Pornpen Pramyothin

Efficacy of Thai medicinal plant extracts against herpes simplex virus type 1 infection in vitro and in vivo

Twenty Thai medicinal plant extracts were evaluated for anti-herpes simplex virus type 1 (HSV-1) activity. Eleven of them inhibited plaque formation of HSV-1 more than 50% at 100microg/ml in a plaque reduction assay. Aglaia odorata, Moringa oleifera, and Ventilago denticulata among the 11 were also effective against thymidine kinase-deficient HSV-1 and phosphonoacetate-resistant HSV-1 strains. These therapeutic efficacies were characterized using a cutaneous HSV-1 infection in mice. The extract of M. oleifera at a dose of 750mg/kg per day significantly delayed the development of skin lesions, prolonged the mean survival times and reduced the mortality of HSV-1 infected mice as compared with 2% DMSO in distilled water (P<0.05). The extracts of A. odorata and V. denticulata were also significantly effective in limiting the development of skin lesions (P<0.05). There were no significant difference between acyclovir and these three plant extracts in the delay of the development of skin lesions and no significant difference between acyclovir and M. oleifera in mean survival times. Toxicity of these plant extracts were not observed in treated mice. Thus, these three plant extracts may be possible candidates of anti-HSV-1 agents.

Anti-herpes simplex virus (HSV-1) activity of oxyresveratrol derived from Thai medicinal plant : Mechanism of action and therapeutic efficacy on cutaneous HSV-1 infection in mice

Oxyresveratrol, a major compound purified from Artocarpus lakoocha, a Thai traditional medicinal plant, was evaluated for its mechanism of action and therapeutic efficacy on cutaneous herpes simplex virus (HSV) infection in mice. The inhibitory concentrations for 50% HSV-1 plaque formation of oxyresveratrol, three clinical isolates, thymidine kinase (TK)-deficient and phosphonoacetic acid (PAA)-resistant HSV-1 were 19.8, 23.3, 23.5, 24.8, 25.5 and 21.7microg/ml, respectively. Oxyresveratrol exhibited the inhibitory activity at the early and late phase of viral replication and inhibited the viral replication with pretreatment in one-step growth assay of HSV-1 and HSV-2. Oxyresveratrol inhibited late protein synthesis at 30microg/ml. The combination of oxyresveratrol and acyclovir (ACV) produced synergistic anti-HSV-1 effect, as characterized by the isobologram of plaque inhibition. Mice orally treated with oxyresveratrol (500mg/kg/dose) dose at 8 h before and three times daily had significant delay in herpetic skin lesion development (P<0.05). Topical application of 30% oxyresveratrol ointment five times daily significantly delayed the development of skin lesions and protected mice from death (P<0.0001).

Protective activity of phyllanthin in ethanol-treated primary culture of rat hepatocytes

AIM OF THE STUDY To investigate the protective effect of phyllanthin (a known principal constituent of Phyllanthus amarus Schum. et Thonn.) on ethanol-induced rat liver cell injury. MATERIALS AND METHODS Primary culture of rat hepatocytes (24h culturing) were pretreated with phyllanthin (1, 2, 3 and 4 microg/ml) for 24h. After 24h pretreatment, cells were treated with ethanol (80 microl/ml) for 2h. RESULTS Ethanol decreased %MTT, increased the release of transaminases (ALT and AST) with the increase in the production of intracellular ROS and lipid peroxidation. Phyllanthin demonstrated its role in protection by antagonizing the above effect induced by ethanol. Phyllanthin also restored the antioxidant capability of rat hepatocytes including level of total glutathione, and activities of superoxide dismutase (SOD) and glutathione reductase (GR) which were reduced by ethanol. CONCLUSIONS These results suggested the hepatoprotective effect of phyllanthin against ethanol-induced oxidative stress causing rat liver cell damage through its antioxidant activity.

Inhibitory activity of oxyresveratrol on wild-type and drug-resistant varicella-zoster virus replication in vitro

The anti-herpes simplex virus (HSV) compound, oxyresveratrol, purified from a Thai traditional medicinal plant of Artocarpus lakoocha, was evaluated for its anti-varicella-zoster virus (VZV) activity. This compound exhibited IC(50) values (50%-inhibitory concentrations for virus plaque formation) of 12.82, 12.80, 12.99 and 12.82 microg/ml against wild type, thymidine kinase-deficient and two types of DNA polymerase mutants with acyclovir-resistance, respectively. Thus oxyresveratrol showed a broad spectrum of anti-VZV activity with a mechanism of action different from that of acyclovir.

Topical cream-based oxyresveratrol in the treatment of cutaneous HSV-1 infection in mice

Anti-herpes simplex virus (HSV) activities of oxyresveratrol in vitro and topical administration in cutaneous HSV-1 infection in mice were examined. The inhibitory concentrations for 50% plaque formation (IC(50)) of oxyresveratrol against HSV-1 clinical isolates and HSV-2 clinical isolates were 20.9-29.5 and 22.2-27.5 μg/ml, respectively. In topical administration in cutaneous HSV-1 infection in mice, 2.5%, 5%, 10% and 20% oxyresveratrol in cream vehicle applied three times daily for 7 days after infection were evaluated and 10% and 20% oxyresveratrol cream were significantly effective in delaying the development of skin lesions and protection from death (P < 0.01). The concentration of 10% oxyresveratrol in cream was significantly more effective than that of 30% oxyresveratrol in vaseline applied three times daily (P < 0.01). Oxyresveratrol cream at 20% was as effective as 5% ACV cream applied three times daily (P < 0.01). Both 10% and 20% oxyresveratrol cream were as effective as that of 5% ACV cream applied two times daily (P > 0.05). Therapeutic efficacy of oxyresveratrol in cream vehicle was dose-dependent and the maximum efficacy observed on day 6 after infection was shown at 10% oxyresveratrol in cream applied three times daily. The frequency of application of 10% oxyresveratrol cream at three, four and five times daily was as effective as that of 5% ACV cream applied five times daily (P > 0.05). These results demonstrated that topical administration of oxyresveratrol in novel cream vehicle reduced the concentration of oxyresveratrol to 10% and was suitable for cutaneous HSV infection.

Endothelium-independent effects of phyllanthin and hypophyllanthin on vascular tension

The purpose of this study was to investigate the modulating effects of phyllanthin and hypophyllanthin on vascular tension, using in the in vitro model of isolated rat aorta. Our results indicated that both phyllanthin and hypophyllanthin significantly relaxed the sustained contraction induced by phenylephrine (PE) in a concentration-dependent manner. In addition, endothelial removal had no significant influence on the vasorelaxation responses of the aortic rings toward these two compounds. Furthermore, both compounds inhibited the contraction of aortic muscle provoked by either PE (1 μM) or KCl (40 mM) as well as the spontaneous contraction of the Ca²⁺-depleted muscle. In high K⁺-Ca²⁺ free solution, phyllanthin (100 μM), but not hypophyllanthin, significantly inhibited the contractile responses upon cumulative addition of CaCl₂. Both compounds (100 μM) significantly inhibited PE-induced contraction in Ca²⁺-free condition, but could not affect caffeine-induced contraction. Taken together, phyllanthin and hypophyllanthin could modulate the vascular tension via the endothelium-independent mechanisms. The modulating effects of both compounds were possibly involved with the blockade of Ca²⁺ entry into vascular smooth muscle cells and inhibition of PE-mediated Ca²⁺ release from sarcoplasmic reticulum.

Combination of multiplex PCR and DHPLC-based strategy for CYP2D6 genotyping scheme in Thais

OBJECTIVE To develop CYP2D6 genotyping scheme for accurate allele calling and reliable estimation of functional allele dosage in Thais. DESIGN AND METHODS We analyzed CYP2D6 copy numbers by pentaplex PCR coupled with semi-quantitative denaturing high performance liquid chromatography (DHPLC)-based technique. Ten common SNPs were genotyped from CYP2D6 gene product using single base extension (SBE) followed by DHPLC analysis. This detection scheme was compared with real-time PCR and conventional PCR-RFLP for cost-effectiveness. RESULTS The distribution of CYP2D6 gene copy numbers in our population ranged from zero (0.69%), one (7.99%), two (60.07%), three (28.13%) and four (3.13%). The most commonly detected SNPs were related to CYP2D6*10 haplotype. CYP2D6*36 in tandem with CYP2D6*10B is the major rearrangement type in Thais (18.75%). CONCLUSIONS Multiplex PCR coupled with DHPLC-based strategy is convenient and reliable method for CYP2D6 genotyping offering sufficient allele coverage for Asians. Both cost and analytical time saving were shown and the method could potentially be modified to accommodate CYP2D6 genotyping in other ethnics.

Phyllanthin and hypophyllanthin inhibit function of P‐gp but not MRP2 in Caco‐2 cells

The purposes of this study were to investigate the inhibitory effects of two lignans, phyllanthin and hypophyllanthin, on the function of P‐glycoprotein (P‐gp) and multidrug resistance protein 2 (MRP2), using the in‐vitro model of Caco‐2 cells. In addition, the effect of prolonged exposure to these two compounds on the expression of active P‐gp was also determined.