Yangseok Kim

Effects of triterpenoids from Poria cocos Wolf on the serotonin type 3A receptor-mediated ion current in Xenopus oocytes

Poria cocos Wolf (P. cocos Wolf) is used to treat chronic gastritis, edema, nephrosis, gastric atony, acute gastroenteric catarrh, dizziness, emesis and vomiting. Triterpenoids are a class of natural compounds produced by P. cocos Wolf that contain acyclic 30-carbon precursors. In this study, we investigated the effect of triterpenoids (PA, Pachymic acid; DA, dehydroeburicoic acid; HA, 3beta-hydroxylanosta-7,9(11),24-trien-21-oic acid) on human 5-hydroxytryptamine 3A (5-HT(3A)) receptor channel activity, which is one of the ligand-gated ion channel families. The two-electrode voltage-clamp technique was used to examine the 5-HT3A mediated current. The inhibitory effect of triterpenoids on 5HT-induced inward current (I(5-HT)) occurred in a concentration dependent and reversible manner. Furthermore, the half-inhibitory concentrations (IC(50)) of PA, DA and HA were 3.2+/-0.2, 5.5+/-0.6 and 1.4+/-0.2 microM, respectively. This corresponded to an order of potency for the inhibition of I(5-HT) in oocytes expressing human 5-HT(3A) receptor of HA>PA>DA. Finally, inhibition of I(5HT) by triterpenoids occurred in a non-competitive manner, while inhibition by HA and PA showed more voltage-dependency. Taken together, these results indicate that triterpenoids may regulate the expressed 5-HT(3A) receptors in Xenopus oocytes. Furthermore, this regulation of the ligand-gated ion channel activity by triterpenoids may be one of the pharmacological actions of P. cocos Wolf.

Gene expression profile analysis of genes in rat hippocampus from antidepressant treated rats using DNA microarray

BackgroundThe molecular and biological mechanisms by which many antidepressants function are based on the monoamine depletion hypothesis. However, the entire cascade of mechanisms responsible for the therapeutic effect of antidepressants has not yet been elucidated.ResultsWe used a genome-wide microarray system containing 30,000 clones to evaluate total RNA that had been isolated from the brains of treated rats to identify the genes involved in the therapeutic mechanisms of various antidepressants, a tricyclic antidepressant (imipramine). a selective serotonin reuptake inhibitor (fluoxetine), a monoamine oxidase inhibitor (phenelzine) and psychoactive herbal extracts of Nelumbinis Semen (NS). To confirm the differential expression of the identified genes, we analyzed the amount of mRNA that was isolated from the hippocampus of rats that had been treated with antidepressants by real-time RT-PCR using primers specific for selected genes of interest. These data demonstrate that antidepressants interfere with the expression of a large array of genes involved in signaling, survival and protein metabolism, suggesting that the therapeutic effect of these antidepressants is very complex. Surprisingly, unlike other antidepressants, we found that the standardized herbal medicine, Nelumbinis Semen, is free of factors that can induce neurodegenerative diseases such as caspase 8, α-synuclein, and amyloid precursor protein. In addition, the production of the inflammatory cytokine, IFNγ, was significantly decreased in rat hippocampus in response to treatment with antidepressants, while the inhibitory cytokine, TGFβ, was significantly enhanced.ConclusionsThese results suggest that antidepressants function by regulating neurotransmission as well as suppressing immunoreactivity in the central nervous system.

Inhibition effects of Vitex rotundifolia on inflammatory gene expression in A549 human epithelial cells.

BACKGROUND Vitex rotundifolia has long been used in traditional medicine to treat asthma and other allergic diseases. OBJECTIVE To evaluate the anti-inflammatory mechanisms of V rotundifolia in cultured A549 human alveolar epithelial cells. METHODS In the present study, A549 cells were stimulated with tumor necrosis factor alpha, interleukin 4, and interleukin 1beta to induce expression of chemokines and adhesion molecules involved in eosinophil chemotaxis. The anti-inflammatory effects of V rotundifolia on stimulated A549 cells were then evaluated by analyzing eotaxin secretion and eosinophil migration. In addition, the effects of V rotundifolia on gene expression profiles in stimulated A549 cells were evaluated by oligonucleotide microarray and real-time reverse transcription-polymerase chain reaction (RTRP). RESULTS The V rotundifolia-treated A549 cells had significantly suppressed eotaxin secretion and eosinophil migration in a dose-dependent manner. In addition, the results of the microarray analysis and RTRP revealed that inflammation-related genes and cell adhesion-related genes were down-regulated in V rotundifolia-treated A549 cells. Furthermore, several genes related to the mitogen-activated protein kinase pathway were down-regulated in V rotundifolia-treated A549 cells. CONCLUSIONS The mechanism responsible for the effects of V rotundifolia on A549 cells is closely associated with regulation of the mitogen-activated protein kinase pathway. Thus, V rotundifolia may be useful in the treatment of asthma and other allergic diseases.

Screening of herbal medicines for the recovery of cisplatin-induced nephrotoxicity

The goal of this study was to quantitatively determine the recovery effects of herbal medicines (HM) on the cisplatin-induced nephrotoxicity. In the present study, the recovery effects of 239 HM on HEK 293 cells that had been damaged by cisplatin were evaluated by a mitochondrial activity MTS assay. After the first round of screening, candidate HM were selected based on a recovery rate of greater than 20%. The efficacy of the selected herbs was then determined by dose response kinetic analysis. Of the extracts evaluated, 7 HM (Paeonia suffruticosa (PS), Curcuma longa (CL), Centipeda minima (CM), Loranthus parasiticus (LP), Pulsatilla dahurica (PD), Sinapis alba (SA), and Scutellaria barbata (SB)) had a strong recovery effect on cisplatin-induced damage in HEK 293 cells. An LDH assay showed that LP, CM, SB, CL, SA, and PS had the best recovery effect, whereas a comet assay indicated that PS, SB, SA, PD, and CL had the best recovery effect. Taken together, these results suggest that SB, CL, PS, and SA are the best candidate HM for the recovery of cisplatin-induced nephrotoxicity. Therefore, additional studies should be conducted to determine if these HM possess novel therapeutic agents that can be used for the prevention or treatment of renal disorders.

Microarray analysis of gene expression profiles in response to treatment with bee venom in lipopolysaccharide activated RAW 264.7 cells.

AIM OF THE STUDY The therapeutic application of bee venom (BV) has been used in traditional medicine to treat diseases such as arthritis, rheumatism and pain. Macrophages produce molecules that are known to play roles in inflammatory responses. MATERIAL AND METHODS We performed microarray analysis to evaluate the global gene expression profiles of RAW264.7 macrophage cells treated with BV. In addition, six genes were subjected to real-time PCR to confirm the results of the microarray. The cells were treated with lipopolysaccharide (LPS) or BV plus LPS for 30 min or 1h. RESULTS 124 genes were found to be up-regulated and 158 were found to be down-regulated in cells that were treated with BV plus LPS for 30 min, whereas 211 genes were up-regulated and 129 were down-regulated in cells that were treated with BV plus LPS for 1h when compared with cells that were treated with LPS alone. Furthermore, the results of real-time PCR were similar to those of the microarray. BV inhibited the expression of specific inflammatory genes that were up-regulated by nuclear factor-kappa B in the presence of LPS, including mitogen-activated protein kinase kinase kinase 8 (MAP3K8), TNF, TNF-alpha-induced protein 3 (TNFAIP3), suppressor of cytokine signaling 3 (SOCS3), TNF receptor-associated factor 1 (TRAF1), JUN, and CREB binding protein (CBP). CONCLUSIONS These results demonstrate the potent activity of BV as a modulator of the LPS-mediated nuclear factor-kappaB (NF-kappaB)/MAPK pathway in activated macrophages. In addition, these results can be used to understand other effects of BV treatment.

Effects of protostane-type triterpenoids on the 5-HT3A receptor-mediated ion current in Xenopus oocytes.

Alisol derivatives are unique protostane-type triterpenoid compounds that are isolated from Alismatis rhizoma, which is a well-known traditional medicine in East Asia. In the present study, we investigated the effects of protostane-type triterpenoids (AA, Alisol A; AB, Alisol B; AB-ac, Alisol B 23-acetate; AC-ac, Alisol C 23-aceteate) on 5-HT-induced currents mediated by the human 5-HT(3)A receptor expressed in Xenopus laevis oocytes. Co-treatment with triterpenoids regulated the 5-HT-induced inward peak current in a concentration-dependent and reversible manner. In addition, regulation of I(5-HT) by triterpenoids occurred in a non-competitive manner. Taken together, these results indicate that triterpenoids may regulate the 5-HT(3)A receptors that are expressed in Xenopus oocytes. Furthermore, this regulation of the ligand-gated ion channel activity by triterpenoids may be one of the pharmacological actions of Alismatis rhizoma.

The α-adrenoceptor mediation of the immunomodulatory effects of electroacupuncture in DNP-KLH immunized mice

Our previous study demonstrated that successive electroacupuncture (EA) at ST36 acupoint reduces IgE production in BALB/c mice immunized with 2,4-dinitrophenylated keyhole limpet protein (DNP-KLH) by suppression of the Th2 cell lineage development. Here, we report that pretreatment of phentolamine (alpha-adrenoceptor antagonist, 10mg/kg, i.p.) completely blocks the EA-induced suppression of antigen-specific and total IgE levels in serum and IL-4 production in anti-CD3 mAb-activated splenocytes in DNP-KLH immunized mice. The results suggest that alpha-adrenoceptor play an important role in mediating the suppressive effects of EA on IgE production and Th2 cell response in DNP-KLH immunized mice.

Panax ginseng C.A. Meyer modulates the levels of MMP3 in S12 murine articular cartilage cell line.

AIM OF THE STUDY The destruction of cartilage in patients with osteoarthritis occurs due to an imbalance between matrix synthesis and degradation. Cartilage degradation is induced by the activation of matrix metalloproteinases (MMPs). Therefore, this study was conducted to evaluate the cartilage protective effect of Panax ginseng C.A. Meyer (PG). MATERIALS AND METHODS S12 cells were treated with various concentrations of extract of PG and gensenosides Rd and Rb(3) for 3h, after which 10 ng/ml interleukin-1beta (IL-1beta) was added to the culture media. The levels of MMP3 in the conditioned media were then evaluated using an enzyme-linked immunosorbent assay (ELISA). In addition, reverse transcriptase-polymerase chain reaction (RT-PCR) was used to evaluate the mRNA expression of Type II Collagen and Pro-collagenase. Furthermore, Western blot analysis was performed to identify the roles that PG played in the ERK and p38 signaling pathways. RESULTS The MMP3 secretion levels of S12 cells were significantly lowered in response to treatment with PG and gensenosides Rd and Rb(3) at a concentration of 100 microg/ml when compared to cells that were treated with IL-1beta. In addition, PG induced the mRNA expression of Type II Collagen dose dependently. Furthermore, phosphorylated p38 and ERK were detected in S12 articular cartilage cell line that was treated with IL-1beta. PG decreased the phosphorylation of p38, but PG did not exert any effect on phospho-ERK. CONCLUSIONS These findings indicate that PG and gensenosides Rd and Rb(3) suppress MMP3 secretion and that gensenosides Rd and Rb(3) are the major elements involved in the suppression of MMP3 by PG. Furthermore, the suppression of MMP3 by PG occurs via the inhibition of phospho-p38 activation. Therefore, PG may exert a protective effect against the cartilage degradation of OA.

Screening of herbal medicines for recovery of acetaminophen-induced nephrotoxicity.

This study was conducted to quantitatively evaluate the recovery effects of herbal medicines on acetaminophen-induced nephrotoxicity. In the present study, the recovery effects of 251 herb medicines on HEK 293 cells that had been damaged by acetaminophen were evaluated using an MTS assay. HEK 293 cells were cultured in 96-well plates and then pretreated with or without 20μM acetaminophen (IC(50) value: 17.5±1.9) for 1h. Next, different herbal medicines were added to the wells, after which the cells were reincubated at 37°C for 24h. After the first round of screening, the candidate herbal medicines were selected based on a recovery rate of greater than 20% and their efficacy were then determined by dose response kinetic analysis. Among these extracts, 8 herbal medicines (Ledebouriella divaricata, Sparganium simplex, Panax ginseng, Aster tataricus, Citrus aurantium, Sanguisorba officianlis, Arisaema consanguineum, and Polygonum aviculare) had a strong recovery effect on acetaminophen-induced damage in HEK 293 cells. Dose response non-linear regression analysis demonstrated that P. aviculare showed the best recovery rate (98%), and that its EC(50) (0.1ng/mL) was the smallest among the screened candidate herbal medicines. Additional studies of these herbal medicines should be conducted to determine if they possess novel therapeutic agents for the prevention or treatment of renal disorders.

Inhibitory effects of Schizandrae Fructus on eotaxin secretion in A549 human epithelial cells and eosinophil migration

Eosinophilia have been implicated in a broad range of diseases, most notably allergic conditions (e.g. asthma, rhinitis and atopic dermatitis) and inflammatory diseases. These diseases are characterized by an accumulation of eosinophils in the affected tissue. Defining the mechanisms that control the recruitment of eosinophil is fundamental to understanding how these diseases progress and identifying a novel target for drug therapy. Accordingly, this study was conducted to evaluate the regulatory effect of Schizandrae Fructus (SF) on the expression of eotaxin, an eosinophil-specific chemokine released in respiratory epithelium following allergic stimulation, as well as its effects on eosinophil migration. To accomplish this, human epithelial lung cells (A549 cell) were stimulated with a combination of TNF-alpha (100ng/ml) and IL-4 (100ng/ml) for 24h. The cells were then restimulated with TNF-alpha (100ng/ml) and IL-1beta (10ng/ml) to induce the expression of chemokines and adhesion molecules involved in eosinophil chemotaxis for another 24h. Next, the samples were treated with various concentrations of Schizandrae Fructus (SF) (1, 10, 100, 1000microg/ml) or one of the major constituents of SF, schizandrin (0.1, 1, 10, 100microg/ml), after which following inhibition effect assay was performed triplicates in three independence. The levels of eotaxin in secreted proteins were suppressed significantly by SF (100 and 1000microg/ml, p<0.01) and schizandrin (10 and 100microg/ml, p<0.01). In addition, SF (1, 10, 100 and 1000microg/ml) decreased mRNA expression levels in A549 cells significantly (p<0.01). Eosinophil recruitment to lung epithelial cells was also reduced by SF, which indicates that eotaxin plays a role in eosinophil recruitment. Furthermore, treatment with SF suppressed the expression of another chemokine, IL-8 (0.1 and 1microg/ml SF, p<0.01), as well as intercellular adhesion molecule-1 (10 and 100microg/ml SF, p<0.01) and vascular cell adhesion molecule-1 (0.1 and 1microg/ml SF, p<0.05), which are all related to eosinophil migration. Taken together, these findings indicate that SF may be a desirable medicinal plant for the treatment of allergic diseases.