapter-inducing interferon ß, and interferon ß levels. Our results demonstrate that DWJ504 prevented intrahepatic lipid accumulation, induced M2 macrophage polarization, and suppressed the toll-like receptor 4-mediated inflammatory signaling pathway. Thus, DWJ504 has therapeutic potential in the prevention of nonalcoholic fatty liver disease.

In vitro synergistic anticancer activity of the combination of T-type calcium channel blocker and chemotherapeutic agent in A549 cells.

As a result of our continuous research, new 3,4-dihydroquinazoline derivative containing ureido group, KCP10043F was synthesized and evaluated for T-type Ca(2+) channel (Cav3.1) blockade, cytotoxicity, and cell cycle arrest against human non-small cell lung (A549) cells. KCP10043F showed both weaker T-type Ca(2+) channel blocking activity and less cytotoxicity against A549 cells than parent compound KYS05090S [4-(benzylcarbamoylmethyl)-3-(4-biphenylyl)-2-(N,N',N'-trimethyl-1,5-pentanediamino)-3,4-dihydroquinazoline 2 hydrochloride], but it exhibited more potent G1-phase arrest than KYS05090S in A549 cells. This was found to be accompanied by the downregulations of cyclin-dependent kinase (CDK) 2, CDK4, CDK6, cyclin D2, cyclin D3, and cyclin E at the protein levels. However, p27(KIP1) as a CDK inhibitor was gradually upregulated at the protein levels and increased recruitment to CDK2, CDK4 and CDK6 after KCP10043F treatment. Based on the strong G1-phase cell cycle arrest of KCP10043F in A549 cells, the combination of KCP10043F with etoposide (or cisplatin) resulted in a synergistic cell death (combination index=0.2-0.8) via the induction of apoptosis compared with either agent alone. Taken together with these overall results and the favorable in vitro ADME (absorption, distribution, metabolism, and excretion) profiles of KCP10043F, therefore, it could be used as a potential agent for the combination therapy on human lung cancer.

Synthesis of (2-amino)ethyl derivatives of quercetin 3-O-methyl ether and their antioxidant and neuroprotective effects.

Reactive oxygen species have been implicated in several diseases, particularly in ischemia-reperfusion injury. Quercetin 3-O-methyl ether has been reported to show potent antioxidant and neuroprotective activity against neuronal damage induced by reactive oxygen species. Several aminoethyl-substituted derivatives of quercetin 3-O-methyl ether have been synthesized to increase water solubility while retaining antioxidant and neuroprotective activity. Among such derivatives, compound 3a shows potent and well-balanced antioxidant activity in three types of cell-free assay systems and has in vivo neuroprotective effects on transient focal ischemic injury induced by the occlusion of the middle cerebral artery in rats.

Chemical constituents from aerial parts of Caryopteris incana and cytoprotective effects in human HepG2 cells.

An ethyl acetate fraction of the aerial parts of Caryopteris incana (Verbenaceae) showed potent cytoprotective effects against damage to HepG2 cells induced by tert-butylhydroperoxide (t-BHP). To search for hepatoprotective components of C. incana, various chromatographic separations of the ethyl acetate soluble fraction of C. incana led to isolation of three phenylpropanoid glycosides, 6‴-O-feruloylincanoside D, 6‴-O-sinapoylincanoside D and caryopteroside, and two iridoid glycosides, incanides A and B, together with 17 known compounds. Structures of these compounds were determined by spectroscopic analyses. The absolute stereochemistry of the caryopteroside was established with the help of circular dichroism data and in comparison with literature data. All isolated substances were determined for their cytoprotective effects against t-BHP-induced toxicity in HepG2 cells. Among the tested compounds, 6'-O-caffeoylacteoside exhibited the most potent cytoprotective activity with an IC50 value of 0.8±0.1 µM against t-BHP-induced toxicity. Structure-activity relationships of the assay results indicated an important role of the catechol moiety in phenylpropanoid, iridoid and flavonoid derivatives in eliciting cytoprotective effects.

Major role of the PI3K/Akt pathway in ischemic tolerance induced by sublethal oxygen-glucose deprivation in cortical neurons in vitro.

Ischemic preconditioning can provide protection to neurons from subsequent lethal ischemia. The molecular mechanisms of neuronal ischemic tolerance, however, are still not well-known. The present study, therefore, examined the role of MAPK and PI3K/Akt pathways in ischemic tolerance induced by preconditioning with sublethal oxygen-glucose deprivation (OGD) in cultured rat cortical neurons. Ischemic tolerance was simulated by preconditioning of the neurons with sublethal 1-h OGD imposed 12 h before lethal 3-h OGD. The time-course studies of relative phosphorylation and expression levels of ERK1/2, JNK and p38 MAPK showed lack of their involvement in ischemic tolerance. However, there were significant increases in Akt phosphorylation levels during the reperfusion period following preconditioned lethal OGD. In addition, Bcl-2 associated death promoter (Bad) and GSK-3ß were also found to be inactivated during that reperfusion period. Finally, treatment with an inhibitor of PI3K, wortmannin, applied from 15 min before and during lethal OGD abolished not only the preconditioning-induced neuroprotection but also the Akt activation. Concomitant with blockade of the Akt activation, PI3K inhibition also resulted in activation of Bad and GSK-3ß. The results suggest that ischemic tolerance induced by sublethal OGD preconditioning is primarily mediated through activation of the PI3K/Akt pathway, but not the MAPK pathway, in rat cortical neurons.

Gypenoside TN-2 ameliorates scopolamine-induced learning deficit in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynostemma pentaphyllum (Thunb.) Makino (GP, family Cucurbitaceae), which contains dammarane saponins as its main constituents, is used in China, Japan, and Korea as a traditional medicine to treat cancer, obesity, arteriosclerosis, asthma and senility. AIM OF THE STUDY: To investigate the memory-enhancing effects of GP, Gypenoside TN-2 (TN-2) was isolated by activity-guided fractionation and administered to scopolamine-induced memory-deficient mice. MATERIALS AND METHODS: The memory-enhancing effects of TN-2 were evaluated using passive avoidance, Y-maze, and Morris water maze tests, and the protein expressions of brain-derived neurotrophic factor (BDNF), cAMP element binding protein (CREB), and p-CREB were determined by immunoblotting. RESULTS: TN-2 inhibited memory and learning deficits in scopolamine treated mice in the passive avoidance test. TN-2 (10, 20, and 40 mg/kg, p.o.) significantly inhibited memory and learning deficits in the passive avoidance test by 40%, 96% and 78%, respectively, and exhibited significant memory-enhancing effects on the Y-maze test and the Morris water maze test. TN-2 also markedly increased BNDF expression and activated the transcription factor CREB in the hippocampi of scopolamine-treated mice. CONCLUSIONS: TN-2 may ameliorate memory and learning deficits by activating the CREB-BDNF pathway.

A new diarylheptanoid glycoside from the stem bark of Alnus hirsuta and protective effects of diarylheptanoid derivatives in human HepG2 cells.

To search for secondary metabolites of Alnus hirsuta (Betulaceae), various chromatographic separations of the ethyl acetate soluble fraction of the stem bark of A. hirsuta led to the isolation of a new diarylheptanoid glycoside, (3R)-1,7-bis-(4-dihydroxyphenyl)-3-heptanol 3-O-beta-D-glucopyranosyl(1-->3)-beta-D-xylopyranoside (13) and twelve diarylheptanoid derivatives, namely, oregonin (1), rubranoside A (2), hirsutanonol 5-O-beta-D-glucopyranoside (3), rubranoside B (4), rubranoside C (5), hirsutanonol (6), hirsutenone (7), (5S)-O-methylhirsutanonol (8), platyphylloside (9), platyphyllonol 5-O-beta-D-xylopyranoside (10), aceroside VII (11) and platyphyllenone (12). Isolates were assessed for their hepatoprotective effects against tert-butylhydroperoxide (t-BHP)-induced toxicity in HepG2 cells. Of these isolates, compounds 1-8 showed significant hepatoprotective effects on t-BHP-induced damage to HepG2 cells, with 8 exhibiting the greatest protective effect (50.7 + or - 3.7% at a concentration of 10 microM).

Lupane glycosides from the leaves of Acanthopanax koreanum.

Three new lupane-type saponins, acankoreosides F--H (1--3) were isolated from the methanol extract of the leaves of Acanthopanax koreanum NAKAI. The structures of these three saponins were established by chemical and spectroscopic analysis as 3alpha,30-dihydroxylup-20(29)-en-23,28-dioic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester (1), 3alpha,30-dihydroxylup-23-al-20(29)-en-28-oic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester (2), and (20S) 3alpha-hydroxylup-23-al-28,29-dioic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester (3), respectively. The effects of the isolates (1-3) on the lipopolysaccharide-induced production of nitric oxide and prostaglandin E2 were evaluated in RAW 264.7 macrophages.

Acteoside inhibits human promyelocytic HL-60 leukemia cell proliferation via inducing cell cycle arrest at G0/G1 phase and differentiation into monocyte.

We investigated the in vitro effects of acteoside on the proliferation, cell cycle regulation and differentiation of HL-60 human promyelocytic leukemia cells. Acteoside inhibited the proliferation of HL-60 cells in a concentration- and time-dependent manner with an IC50, approximately 30 microM. DNA flow cytometric analysis indicated that acteoside blocked cell cycle progression at the G1 phase in HL-60 human promyelocytic leukemia cells. Among the G1 phase cell cycle-related proteins, the levels of cyclin-dependent protein kinase (CDK)2, CDK6, cyclin D1, cyclin D2, cyclin D3 and cyclin E were reduced by acteoside, whereas the steady-state level of CDK4 was unaffected. The protein and mRNA levels of CDK inhibitors (cyclin-dependent kinase inhibitors), such as p21(CIP1/WAF1) and p27(KIP1), were gradually increased after acteoside treatment in a time-dependent manner. In addition, acteoside markedly enhanced the binding of p21(CIP1/WAF1) and p27(KIP1) to CDK4 and CDK6, resulting in the reduction of CDK2, CDK4 and CDK6 activities. Moreover, the hypophosphorylated form of retinoblastoma increased, leading to the enhanced binding of protein retinoblastoma (pRb) and E2F1. Our results further suggest that acteoside is a potent inducer of differentiation of HL-60 cells based on biochemical activities and the expression level of CD14 cell surface antigen. In conclusion, the onset of acteoside-induced G1 arrest of HL-60 cells prior to the differentiation appears to be tightly linked to up-regulation of the p21(CIP1/WAF1) and p27(KIP1) levels and decreases in the CDK2, CDK4 and CDK6 activities. These findings, for the first time, reveal the mechanism underlying the anti-proliferative effect of acteoside on human promyelocytic HL-60 cells.

Neuroprotective effects of kobophenol A against the withdrawal of tropic support, nitrosative stress, and mitochondrial damage in SH-SY5Y neuroblastoma cells.

This study examined the neuroprotective effects of kobophenol A (kob A), oligomeric stillbene, and a resveratrol tetramer. Neuronal death induced by the withdrawal of tropic support was ameliorated by kob A. The protective effect of kob A against nitrosative/oxidative or mitochondrial damages resulted in the inhibition of the ROS, intracellular calcium ion level, and mitochondrial transmembrane potential changes on SH-SY5Y cells.

Synthesis of tyrosinase inhibitory kojic acid derivative.

Kojic acid derivative 2 was synthesized by joining two pyrone rings through an ethylene linkage by Horner-Emmons reaction of phosphonate 6 with aldehyde 7. The intermediates 6 and 7 were derived from kojic acid. The tyrosinase inhibitory activity of 2 was about 8 times more potent (IC(50) = 3.63 microM) than that of kojic acid (IC(50) = 30.61 microM). Compound 2 also exhibited potent melanin synthesis inhibitory activity (19.53% inhibition at 5 mug) indicating that the connection of two pyrone rings of kojic acid through a suitable linker can be an useful strategy for identification of potent tyrosinase inhibitors.