Inhibitory effect of rhynchophylline on human ether-a-go-go related gene channel.

We studied the effects of Chinese traditional medicine rhynchophylline (Rhy) on human ether-a-go-go related gene (HERG) channel and characterized the electrophysiological properties of Rhy's pharmacological effect on HERG channel using Xenopus oocytes. Xenopus oocytes were injected with either 23 nl (5.75 ng) HERG cRNA or 23 nl distilled water. Xenopus oocytes were randomly assigned to receive one of the following different concentrations of Rhy: (1) control, (2)10 mumol/L Rhy, (3)100 mumol/L Rhy, (4) 500 mumol/L Rhy, (5) 1 000 mumol/L Rhy, (6) 10 000 mumol/L Rhy. Cell currents were recorded in oocytes. The peak tail currents of HERG channel were inhibited by Rhy. The inhibition was in a dose-dependent manner [IC(50)=(773.4 +/- 42.5) mumol/L]. Experiment with 100 mumol/L Rhy indicated that the degree of HERG blockade showed some voltage dependence (within -40 mV to -20 mV ). Kinetic analyses revealed that Rhy decreased the rate of channel activation. The findings indicate that Rhy inhibits HERG encoded potassium channels. It may underline the molecular mechanism of myocardial electrophysiological characteristics associated with this drug.

[Relationship between congenital long QT syndrome and Brugada syndrome gene mutation].

OBJECTIVE: To investigate the molecular pathology in families with long QT syndrome (LQTS) including Jervell-Longe-Nielsen syndrome (JLNS) and Romano-ward syndrome (RWS) and Brugada syndrome (BS) in Chinese population. METHODS: Polymerase chain reaction and DNA sequencing were used to screen for KCNQ1, KCNH2, KCNE1, and SCN5A mutation. RESULTS: We identified a novel mutation N1774S in the SCN5A gene of the BS family, a novel mutation G314S in a RWS family which had also been found in Europe, North America, and Japan, and a single nucleotide polymorphisms (SNPs) G643S in the KCNQ1 of the JLNS family. In this JLNS family, another heterozygous novel mutation in exon 2a was found in KCNQ1 of the patients. CONCLUSION: New mutations were found in our experiment, which expand the spectrum of KCNQ1 and SCN5A mutations that cause LQTS and BS.

[Mutation analysis of a Chinese family with inherited long QT syndrome].

OBJECTIVE: To identify the mutation of a Chinese family with inherited long QT syndrome(LQTS). METHODS: The disease-causing gene was tentatively determined in light of the clinical manifestations and electrophysiological properties, and then polymerase chain reaction and DNA sequencing were used for screening and identifying mutation. RESULTS: A missense mutation G940A(G314S) in the KCNQ1 gene was identified, which was the 'hot spot' of long QT syndrome mutation. CONCLUSION: The mutation that is involved with long QT syndrome in Chinese patients is the same as that in the European, American and Japanese patients.

[PCR site-directed mutagenesis of long QT syndrome KCNQ1 gene in vitro].

To study PCR site-directed mutagenesis of long QT syndrome KCNQ1 gene in vitro. The site-directed mutagenesis of LQTS gene KCNQ1 was made by PCR. Two sets of primers were designed according to the sequence of KCNQ1 cDNA, and mismatch was introduced into primers. Mutagenesis was performed in a three-step PCR. The amplified fragments from the third PCR which contained the mutation site were subcloned into the T-vecor PCR2.1. Then the fragments containing the mutation site was obtained from PCR2.1 with restriction enzyme digestion and was inserted into the same restriction site of pIRES2-EGFP-KCNQ1. With Effectene Transfection Reagent, pIRES(2)-EGFP-KCNQ1 was transfected into HEK293 cell. The sequencing analysis showed that the mutation site was correct. Mutation from T to C in 934 site of KCNQ1 cDNA was found. Under the fluorescence microscope, the green fluorescence was spread in the transfected HEK293 cell, meaning the pIRES(2)-EGFP-KCNQ1 containing the mutation site was expressed correctly.