관상동맥 평활근 세포의 이온 전류에 미치는 lysophosphatidylcholine의 효과

Abstract

[한글]

심근조직에 공급되는 혈류량의 크기에 커다란 영향을 끼치고 있는 관상동맥의 긴장도는 평활근 세포막 전압의 분극정도에 커다란 영향을 받고 있으며, 평활근 세포막의 막전압 의존성 K**+ 전류의 활성도의 변화가 이같은 막전압의 크기 결정에 매우 중요한 역할을 하고 있음은 잘 알려져있다. 즉 막전압 의존성 K**+ 전류의 활성이 어떤 원인에 의해서든지 감소하게 되면, 이를 통한 K**+ 이온의 유출이 감소함으로써 세포막 전압이 안정상태에 비해 탈분극되고 이에 따라 평활근 세포막의 Ca**2+ 통로를 통한 Ca**2+ 유입이 증가함으로써 혈관 수축이 유발되게 된다. 죽상경화증 시 관찰되는 관상동맥의 비정상적 수축 현상 역시 죽상경화증 혈관에서 관찰되는 막전압의 비정상적인 탈분극 효과에 의해 나타날 가능성이 있으나 이 같은 막전압의 탈분극을 유발하는 인자나 유발 기전에 대해서는 아직 명확치 않은 실정이다. 따라서 본 실험에서는 토끼 관상동맥 평활근 세포를 이용한 whole patch clamp 방법을 사용하여 죽상경화증 혈관에서 그 농도가 크게 증가한다고 알려진 lysophosphatidylcholine(LPC)이 막전압 의존성 K**+ 전류의 활성에 미치는 효과와 그 작용 기전을 규명함으로써 죽상경화증 혈관에서 관찰되는 비정상적인 관상동맥 수축 현상의 기전을 규명하고자 하였으며, 이들 실험을 통해 얻은 결과는 다음과 같다.

1. 평활근 세포막 전압의 탈분극 정도에 따라 LPC는 외향성 막전류의 크기를 감소 또는 증가시켜 주었으며, Gd**3+ 투여 시 LPC에 의한 막전류 증가효과는 소실되었다.

2. 평활근 세포의 막전압을 탈분극된 상태로 장기간 유지하는 경우 LPC에 의한 막전류 감소 효과는 소실되었다.

3. 막전압 의존성 K**+ 전류의 활성은 첨가한 LPC 농도에 비례하여 감소하였으며, 이때 막전압 의존성 활성 곡선 및 비활성 곡선이 LPC에 의해 오른쪽으로 이동하였다.

4. PKC 차단제인 staurosporine 전처치시 LPC에 의한 막전압 의존성 K**+ 전류 억제 효과가 감소되었다

5. 세포 내액에 직접 첨가한 저농도(10nM) LPC에 의해 막전압 의존성 K**+ 전류가 억제되는 것을 관찰하였다.

이상의 실험 결과로 볼 때 LPC에 의한 막전압 의존성 K**+ 전류의 억제효과는 평활근 세포막 전압의 탈분극 및 이에 따른 세포 외로부터의 Ca**2+유입의 증가를 초래함으로써 죽상경화증 혈관에서 관찰되는 비정상적 혈관수축에 중요한 역할을 한다고 생각되며, 이

같은 막전압 의존성 K**+ 전류 억제효과에는 LPC에 의한 PKC 활성 증가가 중요한 역할을 한다고 생각된다.

Effect of lysophosphatidylcholine on whole cell K**+ current in rabbit coronary

smooth muscle cells

Chang-Heon Yi

Department of Medicine The Graduate School, Yonsei University

(Directed by Associate Professor Duck-Sun Ahn)

Impairment of relaxing response and augmentation of contractile response to

vasoactive substances have been reported in atherosclerotic arteries. These

alterations in vascular reactivity are considered as an underlying mechanism for

the development of acute vasospasm in atherosclerotic coronary artery. Recently, it

has been reported that lysophosphatidylcholine (LPC), an oxidative metabolite of

low density lipoprotein causes this functional abnormality. However, the precise

mechanism of LPC induced change of vascular reactivity is still uncertain. In this

study, to elucidate the underlying mechanisms of abnormal vascular reactivity in

atherosclerotic coronary artery, we directly examined the effect of LPC on whole

cell K**+ current using patch clamp technique in rabbit coronary smooth muscle

cells.

Application of LPC(1μM) showed dual effect on whole cell outward current which

depends on the magnitude of test potentials. At relatively high depolarizing test

potentials(>10mV), LPC increased amplitude of outward current which was blocked by

Gd**3+ , not by iberiotoxin(100nM) and TEA(1mM). Reversal potential of this Gd**3+

-sensitive, LPC-induced current was -9.7±0.6mV. At less depolarizing test

potentials(<10mV), LPC decreased whole cell K**+ currents in a dose dependent

manner(from 0.01 to 10 μM) in the range of -3OmV to +10mV. Half maximal inhibition

of K**+ current was 1.509μM at OmV test potential(n=5). Depolarizing holding

potential (OmV) prevented this LPC-induced inhibition of K**+ current. Steady state

activation and inactivation parameters of K**+ current were significantly shifted

to the positive direction by application of LPC(p<0.01, n=8). Pretreatment of

staurosporine(100nM), a blocker of protein kinase C partially blocked LPC-induced

decrease of K**+ currents. These LPC-induced changes of K**+ current may explain

earlier observations of abnormal vascular reactivity in atherosclerotic coronary

artery.

[영문]

Impairment of relaxing response and augmentation of contractile response to vasoactive substances have been reported in atherosclerotic arteries. These alterations in vascular reactivity are considered as an underlying mechanism for the development of acute vasospasm in atherosclerotic coronary artery. Recently, it has been reported that lysophosphatidylcholine (LPC), an oxidative metabolite of low density lipoprotein causes this functional abnormality. However, the precise mechanism of LPC induced change of vascular reactivity is still uncertain. In this study, to elucidate the underlying mechanisms of abnormal vascular reactivity in atherosclerotic coronary artery, we directly examined the effect of LPC on whole cell K**+ current using patch clamp technique in rabbit coronary smooth muscle cells.

Application of LPC(1μM) showed dual effect on whole cell outward current which depends on the magnitude of test potentials. At relatively high depolarizing test potentials(>10mV), LPC increased amplitude of outward current which was blocked by Gd**3+ , not by iberiotoxin(100nM) and TEA(1mM). Reversal potential of this Gd**3+ -sensitive, LPC-induced current was -9.7±0.6mV. At less depolarizing test

potentials(<10mV), LPC decreased whole cell K**+ currents in a dose dependent manner(from 0.01 to 10 μM) in the range of -3OmV to +10mV. Half maximal inhibition of K**+ current was 1.509μM at OmV test potential(n=5). Depolarizing holding potential (OmV) prevented this LPC-induced inhibition of K**+ current. Steady state activation and inactivation parameters of K**+ current were significantly shifted to the positive direction by application of LPC(p<0.01, n=8). Pretreatment of staurosporine(100nM), a blocker of protein kinase C partially blocked LPC-induced decrease of K**+ currents. These LPC-induced changes of K**+ current may explain earlier observations of abnormal vascular reactivity in atherosclerotic coronary artery.