The vasorelaxation effect of an extract of Chinese medicinal herb, Radix Angelica Pubescentis, in porcine coronary artery Institute: The University of Hong Kong Supervisor: Prof. Ricky Y. K. Man Prof. C. M. Che Presenter: Matthew L. Y. Chan 2006 World Congress on Chinese Medicine 獨活提取物對豬冠狀動脈血管舒張的影響

Pharmaceutical name: Radix Angelicae Pubescentis Latin botanical name: Angelica pubescens Maxim. f. biserrata Shan et Yuan Chinese (English) name: 獨活 (Du Huo) Introduction Chinese Medicinal Herb

anti-arthritis headache thrombosis vessels dilation promotion blood circulation Pharmacological Actions: Introduction Chinese Pharmacopoeia anti-inflammatory * # anti-cancer * # vaso-relaxant # anti-oxidant # Recent Publications extract ingredients * #

Major active ingredients  anti-arrhythmic  vaso-relaxant Osthole (0.5%)  antioxidant Xanthotoxin  anti-cancer Angelmarin Introduction

Dry AP samples Grind Extract trice with 95% ethanol (1:5, w/v) (stir overnight, centrifuge and evaporate to dryness) Store at -20˚C and dissolve in DMSO before use Powdered herb Crude extract 20 mg/ml APE Material Preparation of APE and osthole Store at -20˚C and dissolve in DMSO before use Osthole powder 10 mM Osthole

Organ bath setup Amplifier Force transducer Circulator Organ bath Method

Preparation of the porcine coronary arterial rings Water in Water out 95% O 2 5% CO 2 3 mm coronary arterial ring Measurement of isometric tension change Krebs-Henseleit solution (KHS) (NaCl 120 mM, KCl 4.76 mM, NaHCO 3 25 mM, NaH 2 PO 4 ·2H 2 O, 1.18 mM CaCl mM, MgSO 4 ·7H 2 O 1.18 mM, D-glucose 5.5 mM)

Method U46619 (30 nM) Wash with Krebs Indo (10 μM) Viability test > 80 % relaxation Cumulative addition of drug BK (1 μM) Indo and/or blocker > 4 g contraction 2 g resting tension U46619 (30 nM) Time Tension Schematic diagram of organ bath technique

Results & Discussion Part I Response to APE on porcine coronary arterial rings A Endothelium-intact B Endothelium-disrupted C D

GC GTP cGMP PKG + Vascular smooth muscle cell Relaxation Endothelium NO eNOS L-arginine Conclusion Part I Schematic diagram depicts the proposed mechanisms of APE L-NAME - Endothelium-intact AP E 

Vascular smooth muscle cell GC GTP cGMP PKG + AC ATP cAMP PKA + Relaxation AP E KT5823 Rp-8-Br-cGMPS ODQ KT5720 Rp-8-Br-cAMPS SQ       Schematic diagram depicts the proposed mechanisms of APE Endothelium-disrupted Conclusion Part I

Results & Discussion Part II Response to osthole on porcine coronary arterial rings ∗ p < 0.05 versus L-NAME group Endothelium-intact * F G * ∗ p < 0.05 versus ODQ group ∗ p < 0.05 versus without EC group E *

I Endothelium-disrupted H Results & Discussion Part II Response to osthole on porcine coronary arterial rings ∗ p < 0.05 versus without EC group E *

GC GTP cGMP PKG + Vascular smooth muscle cell Relaxation Endothelium NO eNOS L-arginine Conclusion Part II Schematic diagram depicts the proposed mechanisms of osthole Endothelium-intact Osthol e (10 μM) L-NAME - ODQ -

Vascular smooth muscle cell GC GTP cGMP PKG + AC ATP cAMP PKA + Relaxation Conclusion Part II Schematic diagram depicts the proposed mechanisms of osthole Endothelium-disrupted ODQ - SQ Osthol e  

Future Plan  Continue to study the mechanism of action APE work on, such as K and Ca channel.  Continue to study the mechanism of action osthole work on, such as those downstream pathway of cGMP cascade.

Calculated concentration of osthole in APE Actual concentration of osthole Comparison of the osthole concentration used in the experiment